Evaluating the role of low-speed centrifugation towards transfecting human peripheral blood mononuclear cell culture

Directory of Open Access Journals (Sweden)

M Majumdar

2014-01-01

Full Text Available The conventional method of transfection of suspension cells by chemical has proven to be very difficult. We present a new transfection protocol, wherein, low-speed centrifugation of cell culture plates immediately after adding the lipid: DNA complex significantly enhances the transfection efficiency. Peripheral blood mononuclear cells (PBMCs were transfected with BLOCK-iT™ Fluorescent Oligo (scrambled siRNA and lipofectamine complex using conventional and low-speed centrifugation modified transfection protocols. The efficiency of transfection was determined using flowcytometer and cell viability was checked using MTT assay. Incorporation of low-speed centrifugation significantly enhances the transfection efficiency of BLOCK-iT™ in the suspension culture of PBMCs as compared to conventional transfection method (99.8% vs 28.3%; P < 0.0001, even at a low concentration of 40 picomoles without affecting the cell viability. Centrifugation enhanced transfection (CET technique is simple, time-saving and novel application without compromising the cell viability in the context of recently popular RNA interference in suspension cultures of PBMCs. This undemanding modification might be applicable to a wide variety of cell lines and solve crucial problem of researchers working with RNA interference in suspension cultures.

Comparison among four proposed direct blood culture microbial identification methods using MALDI-TOF MS.

Science.gov (United States)

Bazzi, Ali M; Rabaan, Ali A; El Edaily, Zeyad; John, Susan; Fawarah, Mahmoud M; Al-Tawfiq, Jaffar A

Matrix-assisted laser desorption-ionization time-of-flight (MALDI-TOF) mass spectrometry facilitates rapid and accurate identification of pathogens, which is critical for sepsis patients. In this study, we assessed the accuracy in identification of both Gram-negative and Gram-positive bacteria, except for Streptococcus viridans, using four rapid blood culture methods with Vitek MALDI-TOF-MS. We compared our proposed lysis centrifugation followed by washing and 30% acetic acid treatment method (method 2) with two other lysis centrifugation methods (washing and 30% formic acid treatment (method 1); 100% ethanol treatment (method 3)), and picking colonies from 90 to 180min subculture plates (method 4). Methods 1 and 2 identified all organisms down to species level with 100% accuracy, except for Streptococcus viridans, Streptococcus pyogenes, Enterobacter cloacae and Proteus vulgaris. The latter two were identified to genus level with 100% accuracy. Each method exhibited excellent accuracy and precision in terms of identification to genus level with certain limitations. Copyright © 2016 King Saud Bin Abdulaziz University for Health Sciences. Published by Elsevier Ltd. All rights reserved.

Proliferative and phenotypical characteristics of human adipose tissue-derived stem cells: comparison of Ficoll gradient centrifugation and red blood cell lysis buffer treatment purification methods.

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Najar, Mehdi; Rodrigues, Robim M; Buyl, Karolien; Branson, Steven; Vanhaecke, Tamara; Lagneaux, Laurence; Rogiers, Vera; De Kock, Joery

2014-09-01

Adult human subcutaneous adipose tissue harbors a multipotent stem cell population, the so-called human adipose tissue-derived mesenchymal stromal cells (AT-MSCs). These cells are able to differentiate in vitro into various cell types and possess immunomodulatory features. Yet procedures to obtain AT-MSCs can vary significantly. The two most extensively used AT-MSC purification techniques are (i) density gradient centrifugation using Ficoll and (ii) red blood cell (RBC) lysis buffer treatment of the stromal vascular fraction. In the context of potential clinical cell therapy, the stem cell yield after purification and upon consecutive passages, as well as the purity of the obtained cell population, are of utmost importance. We investigated the expansion capacity and purity of AT-MSCs purified by both procedures immediately after isolation and upon consecutive passages. We also investigated possible purification-dependent differences in their expression of immune-inhibitory factors and cell adhesion molecules. We found that RBC lysis buffer treatment is a more robust and easier method to purify AT-MSCs than density gradient fractionation. However, the resulting AT-MSC-RBC population contains a significantly higher number of CD34(+) cells, particularly during the first passages after plating. From passage 4 onward, no significant differences could be observed between both populations with respect to the immunophenotype, expansion capacity and expression of immune inhibitory factors and cell adhesion molecules. Our data show that RBC lysis buffer treatment may be a good alternative to density fractionation, providing a faster, more robust and easier method to purify AT-MSCs with biologically preserved characteristics. Copyright © 2014 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Direct identification of microorganisms from positive blood cultures using the lysis-filtration technique and matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS): a multicentre study.

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Farina, Claudio; Arena, Fabio; Casprini, Patrizia; Cichero, Paola; Clementi, Massimo; Cosentino, Marina; Degl'Innocenti, Roberto; Giani, Tommaso; Luzzaro, Francesco; Mattei, Romano; Mauri, Carola; Nardone, Maria; Rossolini, Gian Maria; Serna Ortega, Paula Andrea; Vailati, Francesca

2015-04-01

Microbial identification from blood cultures is essential to institute optimal antibiotic therapy and improve survival possibilities. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has been successfully applied to identify bacteria and yeasts from positive blood cultures broths. The aim of this multicentre study was to evaluate the reliability of the lysis-filtration technique associated with MALDI-TOF MS to directly identify microorganisms from 765 positive blood cultures collected in six Italian hospitals. Overall, 675/765 (78.1%) blood isolates were correctly identified at the species level, with significant differences between Gram-negative and Gram-positive bacteria (92.6%, and 69.8%, respectively). Some difficulties arise in identifying Streptococcus pneumoniae, Staphylococcus aureus, yeasts and anaerobes. The lysis-filtration protocol is a suitable procedure in terms of performance in identifying microorganisms, but it is quite expensive and technically time-consuming since the time of filtration is not regular for all the samples. The application of the MALDI-TOF MS technique to the direct microbial identification from positive blood cultures is a very promising approach, even if more experience must be gained to minimize errors and costs.

A simple method for rapid microbial identification from positive monomicrobial blood culture bottles through matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

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Lin, Jung-Fu; Ge, Mao-Cheng; Liu, Tsui-Ping; Chang, Shih-Cheng; Lu, Jang-Jih

2017-06-30

Rapid identification of microbes in the bloodstream is crucial in managing septicemia because of its high disease severity, and direct identification from positive blood culture bottles through matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) can shorten the turnaround time. Therefore, we developed a simple method for rapid microbiological identification from positive blood cultures by using MALDI-TOF MS. We modified previously developed methods to propose a faster, simpler and more economical method, which includes centrifugation and hemolysis. Specifically, our method comprises two-stage centrifugation with gravitational acceleration (g) at 600g and 3000g, followed by the addition of a lysis buffer and another 3000g centrifugation. In total, 324 monomicrobial bacterial cultures were identified. The success rate of species identification was 81.8%, which is comparable with other complex methods. The identification success rate was the highest for Gram-negative aerobes (85%), followed by Gram-positive aerobes (78.2%) and anaerobes (67%). The proposed method requires less than 10 min, costs less than US$0.2 per usage, and facilitates batch processing. We conclude that this method is feasible for clinical use in microbiology laboratories, and can serve as a reference for treatments or further complementary diagnostic testing. Copyright © 2017. Published by Elsevier B.V.

Same day identification and full panel antimicrobial susceptibility testing of bacteria from positive blood culture bottles made possible by a combined lysis-filtration method with MALDI-TOF VITEK mass spectrometry and the VITEK2 system.

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Machen, Alexandra; Drake, Tim; Wang, Yun F Wayne

2014-01-01

Rapid identification and antimicrobial susceptibility testing of microorganisms causing bloodstream infections or sepsis have the potential to improve patient care. This proof-of-principle study evaluates the Lysis-Filtration Method for identification as well as antimicrobial susceptibility testing of bacteria directly from positive blood culture bottles in a clinical setting. A total of 100 non-duplicated positive blood cultures were tested and 1012 microorganism-antimicrobial combinations were assessed. An aliquot of non-charcoal blood culture broth was incubated with lysis buffer briefly before being filtered and washed. Microorganisms recovered from the filter membrane were first identified by using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight VITEK® Mass Spectrometry (VITEK MS). After quick identification from VITEK MS, filtered microorganisms were inoculated to VITEK®2 system for full panel antimicrobial susceptibility testing analysis. Of 100 bottles tested, the VITEK MS resulted in 94.0% correct organism identification to the species level. Compared to the conventional antimicrobial susceptibility testing methods, direct antimicrobial susceptibility testing from VITEK®2 resulted in 93.5% (946/1012) category agreement of antimicrobials tested, with 3.6% (36/1012) minor error, 1.7% (7/1012) major error, and 1.3% (13/1012) very major error of antimicrobials. The average time to identification and antimicrobial susceptibility testing was 11.4 hours by using the Lysis-Filtration method for both VITEK MS and VITEK®2 compared to 56.3 hours by using conventional methods (pdirectly from positive blood culture can be achieved and can be used for appropriate antibiotic therapy and antibiotic stewardship.

Centrifugal blood pump 603

Indian Academy of Sciences (India)

Centrifugal blood pump 603 pressure obtained for real blood, as shown in figure 6, is a little higher than that for glycerin aqua Solution with the same viscosity as blood. This may indicate the effect of slight non-. Newtonian turbulent flow. The radial whirl motion of the impeller was observed by dual laser position sensors.

Same day identification and full panel antimicrobial susceptibility testing of bacteria from positive blood culture bottles made possible by a combined lysis-filtration method with MALDI-TOF VITEK mass spectrometry and the VITEK2 system.

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Alexandra Machen

Full Text Available Rapid identification and antimicrobial susceptibility testing of microorganisms causing bloodstream infections or sepsis have the potential to improve patient care. This proof-of-principle study evaluates the Lysis-Filtration Method for identification as well as antimicrobial susceptibility testing of bacteria directly from positive blood culture bottles in a clinical setting. A total of 100 non-duplicated positive blood cultures were tested and 1012 microorganism-antimicrobial combinations were assessed. An aliquot of non-charcoal blood culture broth was incubated with lysis buffer briefly before being filtered and washed. Microorganisms recovered from the filter membrane were first identified by using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight VITEK® Mass Spectrometry (VITEK MS. After quick identification from VITEK MS, filtered microorganisms were inoculated to VITEK®2 system for full panel antimicrobial susceptibility testing analysis. Of 100 bottles tested, the VITEK MS resulted in 94.0% correct organism identification to the species level. Compared to the conventional antimicrobial susceptibility testing methods, direct antimicrobial susceptibility testing from VITEK®2 resulted in 93.5% (946/1012 category agreement of antimicrobials tested, with 3.6% (36/1012 minor error, 1.7% (7/1012 major error, and 1.3% (13/1012 very major error of antimicrobials. The average time to identification and antimicrobial susceptibility testing was 11.4 hours by using the Lysis-Filtration method for both VITEK MS and VITEK®2 compared to 56.3 hours by using conventional methods (p<0.00001. Thus, the same-day results of microorganism identification and antimicrobial susceptibility testing directly from positive blood culture can be achieved and can be used for appropriate antibiotic therapy and antibiotic stewardship.

Same Day Identification and Full Panel Antimicrobial Susceptibility Testing of Bacteria from Positive Blood Culture Bottles Made Possible by a Combined Lysis-Filtration Method with MALDI-TOF VITEK Mass Spectrometry and the VITEK2 System

Science.gov (United States)

Machen, Alexandra; Drake, Tim; Wang, Yun F. (Wayne)

2014-01-01

Rapid identification and antimicrobial susceptibility testing of microorganisms causing bloodstream infections or sepsis have the potential to improve patient care. This proof-of-principle study evaluates the Lysis-Filtration Method for identification as well as antimicrobial susceptibility testing of bacteria directly from positive blood culture bottles in a clinical setting. A total of 100 non-duplicated positive blood cultures were tested and 1012 microorganism-antimicrobial combinations were assessed. An aliquot of non-charcoal blood culture broth was incubated with lysis buffer briefly before being filtered and washed. Microorganisms recovered from the filter membrane were first identified by using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight VITEK® Mass Spectrometry (VITEK MS). After quick identification from VITEK MS, filtered microorganisms were inoculated to VITEK®2 system for full panel antimicrobial susceptibility testing analysis. Of 100 bottles tested, the VITEK MS resulted in 94.0% correct organism identification to the species level. Compared to the conventional antimicrobial susceptibility testing methods, direct antimicrobial susceptibility testing from VITEK®2 resulted in 93.5% (946/1012) category agreement of antimicrobials tested, with 3.6% (36/1012) minor error, 1.7% (7/1012) major error, and 1.3% (13/1012) very major error of antimicrobials. The average time to identification and antimicrobial susceptibility testing was 11.4 hours by using the Lysis-Filtration method for both VITEK MS and VITEK®2 compared to 56.3 hours by using conventional methods (p<0.00001). Thus, the same-day results of microorganism identification and antimicrobial susceptibility testing directly from positive blood culture can be achieved and can be used for appropriate antibiotic therapy and antibiotic stewardship. PMID:24551067

A differential centrifugation protocol and validation criterion for enhancing mass spectrometry (MALDI-TOF) results in microbial identification using blood culture growth bottles.

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March-Rosselló, G A; Muñoz-Moreno, M F; García-Loygorri-Jordán de Urriés, M C; Bratos-Pérez, M A

2013-05-01

Matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF) is a widely used tool in clinical microbiology for rapidly identifying microorganisms. This technique can be applied directly on positive blood cultures without the need for its culturing, thereby, reducing the time required for microbiological diagnosis. The present study proposes an innovative identification protocol applied to positive blood culture bottles using MALDI-TOF. We have processed 100 positive blood culture bottles, of which 36 of 37 Gram-negative bacteria (97.3 %) were correctly identified directly with 100 % of Enterobacteriaceae and other Gram-negative rods and 87.5 % of non-fermenting Gram-negative rods. We also correctly identified directly 62 of 63 of Gram-positive bacteria (98.4 %) with 100 % of Streptococcus, Enterococcus, and Gram-positive bacilli and 98 % of Staphylococcus. Applying the differential centrifugation protocol at the moment the automatic blood culture incubation system gives a positive reading together with the proposed validation criterion offers 98 % sensitivity (95 % confidence interval: 95.2-100 %). The MALDI-TOF system, thus, provides a rapid and reliable system for identifying microorganisms from blood culture growth bottles.

Enhanced infectivity of bluetongue virus in cell culture by centrifugation.

OpenAIRE

Sundin, D R; Mecham, J O

1989-01-01

The effects of centrifugation of the infection of cell culture with bluetongue virus (BTV) were investigated. Baby hamster kidney cells were infected with BTV with or without centrifugation. Viral antigen was detected by immunofluorescence at 24 h in both centrifuged and noncentrifuged cultures. However, after 24 h of infection, the production of PFU in centrifuged cell cultures was 10- to 20-fold greater than that seen in cultures not centrifuged. In addition, centrifugation enhanced the dir...

Direct blood culturing on solid medium outperforms an automated continuously monitored broth-based blood culture system in terms of time to identification and susceptibility testing

Directory of Open Access Journals (Sweden)

E.A. Idelevich

2016-03-01

Full Text Available Pathogen identification and antimicrobial susceptibility testing (AST should be available as soon as possible for patients with bloodstream infections. We investigated whether a lysis-centrifugation (LC blood culture (BC method, combined with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS identification and Vitek 2 AST, provides a time advantage in comparison with the currently used automated broth-based BC system. Seven bacterial reference strains were added each to 10 mL human blood in final concentrations of 100, 10 and 1 CFU/mL. Inoculated blood was added to the Isolator 10 tube and centrifuged at 3000 g for 30 min, then 1.5 mL sediment was distributed onto five 150-mm agar plates. Growth was observed hourly and microcolonies were subjected to MALDI-TOF MS and Vitek 2 as soon as possible. For comparison, seeded blood was introduced into an aerobic BC bottle and incubated in the BACTEC 9240 automated BC system. For all species/concentration combinations except one, successful identification and Vitek 2 inoculation were achieved even before growth detection by BACTEC. The fastest identification and inoculation for AST were achieved with Escherichia coli in concentrations of 100 CFU/mL and 10 CFU/mL (after 7 h each, while BACTEC flagged respective samples positive after 9.5 h and 10 h. Use of the LC-BC method allows skipping of incubation in automated BC systems and, used in combination with rapid diagnostics from microcolonies, provides a considerable advantage in time to result. This suggests that the usefulness of direct BC on solid medium should be re-evaluated in the era of rapid microbiology.

Container for centrifuging blood

International Nuclear Information System (INIS)

Narra, R.K.

1982-01-01

A container is described for use in drawing patient's blood, centrifuging the blood and then labelling the separated red cells with sup(99m)Tc. It consists of a tube with a central chamber and a lower portion provided with an aperture in which a weir is supported and extends into the central chamber. The weir has a central channel. A resilient plug seals the aperture. (author)

Simple Sample Preparation Method for Direct Microbial Identification and Susceptibility Testing From Positive Blood Cultures.

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Pan, Hong-Wei; Li, Wei; Li, Rong-Guo; Li, Yong; Zhang, Yi; Sun, En-Hua

2018-01-01

Rapid identification and determination of the antibiotic susceptibility profiles of the infectious agents in patients with bloodstream infections are critical steps in choosing an effective targeted antibiotic for treatment. However, there has been minimal effort focused on developing combined methods for the simultaneous direct identification and antibiotic susceptibility determination of bacteria in positive blood cultures. In this study, we constructed a lysis-centrifugation-wash procedure to prepare a bacterial pellet from positive blood cultures, which can be used directly for identification by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) and antibiotic susceptibility testing by the Vitek 2 system. The method was evaluated using a total of 129 clinical bacteria-positive blood cultures. The whole sample preparation process could be completed in identification was 96.49% for gram-negative bacteria and 97.22% for gram-positive bacteria. Vitek 2 antimicrobial susceptibility testing of gram-negative bacteria showed an agreement rate of antimicrobial categories of 96.89% with a minor error, major error, and very major error rate of 2.63, 0.24, and 0.24%, respectively. Category agreement of antimicrobials against gram-positive bacteria was 92.81%, with a minor error, major error, and very major error rate of 4.51, 1.22, and 1.46%, respectively. These results indicated that our direct antibiotic susceptibility analysis method worked well compared to the conventional culture-dependent laboratory method. Overall, this fast, easy, and accurate method can facilitate the direct identification and antibiotic susceptibility testing of bacteria in positive blood cultures.

Evaluation of three sample preparation methods for the direct identification of bacteria in positive blood cultures by MALDI-TOF

OpenAIRE

Tanner, Hannah; Evans, Jason T.; Gossain, Savita; Hussain, Abid

2017-01-01

Background Patient mortality is significantly reduced by rapid identification of bacteria from sterile sites. MALDI-TOF can identify bacteria directly from positive blood cultures and multiple sample preparation methods are available. We evaluated three sample preparation methods and two MALDI-TOF score cut-off values. Positive blood culture bottles with organisms present in Gram stains were prospectively analysed by MALDI-TOF. Three lysis reagents (Saponin, SDS, and SepsiTyper lysis bufer) w...

Simple Sample Preparation Method for Direct Microbial Identification and Susceptibility Testing From Positive Blood Cultures

Directory of Open Access Journals (Sweden)

Hong-wei Pan

2018-03-01

Full Text Available Rapid identification and determination of the antibiotic susceptibility profiles of the infectious agents in patients with bloodstream infections are critical steps in choosing an effective targeted antibiotic for treatment. However, there has been minimal effort focused on developing combined methods for the simultaneous direct identification and antibiotic susceptibility determination of bacteria in positive blood cultures. In this study, we constructed a lysis-centrifugation-wash procedure to prepare a bacterial pellet from positive blood cultures, which can be used directly for identification by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS and antibiotic susceptibility testing by the Vitek 2 system. The method was evaluated using a total of 129 clinical bacteria-positive blood cultures. The whole sample preparation process could be completed in <15 min. The correct rate of direct MALDI-TOF MS identification was 96.49% for gram-negative bacteria and 97.22% for gram-positive bacteria. Vitek 2 antimicrobial susceptibility testing of gram-negative bacteria showed an agreement rate of antimicrobial categories of 96.89% with a minor error, major error, and very major error rate of 2.63, 0.24, and 0.24%, respectively. Category agreement of antimicrobials against gram-positive bacteria was 92.81%, with a minor error, major error, and very major error rate of 4.51, 1.22, and 1.46%, respectively. These results indicated that our direct antibiotic susceptibility analysis method worked well compared to the conventional culture-dependent laboratory method. Overall, this fast, easy, and accurate method can facilitate the direct identification and antibiotic susceptibility testing of bacteria in positive blood cultures.

Direct identification of bacteria in positive blood culture bottles by matrix-assisted laser desorption ionisation time-of-flight mass spectrometry.

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Bernard La Scola

Full Text Available BACKGROUND: With long delays observed between sampling and availability of results, the usefulness of blood cultures in the context of emergency infectious diseases has recently been questioned. Among methods that allow quicker bacterial identification from growing colonies, matrix-assisted laser desorption ionisation time-of-flight (MALDI-TOF mass spectrometry was demonstrated to accurately identify bacteria routinely isolated in a clinical biology laboratory. In order to speed up the identification process, in the present work we attempted bacterial identification directly from blood culture bottles detected positive by the automate. METHODOLOGY/PRINCIPAL FINDINGS: We prospectively analysed routine MALDI-TOF identification of bacteria detected in blood culture by two different protocols involving successive centrifugations and then lysis by trifluoroacetic acid or formic acid. Of the 562 blood culture broths detected as positive by the automate and containing one bacterial species, 370 (66% were correctly identified. Changing the protocol from trifluoroacetic acid to formic acid improved identification of Staphylococci, and overall correct identification increased from 59% to 76%. Lack of identification was observed mostly with viridans streptococci, and only one false positive was observed. In the 22 positive blood culture broths that contained two or more different species, only one of the species was identified in 18 samples, no species were identified in two samples and false species identifications were obtained in two cases. The positive predictive value of bacterial identification using this procedure was 99.2%. CONCLUSIONS/SIGNIFICANCE: MALDI-TOF MS is an efficient method for direct routine identification of bacterial isolates in blood culture, with the exception of polymicrobial samples and viridans streptococci. It may replace routine identification performed on colonies, provided improvement for the specificity of blood culture

Direct identification of bacteria in positive blood culture bottles by matrix-assisted laser desorption ionisation time-of-flight mass spectrometry.

Science.gov (United States)

La Scola, Bernard; Raoult, Didier

2009-11-25

With long delays observed between sampling and availability of results, the usefulness of blood cultures in the context of emergency infectious diseases has recently been questioned. Among methods that allow quicker bacterial identification from growing colonies, matrix-assisted laser desorption ionisation time-of-flight (MALDI-TOF) mass spectrometry was demonstrated to accurately identify bacteria routinely isolated in a clinical biology laboratory. In order to speed up the identification process, in the present work we attempted bacterial identification directly from blood culture bottles detected positive by the automate. We prospectively analysed routine MALDI-TOF identification of bacteria detected in blood culture by two different protocols involving successive centrifugations and then lysis by trifluoroacetic acid or formic acid. Of the 562 blood culture broths detected as positive by the automate and containing one bacterial species, 370 (66%) were correctly identified. Changing the protocol from trifluoroacetic acid to formic acid improved identification of Staphylococci, and overall correct identification increased from 59% to 76%. Lack of identification was observed mostly with viridans streptococci, and only one false positive was observed. In the 22 positive blood culture broths that contained two or more different species, only one of the species was identified in 18 samples, no species were identified in two samples and false species identifications were obtained in two cases. The positive predictive value of bacterial identification using this procedure was 99.2%. MALDI-TOF MS is an efficient method for direct routine identification of bacterial isolates in blood culture, with the exception of polymicrobial samples and viridans streptococci. It may replace routine identification performed on colonies, provided improvement for the specificity of blood culture broths growing viridans streptococci is obtained in the near future.

Stretching single fibrin fibers hampers their lysis.

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Li, Wei; Lucioni, Tomas; Li, Rongzhong; Bonin, Keith; Cho, Samuel S; Guthold, Martin

2017-09-15

Blood clots, whose main structural component is a mesh of microscopic fibrin fibers, experience mechanical strain from blood flow, clot retraction and interactions with platelets and other cells. We developed a transparent, striated and highly stretchable substrate made from fugitive glue (a styrenic block copolymer) to investigate how mechanical strain affects lysis of single, suspended fibrin fibers. In this suspended fiber assay, lysis manifested itself by fiber elongation, thickening (disassembly), fraying and collapse. Stretching single fibrin fibers significantly hampered their lysis. This effect was seen in uncrosslinked and crosslinked fibers. Crosslinking (without stretching) also hampered single fiber lysis. Our data suggest that strain is a novel mechanosensitive factor that regulates blood clot dissolution (fibrinolysis) at the single fiber level. At the molecular level of single fibrin molecules, strain may distort, or hinder access to, plasmin cleavage sites and thereby hamper lysis. Fibrin fibers are the major structural component of a blood clot. We developed a highly stretchable substrate made from fugitive glue and a suspended fibrin fiber lysis assay to investigate the effect of stretching on single fibrin fibers lysis. The key findings from our experiments are: 1) Fibers thicken and elongate upon lysis; 2) stretching strongly reduces lysis; 3) this effect is more pronounced for uncrosslinked fibers; and 4) stretching fibers has a similar effect on reducing lysis as crosslinking fibers. At the molecular level, strain may distort plasmin cleavage sites, or restrict access to those sites. Our results suggest that strain may be a novel mechanobiological factor that regulates fibrinolysis. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

21 CFR 864.9275 - Blood bank centrifuge for in vitro diagnostic use.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Blood bank centrifuge for in vitro diagnostic use. 864.9275 Section 864.9275 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN... Manufacture Blood and Blood Products § 864.9275 Blood bank centrifuge for in vitro diagnostic use. (a...

Egg beater as centrifuge: isolating human blood plasma from whole blood in resource-poor settings.

Science.gov (United States)

Wong, Amy P; Gupta, Malancha; Shevkoplyas, Sergey S; Whitesides, George M

2008-12-01

This paper demonstrates that a hand-powered egg beater can be modified to serve as a centrifuge for separating plasma from human whole blood. Immunoassays used to diagnose infectious diseases often require plasma from whole blood, and obtaining plasma typically requires electrically-powered centrifuges, which are not widely available in resource-limited settings. Human whole blood was loaded into polyethylene (PE) tubing, and the tubing was attached to the paddle of an egg beater. Spinning the paddle pelleted the blood cells to the distal end of the PE tubing; the plasma remained as the supernatant. A cholesterol assay (run on patterned paper) demonstrated the suitability of this plasma for use in diagnostic assays. The physics of the system was also analyzed as a guide for the selection of other rotating systems for use in centrifugation. Egg beaters, polyethylene tubing, and paper are readily available devices and supplies that can facilitate the use of point-of-care diagnostics at sites far from centralized laboratory facilities.

The effect of increased centrifugation temperature on the quality of red-blood-cell concentrates of automated whole blood processing.

Science.gov (United States)

Weinigel, C; Rummler, S; Barz, D

2013-10-01

There are manual and automated methods to separate whole blood (WB) available. The Atreus whole blood processing system is an automated method, which combines centrifugation and expression of components into a single device. A major difference to conventional methods is that centrifugation temperature is not controlled at 22°C. The aim of this study was to examine the influence of increased centrifugation temperatures on the quality of red-blood-cell concentrates (RCC) after active cooling of WB prior to processing. A total of 28 WB were processed: 16 at centrifugation temperatures of up to 28°C (1st protocol) and 12 at 34°C (2nd protocol). RCC quality parameters were tested weekly for 42 days. Red-blood-cell concentrates (RCC) quality complied with the European and German guidelines. Haemolysis was not significantly different throughout storage. Significant statistical differences were detected between both protocols in potassium concentration at the end of storage and in ATP levels at the day of processing. Centrifugation temperatures of up to 34°C are well tolerated by the red blood cells with minimal interference with the RCC quality parameters. © 2013 International Society of Blood Transfusion.

Performance assessment of two lysis methods for direct identification of yeasts from clinical blood cultures using MALDI-TOF mass spectrometry.

Science.gov (United States)

Jeddi, Fakhri; Yapo-Kouadio, Gisèle Cha; Normand, Anne-Cécile; Cassagne, Carole; Marty, Pierre; Piarroux, Renaud

2017-02-01

In cases of fungal infection of the bloodstream, rapid species identification is crucial to provide adapted therapy and thereby ameliorate patient outcome. Currently, the commercial Sepsityper kit and the sodium-dodecyl sulfate (SDS) method coupled with MALDI-TOF mass spectrometry are the most commonly reported lysis protocols for direct identification of fungi from positive blood culture vials. However, the performance of these two protocols has never been compared on clinical samples. Accordingly, we performed a two-step survey on two distinct panels of clinical positive blood culture vials to identify the most efficient protocol, establish an appropriate log score (LS) cut-off, and validate the best method. We first compared the performance of the Sepsityper and the SDS protocols on 71 clinical samples. For 69 monomicrobial samples, mass spectrometry LS values were significantly higher with the SDS protocol than with the Sepsityper method (P < .0001), especially when the best score of four deposited spots was considered. Next, we established the LS cut-off for accurate identification at 1.7, based on specimen DNA sequence data. Using this LS cut-off, 66 (95.6%) and 46 (66.6%) isolates were correctly identified at the species level with the SDS and the Sepsityper protocols, respectively. In the second arm of the survey, we validated the SDS protocol on an additional panel of 94 clinical samples. Ninety-two (98.9%) of 93 monomicrobial samples were correctly identified at the species level (median LS = 2.061). Overall, our data suggest that the SDS method yields more accurate species identification of yeasts, than the Sepsityper protocol. © The Author 2016. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

MALDI-TOF identification of Gram-negative bacteria directly from blood culture bottles containing charcoal: Sepsityper® kits versus centrifugation-filtration method.

Science.gov (United States)

Riederer, Kathleen; Cruz, Kristian; Shemes, Stephen; Szpunar, Susan; Fishbain, Joel T

2015-06-01

Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry has dramatically altered the way microbiology laboratories identify clinical isolates. Direct blood culture (BC) detection may be hampered, however, by the presence of charcoal in BC bottles currently in clinical use. This study evaluates an in-house process for extraction and MALDI-TOF identification of Gram-negative bacteria directly from BC bottles containing charcoal. Three hundred BC aliquots were extracted by a centrifugation-filtration method developed in our research laboratory with the first 96 samples processed in parallel using Sepsityper® kits. Controls were colonies from solid media with standard phenotypic and MALDI-TOF identification. The identification of Gram-negative bacteria was successful more often via the in-house method compared to Sepsityper® kits (94.7% versus 78.1%, P≤0.0001). Our in-house centrifugation-filtration method was further validated for isolation and identification of Gram-negative bacteria (95%; n=300) directly from BC bottles containing charcoal. Copyright © 2015 Elsevier Inc. All rights reserved.

Lysis of Gymnodinium breve by cultures of the green alga Nannochloris eucaryotum.

Science.gov (United States)

Pérez, E; Sawyers, W G; Martin, D F

2001-01-01

Laboratory cultures of Florida's red tide organism, Gymnodinium breve, were killed by the green alga Nannochloris eucaryotum. Studies involved organism-organism interaction as well as organism-cell-free culture (N. eucaryotum) interaction. Both studies demonstrated that N. eucaryotum adversely affected Florida's red tide organism. The lysis has been attributed to compounds called APONINs (apparent oceanic naturally occurring cytolins). N. eucaryotum crude APONIN was extracted from cell-free cultures, partially purified and fractionated. The fractions were bioassayed against G. breve, and 'fingerprints' of the deleterious fractions were obtained.

Kinetically limited differential centrifugation as an inexpensive and readily available alternative to centrifugal elutriation.

Science.gov (United States)

Tan, Jinwang; Lee, Byung-Doo; Polo-Parada, Luis; Sengupta, Shramik

2012-08-01

When separating two species with similar densities but differing sedimentation velocities (because of differences in size), centrifugal elutriation is generally the method of choice. However, a major drawback to this approach is the requirement for specialized equipment. Here, we present a new method that achieves similar separations using standard benchtop centrifuges by loading the seperands as a layer on top of a dense buffer of a specified length, and running the benchtop centrifugation process for a calculated amount of time, thereby ensuring that all faster moving species are collected at the bottom, while all slower moving species remain in the buffer. We demonstrate the use of our procedure to isolate bacteria from blood culture broth (a mixture of bacterial growth media, blood, and bacteria).

Effect of hypergravity on catecholamine levels in telemetrically collected blood of rats during centrifugation

Science.gov (United States)

Kvetnansky, R.; Petrak, J.; Mravec, B.; Tillinger, A.; Jurani, M.; Baranovska, M.; Hapala, I.; Frollo, I.

2005-08-01

Hypergravity is known to activate the sympathoadrenal system (SAS). Rats subjected to various accelerations (+G) exhibited increased levels of plasma epinephrine (EPI) and partly also norepinephrine (NE). However, the collection of blood was performed after centrifugation finished and therefore plasma NE and EPI levels could have been affected by the process of deceleration. The aim of this study was to evaluate plasma EPI and NE levels in blood collected directly during the centrifugation after reaching different +G, using newly developed remote controlled equipment. Such telemetrically regulated equipment for multiple blood sampling allows us to investigate selective effects of hypergravity during centrifugation. All animals had a polyethylene tubing in the tail artery which was connected to a pre-programmed device for three blood withdrawals (0.6 ml each) into individual syringes, performed at any chosen time intervals. Plasma EPI and NE levels were measured at hypergravity between +1G - +5G. Plasma EPI levels showed a huge, hypergravity dependent increase at the interval of 10-20 min. After the blood collection was completed, the centrifuge was turned off and another blood sampling was performed immediately after the centrifuge stopped (10 min). In these samples plasma EPI levels showed a significant reduction compared to the 20 min interval of centrifugation but the EPI levels at 4G-6G were still significantly elevated compared to pre- centrifugation levels. Plasma NE levels showed less pronounced changes (increased after 6G only) with a slower return to control levels.Thus, our data has shown completely different responses of the adrenomedullary (epinephrine) and sympathoneural (norepinephrine) systems to hypergravitation. This data shows that the increased gravitation and not the stressful situations connected with centrifugation is the factor responsible for massive activation of the adrenomedullary system. The mechanism of small activation of the

Lab-on-a-Disc Platform for Automated Chemical Cell Lysis

Directory of Open Access Journals (Sweden)

Moo-Jung Seo

2018-02-01

Full Text Available Chemical cell lysis is an interesting topic in the research to Lab-on-a-Disc (LOD platforms on account of its perfect compatibility with the centrifugal spin column format. However, standard procedures followed in chemical cell lysis require sophisticated non-contact temperature control as well as the use of pressure resistant valves. These requirements pose a significant challenge thereby making the automation of chemical cell lysis on an LOD extremely difficult to achieve. In this study, an LOD capable of performing fully automated chemical cell lysis is proposed, where a combination of chemical and thermal methods has been used. It comprises a sample inlet, phase change material sheet (PCMS-based temperature sensor, heating chamber, and pressure resistant valves. The PCMS melts and solidifies at a certain temperature and thus is capable of indicating whether the heating chamber has reached a specific temperature. Compared to conventional cell lysis systems, the proposed system offers advantages of reduced manual labor and a compact structure that can be readily integrated onto an LOD. Experiments using Salmonella typhimurium strains were conducted to confirm the performance of the proposed cell lysis system. The experimental results demonstrate that the proposed system has great potential in realizing chemical cell lysis on an LOD whilst achieving higher throughput in terms of purity and yield of DNA thereby providing a good alternative to conventional cell lysis systems.

Lab-on-a-Disc Platform for Automated Chemical Cell Lysis.

Science.gov (United States)

Seo, Moo-Jung; Yoo, Jae-Chern

2018-02-26

Chemical cell lysis is an interesting topic in the research to Lab-on-a-Disc (LOD) platforms on account of its perfect compatibility with the centrifugal spin column format. However, standard procedures followed in chemical cell lysis require sophisticated non-contact temperature control as well as the use of pressure resistant valves. These requirements pose a significant challenge thereby making the automation of chemical cell lysis on an LOD extremely difficult to achieve. In this study, an LOD capable of performing fully automated chemical cell lysis is proposed, where a combination of chemical and thermal methods has been used. It comprises a sample inlet, phase change material sheet (PCMS)-based temperature sensor, heating chamber, and pressure resistant valves. The PCMS melts and solidifies at a certain temperature and thus is capable of indicating whether the heating chamber has reached a specific temperature. Compared to conventional cell lysis systems, the proposed system offers advantages of reduced manual labor and a compact structure that can be readily integrated onto an LOD. Experiments using Salmonella typhimurium strains were conducted to confirm the performance of the proposed cell lysis system. The experimental results demonstrate that the proposed system has great potential in realizing chemical cell lysis on an LOD whilst achieving higher throughput in terms of purity and yield of DNA thereby providing a good alternative to conventional cell lysis systems.

Normal human serum (HS) prevents oxidant-induced lysis of cultured endothelial cells (ECs)

International Nuclear Information System (INIS)

Callahan, K.S.; Harlan, J.M.

1986-01-01

Most studies demonstrating oxidant lysis of cultured ECs are performed in serum-free media or media containing low concentrations of bovine serum. The authors found that HS protects human and bovine ECs from lysis caused by reagent H 2 O 2 or glucose/glucose oxidase (GO)-generated H 2 O 2 . EC injury was assessed by 51 Cr release, cell detachment, or trypan blue dye exclusion. Protective HS activity was dose-dependent with concentrations greater than or equal to 25% preventing lethal injury. Cytotoxicity at 24 hrs, induced by 20 mU/ml GO, was 90.1 +/- 5.2% without HS vs 1.7 +/- 4.6% with 25% HS present (20 exp). Similar protection was observed with heparinized plasma. Of note, comparable concentrations of bovine serum were devoid of protective activity. Addition of fatty acid-free albumin to the media was also without protective effect. Preliminary characterization showed HS activity was stable to 60 0 C for 30 min, non-dialyzable at 25,000 MW cutoff, and retained in delipidated serum. The HS protection was not merely due to scavenging of exogenous H 2 O 2 as A23187-induced EC lysis was also prevented by HS. Protective activity was not reproduced by purified cerruloplasmin or transferrin. In conclusion, unidentified factor(s) present in HS protect cultured ECs from oxidant-induced lysis. Since endothelium is normally exposed to 100% plasma, the authors suggest that in vitro studies of oxidant-mediated injury be performed in the presence of HS. Factor(s) in HS may play an important role in modulating oxidant-induced vascular injury in vivo

Leukoreduction by centrifugation does not eliminate Trypanosoma cruzi from infected blood units.

Science.gov (United States)

Dzib, Doris; Hernández, Virginia Peña; Ake, Baldemar Canche; López, Ruth Alacantara; Monteón, Victor Manuel

2009-06-01

Current strategies to prevent transfusion-associated Chagas disease include the identification of Trypanosoma cruzi-infected blood donors through questionnaires and serologic tests. There are other procedures such as leukoreduction that prevent the transmission of infectious agents associated to white cells. The objective of the present work was to estimate the seroprevalence, evaluate the efficacy of leukoreduction by centrifugation to eliminate T. cruzi in infected blood units, and the correlation of immunoglobulin G (IgG) subclasses of seropositive blood donors with chronic chagasic cardiopathy. Over a period of 14 months, 33 out of 6600 blood donors (0.5%) at Centro Estatal de la Transfusión Sanguínea in Campeche State, México were seropositive for T. cruzi. Twenty seropositive blood units were submitted through leukoreduction by centrifugation, and in the fractions generated (red cell fraction, platelets, and the buffy-coat), we searched for the presence of T. cruzi using specific polymerase chain reaction. We detected parasite DNA in 50% to 60% of the fractions tested, suggesting that leukoreduction by centrifugation does not eliminate the microorganisms in the infected blood unit. We also observed that the level of IgG2 and IgG4 subclasses specific for T. cruzi in seropositive blood donors was lower than in chronic cardiopathic chagasic patients. In conclusion, leukoreduction by centrifugation has a limited role in eliminating T. cruzi in infected blood supply, and the low level of specific IgG2 and IgG4 could be a marker in the indeterminate phase of infection.

Direct identification of microorganisms from positive blood cultures by MALDI-TOF MS using an in-house saponin method.

Science.gov (United States)

Yonetani, Shota; Ohnishi, Hiroaki; Ohkusu, Kiyofumi; Matsumoto, Tetsuya; Watanabe, Takashi

2016-11-01

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a fast and reliable method for the identification of bacteria. A MALDI Sepsityper kit is generally used to prepare samples obtained directly from culture bottles. However, the relatively high cost of this kit is a major obstacle to introducing this method into routine clinical use. In this study, the accuracies of three different preparation methods for rapid direct identification of bacteria from positive blood culture bottles by MALDI-TOF MS analysis were compared. In total, 195 positive bottles were included in this study. Overall, 78.5%, 68.7%, and 76.4% of bacteria were correctly identified to the genus level (score ≥1.7) directly from positive blood cultures using the Sepsityper, centrifugation, and saponin methods, respectively. The identification rates using the Sepsityper and saponin methods were significantly higher than that using the centrifugation method (Sepsityper vs. centrifugation, pdirectly from blood culture bottles, and could be a less expensive alternative to the Sepsityper method. Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Sporothrix schenckii fungemia without disseminated sporotrichosis.

OpenAIRE

Kosinski, R M; Axelrod, P; Rex, J H; Burday, M; Sivaprasad, R; Wreiole, A

1992-01-01

Fungemia is a rare complication of Sporothrix schenckii infection and has always been associated with disseminated sporotrichosis. We describe an immunocompetent patient with localized lymphocutaneous sporotrichosis from whose blood the fungus was isolated. A lysis-centrifugation blood culture system may have improved our ability to detect low-level S. schenckii fungemia.

Tuberculosis diagnosis and multidrug resistance testing by direct sputum culture in selective broth without decontamination or centrifugation.

Science.gov (United States)

Grandjean, Louis; Martin, Laura; Gilman, Robert H; Valencia, Teresa; Herrera, Beatriz; Quino, Willi; Ramos, Eric; Rivero, Maribel; Montoya, Rosario; Escombe, A Roderick; Coleman, David; Mitchison, Denis; Evans, Carlton A

2008-07-01

Tuberculosis culture usually requires sputum decontamination and centrifugation to prevent cultures from being overgrown by contaminating bacteria and fungi. However, decontamination destroys many tuberculous bacilli, and centrifugation often is not possible in resource-poor settings. We therefore assessed the performance of Mycobacterium tuberculosis culture with unprocessed samples plated directly by using tuberculosis-selective media and compared this procedure to conventional culture using centrifuge decontamination. Quadruplicate aliquots of strain H37RV were cultured in 7H9 broth with and without selective antimicrobials and after centrifuge decontamination. The subsequent comparison was made with 715 sputum samples. Split paired sputum samples were cultured conventionally with centrifuge decontamination and by direct culture in tuberculosis-selective media containing antibiotics. Centrifuge decontamination reduced tuberculosis H37RV colonies by 78% (P laboratories this deficit may be outweighed by the ease of use.

Hypergravity-induced increase in plasma catecholamine and corticosterone levels in telemetrically collected blood of rats during centrifugation.

Science.gov (United States)

Petrak, Juraj; Mravec, Boris; Jurani, Marian; Baranovska, Magda; Tillinger, Andrej; Hapala, Ivan; Frollo, Ivan; Kvetnanský, Richard

2008-12-01

Rats subjected to various accelerations (+G) exhibited increased levels of plasma epinephrine (EPI), norepinephrine (NE), and corticosterone. However, the collection of blood was performed after a centrifugation finished, and therefore the levels could be affected by the process of deceleration. The aim of this study was to evaluate plasma EPI, NE, and corticosterone levels in blood collected directly during centrifugation after reaching different G (2-6), using newly developed remote-controlled equipment. Animals placed into the centrifuge cabins had inserted polyethylene tubing in the tail artery, which was connected with a preprogrammed device for blood withdrawals. Plasma EPI, NE, and corticosterone levels were measured at different time intervals of hypergravity of 2-6G. Plasma EPI levels showed a huge, hypergravity-level-dependent increase. After the last blood collection was completed during hypergravity, the centrifuge was turned off and another blood sampling was performed immediately after the centrifuge stopped (10 min). In these samples, plasma EPI showed significantly lower levels compared to centrifugation intervals. Plasma NE levels were significantly increased after 6G only. The increase in plasma corticosterone was dependent on level of G, however after the centrifuge stopped, corticosterone levels remained elevated. Thus, our data show that hypergravity highly activates the adrenomedullary and hypothalamo-pituitary-adrenocortical systems, whereas the sympathoneural system is activated only at high hypergravity. Immediately after centrifugation is over, EPI levels quickly return to control values. Our technique of blood collection during centrifugation allows assessment of the real hormonal levels at the particular hypergravity value.

Mass entrapment and lysis of Mesodinium rubrum cells in mucus threads observed in cultures with Dinophysis

DEFF Research Database (Denmark)

Ojamäe, Karin; Hansen, Per Juel; Lips, Inga

2016-01-01

The entrapment and death of the ciliate Mesodinium rubrum in the mucus threads in cultures with Dinophysis is described and quantified. Feeding experiments with different concentrations and predator–prey ratios of Dinophysis acuta, Dinophysis acuminata and M. rubrum to study the motility loss...... and aggregate formation of the ciliates and the feeding behaviour of Dinophysis were carried out. In cultures of either Dinophysis species, the ciliates became entrapped in the mucus, which led to the formation of immobile aggregates of M. rubrum and subsequent cell lysis. The proportion of entrapped ciliates...... was influenced by the concentration of Dinophysis and the ratio of predator and prey in the cultures. At high cell concentrations of prey (136 cells mL−1) and predator (100 cells mL−1), a maximum of 17% of M. rubrum cells became immobile and went through cell lysis. Ciliates were observed trapped in the mucus...

Flow cytometric assay for analysis of cytotoxic effects of potential drugs on human peripheral blood leukocytes

Science.gov (United States)

Nieschke, Kathleen; Mittag, Anja; Golab, Karolina; Bocsi, Jozsef; Pierzchalski, Arkadiusz; Kamysz, Wojciech; Tarnok, Attila

2014-03-01

Toxicity test of new chemicals belongs to the first steps in the drug screening, using different cultured cell lines. However, primary human cells represent the human organism better than cultured tumor derived cell lines. We developed a very gentle toxicity assay for isolation and incubation of human peripheral blood leukocytes (PBL) and tested it using different bioactive oligopeptides (OP). Effects of different PBL isolation methods (red blood cell lysis; Histopaque isolation among others), different incubation tubes (e.g. FACS tubes), anticoagulants and blood sources on PBL viability were tested using propidium iodide-exclusion as viability measure (incubation time: 60 min, 36°C) and flow cytometry. Toxicity concentration and time-depended effects (10-60 min, 36 °C, 0-100 μg /ml of OP) on human PBL were analyzed. Erythrocyte lysis by hypotonic shock (dH2O) was the fastest PBL isolation method with highest viability (>85%) compared to NH4Cl-Lysis (49%). Density gradient centrifugation led to neutrophil granulocyte cell loss. Heparin anticoagulation resulted in higher viability than EDTA. Conical 1.5 mL and 2 mL micro-reaction tubes (both polypropylene (PP)) had the highest viability (99% and 97%) compared to other tubes, i.e. three types of 5.0 mL round-bottom tubes PP (opaque-60%), PP (blue-62%), Polystyrene (PS-64%). Viability of PBL did not differ between venous and capillary blood. A gentle reproducible preparation and analytical toxicity-assay for human PBL was developed and evaluated. Using our assay toxicity, time-course, dose-dependence and aggregate formation by OP could be clearly differentiated and quantified. This novel assay enables for rapid and cost effective multiparametric toxicological screening and pharmacological testing on primary human PBL and can be adapted to high-throughput-screening.°z

Effect of Isolation Techniques on Viability of Bovine Blood Neutrophils

Directory of Open Access Journals (Sweden)

P. Sláma

2006-01-01

Full Text Available The effect of selected isolation methods on the viability of neutrophil granulocytes (neutrophils from the blood of healthy Holstein x Bohemian Red Pied crossbred heifers was evaluated. Two methods of neutrophil isolation were used: a neutrophil isolation on the basis of hypotonic erythrocyte lysis (in two variants: after the erythrocyte lysis proper, the cells were centrifuged at either 200 g or 1000 g, and b neutrophil isolation with FACS Lysing Solution as the lysing agent. The viability of the isolated neutrophils was evaluated on the basis of apoptosis and necrosis. The results obtained with flow cytometry (FCM suggest that, from the isolation techniques used, the method based on FACS Lysing Solution impaired the neutrophil viability least. After the application of this method, 5.36 ± 2.15% of neutrophils were apoptotic and 0.51 ± 0.12% were necrotic. In contrast, when the hypotonic erythrocyte lysis was used, the proportion of apoptotic neutrophils amounted to 42.14 ± 7.12% and 49.00 ± 14.70%, respectively, and 41.12 ± 5.55% and 36.91 ± 24.38% respectively of necrotic neutrophils (P < 0.01. This was also confirmed by the light microscopy. After the isolation with FASC Lysing Solution, 1.92 ± 1.74% of neutrophils were apoptotic and 1.05 ± 0.76% were necrotic, as distinct from after the hypotonic erythrocyte lysis where 9.43 ± 3.69% of neutrophils were apoptotic and 12.67 ± 4.74% of necrotic after centrifugation at 200 g, while 12.60 ± 4.35 were apoptotic and 14.96 ± 12.64% were necrotic after centrifugation at 1000 g. It follows from the above-mentioned data that hypotonic lysis is not a suitable method for the isolation of neutrophils, as the method itself markedly affects cell viability.

Detection and quantification of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis and Streptococcus oralis in blood samples with different microbiological identification methods: An in vitro study.

Science.gov (United States)

Marin, María José; Ambrosio, Nagore; Virto, Leire; Diz, Pedro; Álvarez, Maximiliano; Herrera, David; Sanz, Mariano; Figuero, Elena

2017-02-01

Culture-based methods (culture broth bottles or lysis methods) have been the standard for detecting bacteremia. More recently, quantitative polymerase chain reaction (qPCR) was proposed as a more sensitive and specific test although none of them has been validated for the identification of periodontal pathogens (fastidious growing bacteria) in blood samples. To compare the ability to detect and quantify Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis and Streptococcus oralis (alone or in combination) in blood samples with three culture techniques [direct anaerobic culturing (DAC), haemo-culture (BACTEC), and lysis-centrifugation (LC)] and a non-culture dependent approach (qPCR) in an in vitro study. Blood samples from 12 periodontally healthy volunteers were contaminated with three concentrations [10 4 ,10 2 and 10 1 colony forming units (CFU)/mL] of A. actinomycetemcomitans, P. gingivalis and S. oralis, alone or in combination. Samples were analysed by DAC, BACTEC, LC and qPCR. Sensitivity, specificity, predictive values, kappa index and Lińs correlation coefficients were calculated. DAC, LC and qPCR were able to detect the three target species at all concentrations. An excellent concordance (correlation coefficient r: 0.92-1) was observed between DAC and the reference standard (sensitivity raging 93.33-100% and specificity 88.89-100%) values. BACTEC was not able to identify P. gingivalis in any of the performed experiments. qPCR provided false negative results for S.oralis. DAC showed the best results for the proper identification and quantification of A. actinomycetemcomitans, P. gingivalis and S. oralis, alone or in combination, in blood samples. Copyright © 2016 Elsevier Ltd. All rights reserved.

A portable system for processing donated whole blood into high quality components without centrifugation.

Science.gov (United States)

Gifford, Sean C; Strachan, Briony C; Xia, Hui; Vörös, Eszter; Torabian, Kian; Tomasino, Taylor A; Griffin, Gary D; Lichtiger, Benjamin; Aung, Fleur M; Shevkoplyas, Sergey S

2018-01-01

The use of centrifugation-based approaches for processing donated blood into components is routine in the industrialized world, as disparate storage conditions require the rapid separation of 'whole blood' into distinct red blood cell (RBC), platelet, and plasma products. However, the logistical complications and potential cellular damage associated with centrifugation/apheresis manufacturing of blood products are well documented. The objective of this study was to evaluate a proof-of-concept system for whole blood processing, which does not employ electromechanical parts, is easily portable, and can be operated immediately after donation with minimal human labor. In a split-unit study (n = 6), full (~500mL) units of freshly-donated whole blood were divided, with one half processed by conventional centrifugation techniques and the other with the new blood separation system. Each of these processes took 2-3 hours to complete and were performed in parallel. Blood products generated by the two approaches were compared using an extensive panel of cellular and plasma quality metrics. Comparison of nearly all RBC parameters showed no significant differences between the two approaches, although the portable system generated RBC units with a slight but statistically significant improvement in 2,3-diphosphoglyceric acid concentration (p applications in remote or resource-limited settings, or for patients requiring highly functional platelet product.

All-in-one nanowire-decorated multifunctional membrane for rapid cell lysis and direct DNA isolation.

KAUST Repository

So, Hongyun; Lee, Kunwoo; Murthy, Niren; Pisano, Albert P

2014-01-01

This paper describes a handheld device that uses an all-in-one membrane for continuous mechanical cell lysis and rapid DNA isolation without the assistance of power sources, lysis reagents, and routine centrifugation. This nanowire-decorated multifunctional membrane was fabricated to isolate DNA by selective adsorption to silica surface immediately after disruption of nucleus membranes by ultrasharp tips of nanowires for a rapid cell lysis, and it can be directly assembled with commercial syringe filter holders. The membrane was fabricated by photoelectrochemical etching to create microchannel arrays followed by hydrothermal synthesis of nanowires and deposition of silica. The proposed membrane successfully purifies high-quality DNA within 5 min, whereas a commercial purification kit needs more than an hour.

All-in-one nanowire-decorated multifunctional membrane for rapid cell lysis and direct DNA isolation.

KAUST Repository

So, Hongyun

2014-11-24

This paper describes a handheld device that uses an all-in-one membrane for continuous mechanical cell lysis and rapid DNA isolation without the assistance of power sources, lysis reagents, and routine centrifugation. This nanowire-decorated multifunctional membrane was fabricated to isolate DNA by selective adsorption to silica surface immediately after disruption of nucleus membranes by ultrasharp tips of nanowires for a rapid cell lysis, and it can be directly assembled with commercial syringe filter holders. The membrane was fabricated by photoelectrochemical etching to create microchannel arrays followed by hydrothermal synthesis of nanowires and deposition of silica. The proposed membrane successfully purifies high-quality DNA within 5 min, whereas a commercial purification kit needs more than an hour.

A Centrifugal Microfluidic Platform That Separates Whole Blood Samples into Multiple Removable Fractions Due to Several Discrete but Continuous Density Gradient Sections

Science.gov (United States)

Moen, Scott T.; Hatcher, Christopher L.; Singh, Anup K.

2016-01-01

We present a miniaturized centrifugal platform that uses density centrifugation for separation and analysis of biological components in small volume samples (~5 μL). We demonstrate the ability to enrich leukocytes for on-disk visualization via microscopy, as well as recovery of viable cells from each of the gradient partitions. In addition, we simplified the traditional Modified Wright-Giemsa staining by decreasing the time, volume, and expertise involved in the procedure. From a whole blood sample, we were able to extract 95.15% of leukocytes while excluding 99.8% of red blood cells. This platform has great potential in both medical diagnostics and research applications as it offers a simpler, automated, and inexpensive method for biological sample separation, analysis, and downstream culturing. PMID:27054764

Rapid prototyping of centrifugal microfluidic modules for point of care blood testing

CSIR Research Space (South Africa)

Madzivhandila, Phophi

2016-11-01

Full Text Available We present modular centrifugal microfluidic devices that enable a series of blood tests to be performed towards a full blood count. The modular approach allows for rapid prototyping of device components in a generic format to complete different...

Virus-specific HLA-restricted lysis of herpes simplex virus-infected human monocytes and macrophages mediated by cytotoxic T lymphocytes

Energy Technology Data Exchange (ETDEWEB)

Torpey, D.J. III

1987-01-01

Freshly-isolated peripheral blood human monocytes and 5 day in vitro cultured macrophages were infected with herpes simplex virus type 1 (HSV-1), labeled with /sup 51/Cr, and used as target cells in a 12-14 hour cell-mediated cytotoxicity assay. Mononuclear leukocytes (MNL) from HSV-1 non-immune individuals, whether unstimulated or stimulated with HSV-1 antigen, did not mediate significant lysis of either target cell. HSV-immune MNL, both freshly-isolated and cultured for 5 days without antigen, demonstrated only low levels of natural killer (NK) cell-mediate lysis. MNL from HSV-immune individuals incubated for 5 days in vitro with HSV-1 antigen mediated significant virus-specific lysis of both target cells. Mean virus-specific lysis of autologous monocytes was 8.5(/+-/2.0)% compared to a three-fold greater virus-specific lysis of autologous macrophages. Greater than 70% of this lytic activity was mediated by Leu-11-negative, T3-positive cytotoxic T lymphocytes (CTL). Allogeneic target cells lacking a common HLA determinant were not significantly lysed while T8-positive CTL mediated infrequent lysis of target cells sharing a common HLA-A and/or HLA-B determinant. T4-positive lymphocytes were demonstrated to be the predominant cell mediating lysis of autologous target cells and allogeneic target cells sharing both HLA-A and/or HLA-B plus HLA-DR determinants with the CTL; the T4-positive cell was the sole CTL mediator of lysis of allogeneic target cells having a common HLA-DR determinant.

Virus-specific HLA-restricted lysis of herpes simplex virus-infected human monocytes and macrophages mediated by cytotoxic T lymphocytes

International Nuclear Information System (INIS)

Torpey, D.J. III.

1987-01-01

Freshly-isolated peripheral blood human monocytes and 5 day in vitro cultured macrophages were infected with herpes simplex virus type 1 (HSV-1), labeled with 51 Cr, and used as target cells in a 12-14 hour cell-mediated cytotoxicity assay. Mononuclear leukocytes (MNL) from HSV-1 non-immune individuals, whether unstimulated or stimulated with HSV-1 antigen, did not mediate significant lysis of either target cell. HSV-immune MNL, both freshly-isolated and cultured for 5 days without antigen, demonstrated only low levels of natural killer (NK) cell-mediate lysis. MNL from HSV-immune individuals incubated for 5 days in vitro with HSV-1 antigen mediated significant virus-specific lysis of both target cells. Mean virus-specific lysis of autologous monocytes was 8.5(/+-/2.0)% compared to a three-fold greater virus-specific lysis of autologous macrophages. Greater than 70% of this lytic activity was mediated by Leu-11-negative, T3-positive cytotoxic T lymphocytes (CTL). Allogeneic target cells lacking a common HLA determinant were not significantly lysed while T8-positive CTL mediated infrequent lysis of target cells sharing a common HLA-A and/or HLA-B determinant. T4-positive lymphocytes were demonstrated to be the predominant cell mediating lysis of autologous target cells and allogeneic target cells sharing both HLA-A and/or HLA-B plus HLA-DR determinants with the CTL; the T4-positive cell was the sole CTL mediator of lysis of allogeneic target cells having a common HLA-DR determinant

Microbial identification and automated antibiotic susceptibility testing directly from positive blood cultures using MALDI-TOF MS and VITEK 2.

Science.gov (United States)

Wattal, C; Oberoi, J K

2016-01-01

The study addresses the utility of Matrix Assisted Laser Desorption/Ionisation Time-Of-Flight mass spectrometry (MALDI-TOF MS) using VITEK MS and the VITEK 2 antimicrobial susceptibility testing (AST) system for direct identification (ID) and timely AST from positive blood culture bottles using a lysis-filtration method (LFM). Between July and December 2014, a total of 140 non-duplicate mono-microbial blood cultures were processed. An aliquot of positive blood culture broth was incubated with lysis buffer before the bacteria were filtered and washed. Micro-organisms recovered from the filter were first identified using VITEK MS and its suspension was used for direct AST by VITEK 2 once the ID was known. Direct ID and AST results were compared with classical methods using solid growth. Out of the 140 bottles tested, VITEK MS resulted in 70.7 % correct identification to the genus and/ or species level. For the 103 bottles where identification was possible, there was agreement in 97 samples (94.17 %) with classical culture. Compared to the routine method, the direct AST resulted in category agreement in 860 (96.5 %) of 891 bacteria-antimicrobial agent combinations tested. The results of direct ID and AST were available 16.1 hours before those of the standard approach on average. The combined use of VITEK MS and VITEK 2 directly on samples from positive blood culture bottles using a LFM technique can result in rapid and reliable ID and AST results in blood stream infections to result in early institution of targeted treatment. The combination of LFM and AST using VITEK 2 was found to expedite AST more reliably.

Numerical Calculation for Whirling Motion of a Centrifugal Blood Pump with Conical Spiral Groove Bearings

Science.gov (United States)

Shigemaru, Daichi; Tsukamoto, Hiroshi

2010-06-01

Whirling motion of a pump impeller was calculated for the centrifugal blood pump with Conical Spiral Groove Bearings to get a criterion for the instability of impeller whirling motion. The motion of the centrifugal blood pump impeller was calculated based on a spring damping model, and unsteady flow in the pump was computed using the commercial CFD package ANSYS CFX. Also the whirling motion of rotating impeller was measured using two displacement sensors fixed to the blood pump casing. The numerical calculations were done for the blood pump impeller with conical spiral groove bearings, and impeller whirling motion was evaluated.

Hypochlorite- and hypobromite-mediated radical formation and its role in cell lysis

DEFF Research Database (Denmark)

Hawkins, C L; Brown, B E; Davies, Michael Jonathan

2001-01-01

. In this study it is shown that HOBr induces red blood cell lysis at approximately 10-fold lower concentrations than HOCl, whereas with monocyte (THP1) and macrophage (J774) cells HOCl and HOBr induce lysis at similar concentrations. The role of radical formation during lysis has been investigated by EPR spin...... trapping, and it is shown that reaction of both oxidants with each cell type generates cell-derived radicals. Red blood cells exposed to nonlytic doses of HOCl generate novel nitrogen-centered radicals whose formation is GSH dependent. In contrast, HOBr gives rise to nitrogen-centered, membrane......-derived protein radicals. With lytic doses of either oxidant, protein (probably hemoglobin)-derived, nitrogen-centered radicals are observed. Unlike the red blood cells, treatment of monocytes and macrophages with HOCl gives significant radical formation only under conditions where cell lysis occurs concurrently...

Centrifuge apparatus

Science.gov (United States)

Sartory, Walter K.; Eveleigh, John W.

1976-01-01

A method and apparatus for operating a continuous flow blood separation centrifuge are provided. The hematocrit of the entrant whole blood is continuously maintained at an optimum constant value by the addition of plasma to the entrant blood. The hematocrit of the separated red cells is monitored to indicate the degree of separation taking place, thereby providing a basis for regulating the flow through the centrifuge.

Numerical analysis of the internal flow field in screw centrifugal blood pump based on CFD

Science.gov (United States)

Han, W.; Han, B. X.; Y Wang, H.; Shen, Z. J.

2013-12-01

As to the impeller blood pump, the high speed of the impeller, the local high shear force of the flow field and the flow dead region are the main reasons for blood damage. The screw centrifugal pump can effectively alleviate the problems of the high speed and the high shear stress for the impeller. The softness and non-destructiveness during the transfer process can effectively reduce the extent of the damage. By using CFD software, the characteristics of internal flow are analyzed in the screw centrifugal pump by exploring the distribution rules of the velocity, pressure and shear deformation rate of the blood when it flows through the impeller and the destructive effects of spiral blades on blood. The results show that: the design of magnetic levitation solves the sealing problems; the design of regurgitation holes solves the problem of the flow dead zone; the magnetic levitated microcirculation screw centrifugal pump can effectively avoid the vortex, turbulence and high shear forces generated while the blood is flowing through the pump. Since the distribution rules in the velocity field, pressure field and shear deformation rate of the blood in the blood pump are comparatively uniform and the gradient change is comparatively small, the blood damage is effectively reduced.

Application of CFD in Bioprocessing: Separation of mammalian cells using disc stack centrifuge during production of biotherapeutics.

Science.gov (United States)

Shekhawat, Lalita Kanwar; Sarkar, Jayati; Gupta, Rachit; Hadpe, Sandeep; Rathore, Anurag S

2018-02-10

Centrifugation continues to be one of the most commonly used unit operations for achieving efficient harvest of the product from the mammalian cell culture broth during production of therapeutic monoclonal antibodies (mAbs). Since the mammalian cells are known to be shear sensitive, optimal performance of the centrifuge requires a balance between productivity and shear. In this study, Computational Fluid Dynamics (CFD) has been successfully used as a tool to facilitate efficient optimization. Multiphase Eulerian-Eulerian model coupled with Gidaspow drag model along with Eulerian-Eulerian k-ε mixture turbulence model have been used to quantify the complex hydrodynamics of the centrifuge and thus evaluate the turbulent stresses generated by the centrifugal forces. An empirical model has been developed by statistical analysis of experimentally observed cell lysis data as a function of turbulent stresses. An operating window that offers the optimal balance between high productivity, high separation efficiency, and low cell damage has been identified by use of CFD modeling. Copyright © 2017 Elsevier B.V. All rights reserved.

Preparation of positive blood cultures for direct MALDI-ToF MS identification.

Science.gov (United States)

Robinson, Andrew M; Ussher, James E

2016-08-01

MALDI-ToF MS can be used to identify microorganisms directly from blood cultures. This study compared two methods of sample preparation. Similar levels of genus- (91% vs 90%) and species-level identifications (79% vs 74%) were obtained with differential centrifugation and SDS methods. The SDS method is faster and requires minimal handling. Copyright © 2016 Elsevier B.V. All rights reserved.

The production of collagenase by adherent mononuclear cells cultured from human peripheral blood.

Science.gov (United States)

Louie, J S; Weiss, J; Ryhänen, L; Nies, K M; Rantala-Ryhänen, S; Uitto, J

1984-12-01

Mononuclear cells were isolated from human peripheral blood by Ficoll-Hypaque centrifugation, and the cells adherent to plastic substrata were cultured in serum-free media supplemented with lactalbumin hydrolysate. These cell cultures, which consisted predominantly of monocyte-macrophages as judged by nonspecific esterase staining, accumulated collagenase in the medium. This collagenase resembled other vertebrate collagenases in that it cleaved native triple-helical type I collagen at a locus 3/4-length away from the amino-terminal end of the molecule. The collagenase activity was inhibited by Na2EDTA, dithiothreitol, and fetal calf serum, while the addition of Ca++ or N-ethylmaleimide enhanced the enzyme activity. The accumulation of collagenase in the culture media was markedly enhanced by the incubation of cells with concanavalin A or phorbol myristic acetate. In the presence of cycloheximide, the levels of collagenase activity were markedly reduced, suggesting that active protein synthesis was required to express the enzyme activity. In additional experiments, monocytes were further purified by counterflow centrifugation-elutriation. The collagenase production was markedly increased in cultures enriched in monocyte-macrophages and devoid of polymorphonuclear leukocytes. The accumulation of collagenase in monocyte cultures incubated for 48 hours in the presence of concanavalin A or phorbol myristic acetate was of the same order of magnitude as in parallel cultures containing the same number of polymorphonuclear leukocytes purified by Ficoll-Hypaque centrifugation and Plasmagel sedimentation.(ABSTRACT TRUNCATED AT 250 WORDS)

Rapid identification of microorganisms from positive blood cultures by MALDI-TOF mass spectrometry subsequent to very short-term incubation on solid medium.

Science.gov (United States)

Idelevich, E A; Schüle, I; Grünastel, B; Wüllenweber, J; Peters, G; Becker, K

2014-10-01

Rapid identification of the causative microorganism is important for appropriate antimicrobial therapy of bloodstream infections. Bacteria from positive blood culture (BC) bottles are not readily available for identification by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Lysis and centrifugation procedures suggested for direct MALDI-TOF MS from positive BCs without previous culture are associated with additional hands-on processing time and costs. Here, we describe an alternative approach applying MALDI-TOF MS from bacterial cultures incubated very briefly on solid medium. After plating of positive BC broth on Columbia blood agar (n = 165), MALDI-TOF MS was performed after 1.5, 2, 3, 4, 5, 6, 7, 8, 12 and (for control) 24 h of incubation until reliable identification to the species level was achieved (score ≥2.0). Mean incubation time needed to achieve species-level identification was 5.9 and 2.0 h for Gram-positive aerobic cocci (GPC, n = 86) and Gram-negative aerobic rods (GNR, n = 42), respectively. Short agar cultures with incubation times ≤2, ≤4, ≤6, ≤8 and ≤12 h yielded species identification in 1.2%, 18.6%, 64.0%, 96.5%, 98.8% of GPC, and in 76.2%, 95.2%, 97.6%, 97.6%, 97.6% of GNR, respectively. Control species identification at 24 h was achieved in 100% of GPC and 97.6% of GNR. Ethanol/formic acid protein extraction performed for an additional 34 GPC isolates cultivated from positive BCs showed further reduction in time to species identification (3.1 h). MALDI-TOF MS using biomass subsequent to very short-term incubation on solid medium allows very early and reliable bacterial identification from positive BCs without additional time and cost expenditure. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

Centrifugation after irradiation of red blood cells does not accelerate haemolysis.

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Weiss, Dominik R; Goehring, Jasmin; Weisbach, Volker; Strasser, Erwin F; Ringwald, Juergen; Zimmermann, Robert; Eckstein, Reinhold

2011-01-01

For intrauterine transfusion and some other rare indications, irradiation and washing or adjustment to an elevated haematocrit is necessary. No data are currently available indicating whether irradiation of red blood cell concentrates (RBCs) might impair the mechanical stability of erythrocytes during centrifugation leading to elevated haemolysis. Consequently, if irradiation and centrifugation of RBCs is necessary, there is no definitive recommendation about the preferred sequence of steps. We divided 20 RBC units that were not older than 9 days into two subunits. These subunits were prepared to yield irradiated RBCs with an elevated haematocrit, as they are used for intrauterine transfusion. One subunit was centrifuged and then irradiated, the other subunit was irradiated and then centrifuged. The units were evaluated in vitro before preparation and on days 1 and 7. We could not find any difference in the haemolysis rate, extracellular LDH or alpha-HBDH between the two groups of RBCs. This observation indicates that centrifugation after irradiation of RBCs does not accelerate haemolysis. A similar ATP content in the two subunits demonstrated no difference in energy metabolism. The extracellular potassium concentration was significantly lower in the subunits washed after irradiation. There is no difference in the haemolysis caused by centrifugation between irradiated and non-irradiated RBCs. However, it is well known that washing RBCs after irradiation significantly lowers the potassium content. Summarising these two findings leads to the conclusion that it is optimal first to irradiate and then to wash RBCs.

Molecular dosimetry of DNA damage caused by alkylation. I. Single-strand breaks induced by ethylating agents in cultured mammalian cells in relation to survival

NARCIS (Netherlands)

Abbondandolo, A.; Dogliotti, E.; Lohman, P.H.M.; Berends, F.

1982-01-01

Cultured Chinese hamster ovary cells were treated with ethylating agents. DNA lesions giving rise to single-strand breaks (ssb) or alkali-labile sites were measured by centrifugation in alkaline sucrose gradients after lysis in alkali. 4 agents with different tendencies to ethylate preferentially

In vivo evaluation of centrifugal blood pump for cardiopulmonary bypass-Spiral Pump.

Science.gov (United States)

da Silva, Cibele; da Silva, Bruno Utiyama; Leme, Juliana; Uebelhart, Beatriz; Dinkhuysen, Jarbas; Biscegli, José F; Andrade, Aron; Zavaglia, Cecília

2013-11-01

The Spiral Pump (SP), a centrifugal blood pump for cardiopulmonary bypass (CPB), has been developed at the Dante Pazzanese Institute of Cardiology/Adib Jatene Foundation laboratories, with support from Sintegra Company (Pompeia, Brazil). The SP is a disposable pump with an internal rotor-a conically shaped fuse with double entrance threads. This rotor is supported by two ball bearings, attached to a stainless steel shaft fixed to the housing base. Worm gears provide axial motion to the blood column, and the rotational motion of the conically shaped impeller generates a centrifugal pumping effect, improving pump efficiency without increasing hemolysis. In vitro tests were performed to evaluate the SP's hydrodynamic performance, and in vivo experiments were performed to evaluate hemodynamic impact during usual CPB. A commercially available centrifugal blood pump was used as reference. In vivo experiments were conducted in six male pigs weighing between 60 and 90 kg, placed on CPB for 6 h each. Blood samples were collected just before CPB (T0) and after every hour of CPB (T1-T6) for hemolysis determination and laboratory tests (hematological and biochemical). Values of blood pressure, mean flow, pump rotational speed, and corporeal temperature were recorded. Also, ergonomic conditions were recorded: presence of noise, difficulty in removing air bubbles, trouble in installing the pump in the drive module (console), and difficulties in mounting the CPB circuit. Comparing the laboratory and hemolysis results for the SP with those of the reference pump, we can conclude that there is no significant difference between the two devices. In addition, reports made by medical staff and perfusionists described a close similarity between the two devices. During in vivo experiments, the SP maintained blood flow and pressure at physiological levels, consistent with those applied in cardiac surgery with CPB, without presenting any malfunction. Also, the SP needed lower rotational

Two-Dimensional Computational Flow Analysis and Frictional Characteristics Model for Red Blood Cell under Inclined Centrifuge Microscopy

Science.gov (United States)

Funamoto, Kenichi; Hayase, Toshiyuki; Shirai, Atsushi

Simplified two-dimensional flow analysis is performed in order to simulate frictional characteristics measurement of red blood cells moving on a glass plate in a medium with an inclined centrifuge microscope. Computation under various conditions reveals the influences of parameters on lift, drag, and moment acting on a red blood cell. Among these forces, lift appears only when the cell is longitudinally asymmetric. By considering the balance of forces, the frictional characteristics of the red blood cell are modeled as the sum of Coulomb friction and viscous drag. The model describes the possibility that the red blood cell deforms to expand in the front side in response to the inclined centrifugal force. When velocity exceeds some critical value, the lift overcomes the normal centrifugal force component, and the thickness of the plasma layer between the cell and the glass plate increases from the initial value of the plasma protein thickness.

CFD simulation of centrifugal cells washers.

Science.gov (United States)

Kellet, Beth E; Binbing, Han; Dandy, David S; Wickramasinghe, S Ranil

2004-01-01

The feasibility of using computational fluid dynamics to guide the design of better centrifuges for processing shed blood is explored here. The velocity field and the rate of protein removal from the shed blood have been studied. The results indicate that computational fluid dynamics could help screen preliminary centrifuge bowl designs thus reducing the number of initial experimental tests required when developing new centrifuge bowls. Though the focus of this work is on washing shed blood the methods developed here are applicable to the design of centrifuge bowls for other blood processing applications.

Measurement of endotoxin levels in blood of hemodialysis Patients by 'Lal' test and comparision of its efficacy with blood culture

Directory of Open Access Journals (Sweden)

Gh Vazirzadeh

2006-01-01

Full Text Available Introduction: Presently, bacteremia is the principal cause of morbidity in patients undergoing hemodialysis. Gram-negative bacteria account for approximately 50 percent of documented infections. Endotoxins released during lysis of gram negative bacteremia result in inflammatory and defense response by the body and if not treated promptly result in septic shock and ultimately death of the patient. This study describes the detection of endotoxins in blood of patients with bacteremia due to gram - negative bacteria by LAL test. Method: Blood samples of 278 hemodialysis patients were analyzed in this study and pathogens were isolated from blood culture samples. Then, their antibiotic sensitivity was determined. In patients with positive blood culture, endotoxin levels were measured by LAL-test. Results: Frequency of bacteremia in patients was 13.6% . The prevalence of gram – negative bacteremia was 44.7%. E coli were the major pathogens, while staphylococcus aureus was the most common gram positive bacterium. Endotoxin was detected in 15 patients (3.8 ± 1.08 EU/ml . The sensitivity and specificity of endotoxins for gram – negative bacteremia were 88% and 95%, respectively. Conclusion: The results indicate that the LAL method is a fast, sensitive and simple method. There was no significant difference between the results of blood culture and LAL – test ( P > 0.05 .

A novel method of selective removal of human DNA improves PCR sensitivity for detection of Salmonella Typhi in blood samples.

Science.gov (United States)

Zhou, Liqing; Pollard, Andrew J

2012-07-27

Enteric fever is a major public health problem, causing an estimated 21million new cases and 216,000 or more deaths every year. Current diagnosis of the disease is inadequate. Blood culture only identifies 45 to 70% of the cases and is time-consuming. Serological tests have very low sensitivity and specificity. Clinical samples obtained for diagnosis of enteric fever in the field generally have blood, so that even PCR-based methods, widely used for detection of other infectious diseases, are not a straightforward option in typhoid diagnosis. We developed a novel method to enrich target bacterial DNA by selective removal of human DNA from blood samples, enhancing the sensitivity of PCR tests. This method offers the possibility of improving PCR assays directly using clinical specimens for diagnosis of this globally important infectious disease. Blood samples were mixed with ox bile for selective lysis of human blood cells and the released human DNA was then digested with addition of bile resistant micrococcal nuclease. The intact Salmonella Typhi bacteria were collected from the specimen by centrifugation and the DNA extracted with QIAamp DNA mini kit. The presence of Salmonella Typhi bacteria in blood samples was detected by PCR with the fliC-d gene of Salmonella Typhi as the target. Micrococcal nuclease retained activity against human blood DNA in the presence of up to 9% ox bile. Background human DNA was dramatically removed from blood samples through the use of ox bile lysis and micrococcal nuclease for removal of mammalian DNA. Consequently target Salmonella Typhi DNA was enriched in DNA preparations and the PCR sensitivity for detection of Salmonella Typhi in spiked blood samples was enhanced by 1,000 fold. Use of a combination of selective ox-bile blood cell lysis and removal of human DNA with micrococcal nuclease significantly improves PCR sensitivity and offers a better option for improved typhoid PCR assays directly using clinical specimens in diagnosis of

Acute hydrodynamic damage induced by SPLITT fractionation and centrifugation in red blood cells.

Science.gov (United States)

Urbina, Adriana; Godoy-Silva, Ruben; Hoyos, Mauricio; Camacho, Marcela

2016-05-01

Though blood bank processing traditionally employs centrifugation, new separation techniques may be appealing for large scale processes. Split-flow fractionation (SPLITT) is a family of techniques that separates in absence of labelling and uses very low flow rates and force fields, and is therefore expected to minimize cell damage. However, the hydrodynamic stress and possible consequent damaging effects of SPLITT fractionation have not been yet examined. The aim of this study was to investigate the hydrodynamic damage of SPLITT fractionation to human red blood cells, and to compare these effects with those induced by centrifugation. Peripheral whole blood samples were collected from healthy volunteers. Samples were diluted in a buffered saline solution, and were exposed to SPLITT fractionation (flow rates 1-10 ml/min) or centrifugation (100-1500 g) for 10 min. Cell viability, shape, diameter, mean corpuscular hemoglobin, and membrane potential were measured. Under the operating conditions employed, both SPLITT and centrifugation maintained cell viability above 98%, but resulted in significant sublethal damage, including echinocyte formation, decreased cell diameter, decreased mean corpuscular hemoglobin, and membrane hyperpolarization which was inhibited by EGTA. Wall shear stress and maximum energy dissipation rate showed significant correlation with lethal and sublethal damage. Our data do not support the assumption that SPLITT fractionation induces very low shear stress and is innocuous to cell function. Some changes in SPLITT channel design are suggested to minimize cell damage. Measurement of membrane potential and cell diameter could provide a new, reliable and convenient basis for evaluation of hydrodynamic effects on different cell models, allowing identification of optimal operating conditions on different scales. Copyright © 2016 Elsevier B.V. All rights reserved.

EVAHEART: an implantable centrifugal blood pump for long-term circulatory support.

Science.gov (United States)

Yamazaki, Kenji; Kihara, Shinichiro; Akimoto, Takehide; Tagusari, Osamu; Kawai, Akihiko; Umezu, Mitsuo; Tomioka, Jun; Kormos, Robert L; Griffith, Bartley P; Kurosawa, Hiromi

2002-11-01

We developed "EVAHEART": a compact centrifugal blood pump system as an implantable left ventricular assist device for long-term circulatory support. The 55 x 64 mm pump is made from pure titanium, and weighs 370 g. The entire blood-contacting surface is covered with an anti-thrombogenic coating of diamond like carbon (DLC) or 2-methacryloyloxyethyl phosphorylcholine (MPC) to improve blood compatibility. Flows exceeding 12 L/min against 100 mmHg pressure at 2600 rpm was measured. A low-temperature mechanical seal with recirculating cooling system is used to seal the shaft. EVAHEART demonstrated an acceptably low hemolysis rate with normalized index of hemolysis of 0.005 +/- 0.002 g/100L. We evaluated the pump in long-term in-vivo experiments with seven calves. Via left thoracotomy, we conducted left ventricular apex-descending aorta bypass, placing the pump in the left thoracic cavity. Pump flow rates was maintained at 5-9 L/min, pump power consumption remained stable at 9-10 W in all cases, plasma free Hb levels were less than 15 mg/dl, and the seal system showed good seal capability throughout the experiments. The calves were sacrificed on schedule on postoperative day 200, 222, 142, 90, 151, 155, and 133. No thrombi formed on the blood contacting surface with either the DLC or MPC coating, and no major organ thromboembolisms occurred except for a few small renal infarcts. EVAHEART centrifugal blood pump demonstrated excellent performance in long-term in-vivo experiments.

Centrifugation-free washing: A novel approach for removing immunoglobulin A from stored red blood cells.

Science.gov (United States)

Vörös, Eszter; Piety, Nathaniel Z; Strachan, Briony C; Lu, Madeleine; Shevkoplyas, Sergey S

2018-08-01

Washed red blood cells (RBCs) are indicated for immunoglobulin A (IgA) deficient recipients. Centrifugation-based cell processors commonly used by hospital blood banks cannot consistently reduce IgA below the recommended levels, hence double washing is frequently required. Here, we describe a prototype of a simple, portable, disposable system capable of washing stored RBCs without centrifugation, while reducing IgA below 0.05 mg/dL in a single run. Samples from RBC units (n = 8, leukoreduced, 4-6 weeks storage duration) were diluted with normal saline to a hematocrit of 10%, and then washed using either the prototype washing system, or via conventional centrifugation. The efficiency of the two washing methods was quantified and compared by measuring several key in vitro quality metrics. The prototype of the washing system was able to process stored RBCs at a rate of 300 mL/hour, producing a suspension of washed RBCs with 43 ± 3% hematocrit and 86 ± 7% cell recovery. Overall, the two washing methods performed similarly for most measured parameters, lowering the concentration of free hemoglobin by >4-fold and total free protein by >10-fold. Importantly, the new washing system reduced the IgA level to 0.02 ± 0.01 mg/mL, a concentration 5-fold lower than that produced by conventional centrifugation. This proof-of-concept study showed that centrifugation may be unnecessary for washing stored RBCs. A simple, disposable, centrifugation-free washing system could be particularly useful in smaller medical facilities and resource limited settings that may lack access to centrifugation-based cell processors. © 2017 Wiley Periodicals, Inc.

Impact of Ficoll density gradient centrifugation on major and trace element concentrations in erythrocytes and blood plasma.

Science.gov (United States)

Lu, Ying; Ahmed, Sultan; Harari, Florencia; Vahter, Marie

2015-01-01

Ficoll density gradient centrifugation is widely used to separate cellular components of human blood. We evaluated the suitability to use erythrocytes and blood plasma obtained from Ficoll centrifugation for assessment of elemental concentrations. We determined 22 elements (from Li to U) in erythrocytes and blood plasma separated by direct or Ficoll density gradient centrifugation, using inductively coupled plasma mass spectrometry. Compared with erythrocytes and blood plasma separated by direct centrifugation, those separated by Ficoll had highly elevated iodine and Ba concentration, due to the contamination from the Ficoll-Paque medium, and about twice as high concentrations of Sr and Mo in erythrocytes. On the other hand, the concentrations of Ca in erythrocytes and plasma were markedly reduced by the Ficoll separation, to some extent also Li, Co, Cu, and U. The reduced concentrations were probably due to EDTA, a chelator present in the Ficoll medium. Arsenic concentrations seemed to be lowered by Ficoll, probably in a species-specific manner. The concentrations of Mg, P, S, K, Fe, Zn, Se, Rb, and Cs were not affected in the erythrocytes, but decreased in plasma. Concentrations of Mn, Cd, and Pb were not affected in erythrocytes, but in plasma affected by EDTA and/or pre-analytical contamination. Ficoll separation changed the concentrations of Li, Ca, Co, Cu, As, Mo, I, Ba, and U in erythrocytes and blood plasma, Sr in erythrocytes, and Mg, P, S, K, Fe, Zn, Se, Rb and Cs in blood plasma, to an extent that will invalidate evaluation of deficiencies or excess intakes. Copyright © 2014 Elsevier GmbH. All rights reserved.

[Hemolysis Performance Analysis of the Centrifugal Maglev Blood Pump].

Science.gov (United States)

Wang, Yiwen; Zhang, Fan; Fang, Yuan; Dong, Baichuan; Zhou, Liang

2016-05-01

In order to analyze and study the hemolysis performance of the centrifugal maglev blood pump, which was designed by ourselves, this paper built the mathematical model and computational fluid dynamics analyzed it using Fluent. Then we set up the in vitro hemolysis experiment platform, in case of the design condition, the content of free hemoglobin and hematocrit in plasma were measured in a certain time interval, and calculated the normalized index of hemolysis of the blood pump. The numerical simulation results show the internal static pressure distribution is smooth inside the pump, the wal shear stress inside the pump is less than 150 Pa. Therefore, the red blood cel damage and exposure time is independent. The normalized index of hemolysis is (0.002 9±0.000 7) mg/L, which is in accordance with human physiological requirement.

Modeling centrifugal cell washers using computational fluid dynamics.

Science.gov (United States)

Kellet, Beth E; Han, Binbing; Dandy, David S; Wickramasinghe, S Ranil

2004-11-01

Reinfusion of shed blood during surgery could avoid the need for blood transfusions. Prior to reinfusion of the red blood cells, the shed blood must be washed in order to remove leukocytes, platelets, and other contaminants. Further, the hematocrit of the washed blood must be increased. The feasibility of using computational fluid dynamics (CFD) to guide the design of better centrifuges for processing shed blood is explored here. The velocity field within a centrifuge bowl and the rate of protein removal from the shed blood has been studied. The results obtained indicate that CFD could help screen preliminary centrifuge bowl designs, thus reducing the number of initial experimental tests required when developing new centrifuge bowls. Although the focus of this work is on washing shed blood, the methods developed here are applicable to the design of centrifuge bowls for other blood-processing applications.

Method and apparatus for automated processing and aliquoting of whole blood samples for analysis in a centrifugal fast analyzer

Science.gov (United States)

Burtis, C.A.; Johnson, W.F.; Walker, W.A.

1985-08-05

A rotor and disc assembly for use in a centrifugal fast analyzer. The assembly is designed to process multiple samples of whole blood followed by aliquoting of the resultant serum into precisely measured samples for subsequent chemical analysis. The assembly requires minimal operator involvement with no mechanical pipetting. The system comprises: (1) a whole blood sample disc; (2) a serum sample disc; (3) a sample preparation rotor; and (4) an analytical rotor. The blood sample disc and serum sample disc are designed with a plurality of precision bore capillary tubes arranged in a spoked array. Samples of blood are loaded into the blood sample disc by capillary action and centrifugally discharged into cavities of the sample preparation rotor where separation of serum and solids is accomplished. The serum is loaded into the capillaries of the serum sample disc by capillary action and subsequently centrifugally expelled into cuvettes of the analyticaly rotor for conventional methods. 5 figs.

Method and apparatus for automated processing and aliquoting of whole blood samples for analysis in a centrifugal fast analyzer

Science.gov (United States)

Burtis, Carl A.; Johnson, Wayne F.; Walker, William A.

1988-01-01

A rotor and disc assembly for use in a centrifugal fast analyzer. The assembly is designed to process multiple samples of whole blood followed by aliquoting of the resultant serum into precisely measured samples for subsequent chemical analysis. The assembly requires minimal operator involvement with no mechanical pipetting. The system comprises (1) a whole blood sample disc, (2) a serum sample disc, (3) a sample preparation rotor, and (4) an analytical rotor. The blood sample disc and serum sample disc are designed with a plurality of precision bore capillary tubes arranged in a spoked array. Samples of blood are loaded into the blood sample disc in capillary tubes filled by capillary action and centrifugally discharged into cavities of the sample preparation rotor where separation of serum and solids is accomplished. The serum is loaded into the capillaries of the serum sample disc by capillary action and subsequently centrifugally expelled into cuvettes of the analytical rotor for analysis by conventional methods.

Modeling industrial centrifugation of mammalian cell culture using a capillary based scale-down system.

Science.gov (United States)

Westoby, Matthew; Rogers, Jameson K; Haverstock, Ryan; Romero, Jonathan; Pieracci, John

2011-05-01

Continuous-flow centrifugation is widely utilized as the primary clarification step in the recovery of biopharmaceuticals from cell culture. However, it is a challenging operation to develop and characterize due to the lack of easy to use, small-scale, systems that can be used to model industrial processes. As a result, pilot-scale continuous centrifugation is typically employed to model large-scale systems requiring a significant amount of resources. In an effort to reduce resource requirements and create a system which is easy to construct and utilize, a capillary shear device, capable of producing energy dissipation rates equivalent to those present in the feed zones of industrial disk stack centrifuges, was developed and evaluated. When coupled to a bench-top, batch centrifuge, the capillary device reduced centrate turbidity prediction error from 37% to 4% compared to using a bench-top centrifuge alone. Laboratory-scale parameters that are analogous to those routinely varied during industrial-scale continuous centrifugation were identified and evaluated for their utility in emulating disk stack centrifuge performance. The resulting relationships enable bench-scale process modeling of continuous disk stack centrifuges using an easily constructed, scalable, capillary shear device coupled to a typical bench-top centrifuge. Copyright © 2010 Wiley Periodicals, Inc.

Simple tube centrifugation for processing platelet-rich plasma in the horse.

Science.gov (United States)

Fontenot, Robin L; Sink, Carolyn A; Werre, Stephen R; Weinstein, Nicole M; Dahlgren, Linda A

2012-12-01

This study evaluated the quality and bacteriologic safety of platelet-rich plasma (PRP) produced by 3 simple, inexpensive tube centrifugation methods and a commercial system. Citrated equine blood collected from 26 normal horses was processed by 4 methods: blood collection tubes centrifuged at 1200 and 2000 × g, 50-mL conical tube, and a commercial system. White blood cell (WBC), red blood cell (RBC), and platelet counts and mean platelet volume (MPV) were determined for whole blood and PRP, and aerobic and anaerobic cultures were performed. Mean platelet concentrations ranged from 1.55- to 2.58-fold. The conical method yielded the most samples with platelet concentrations greater than 2.5-fold and within the clinically acceptable range of > 250,000 platelets/μL. White blood cell counts were lowest with the commercial system and unacceptably high with the blood collection tubes. The conical tube method may offer an economically feasible and comparatively safe alternative to commercial PRP production systems.

Centrifugation: an important pre-analytic procedure that influences plasma microRNA quantification during blood processing.

Science.gov (United States)

Zheng, Xiao-Hui; Cui, Cui; Zhou, Xin-Xi; Zeng, Yi-Xin; Jia, Wei-Hua

2013-12-01

Circulating microRNAs are robustly present in plasma or serum and have become a research focus as biomarkers for tumor diagnosis and prognosis. Centrifugation is a necessary procedure for obtaining high-quality blood supernatant. Herein, we investigated one-step and two-step centrifugations, two centrifugal methods routinely used in microRNA study, to explore their effects on plasma microRNA quantification. The microRNAs obtained from one-step and two-step centrifugations were quantified by microarray and TaqMan-based real-time quantitative polymerase chain reaction (Q-PCR). Dynamic light scattering was performed to explore the difference underlying the two centrifugal methods. The results from the microarray containing 1,347 microRNAs showed that the signal detection rate was greatly decreased in the plasma sample prepared by two-step centrifugation. More importantly, the microRNAs missing in this plasma sample could be recovered and detected in the precipitate generated from the second centrifugation. Consistent with the results from microarray, a marked decrease of three representative microRNAs in two-step centrifugal plasma was validated by Q-PCR. According to the size distribution of all nanoparticles in plasma, there were fewer nanoparticles with size >1,000 nm in two-step centrifugal plasma. Our experiments directly demonstrated that different centrifugation methods produced distinct quantities of plasma microRNAs. Thus, exosomes or protein complexes containing microRNAs may be involved in large nanoparticle formation and may be precipitated after two-step centrifugation. Our results remind us that sample processing methods should be first considered in conducting research.

Effect of centrifuge test on blood serum lipids index of cadet pilots.

Science.gov (United States)

Wochyński, Zbigniew; Kowalczuk, Krzysztof; Kłossowski, Marek; Sobiech, Krzysztof A

2016-01-01

This study aimed at investigating the relationship between the lipid index (WS) in the examined cadets and duration of exposure to +Gz in the human centrifuge. The study involved 19 first-year cadets of the Polish Air Force Academy in Dęblin. Tests in the human centrifuge were repeated twice, i.e. prior to (test I) and 45 days after (test II). After exposure to +Gz, the examined cadets were divided into 2 groups. Group I (N=11) included cadets subjected to a shorter total duration of exposure to +Gz, while group II (N=8) included cadets with a longer total duration of exposure to +Gz. Total cholesterol (TC), high density lipoprotein (HDL), triglycerides (TG), and apolipoproteins A1 and B were assayed in blood serum prior to (assay A) and after (assay B) both exposures to +Gz. Low density lipoprotein (LDL) level was estimated from the Friedewald formula. WS is an own mathematical algorithm. WS was higher in group II, assay A - 10.0 and B - 10.08 of test I in the human centrifuge than in group I where the WS values were 6.91 and 6.96, respectively. WS was also higher in group II in assay A - 10.0 and B -10.1 of test II in the human centrifuge than in group I - 6.96 and 6.80, respectively. The higher value of WS in group II, both after the first and second exposure to +Gz in human centrifuge, in comparison with group I, indicated its usefulness for determination of the maximum capability of applying acceleration of the interval type during training in the human centrifuge.

Rapid kinetics of lysis in human natural cell-mediated cytotoxicity: some implications

International Nuclear Information System (INIS)

Bloom, E.T.; Babbitt, J.T.

1983-01-01

The entire lytic process of natural cell-mediated cytotoxicity against sensitive target cells can occur rapidly, within minutes. This was demonstrated by 51 chromium release and in single-cell assays. At the cellular level, most of the target cell lysis occurred within 15-30 min after binding to effector cells. The enriched natural killer cell subpopulation of lymphocytes obtained by Percoll density gradient centrifugation (containing greater than 70% large granular lymphocytes (LGL)) was the most rapidly lytic population by 51 chromium release. However, in the single-cell assay, the rate of lysis of bound target cells was quite similar for the LGL-enriched effector subpopulation and the higher density subpopulation of effector cells recognized previously. Both the light and dense effector cells contained similar numbers of target binding cells. Therefore, that the light subpopulation effected lysis more rapidly and to a greater extent than the dense subpopulation suggested that the low-density effector cells probably recycled more rapidly than those of higher density. This was corroborated by the finding that when conjugates were formed at 29 degrees C for the single-cell assay, a significant number of dead unconjugated targets could be observed only on the slides made with the LGL-enriched effector cells but not on those made with dense effector cell. Lysis continued to increase in the chromium-release assay probably because of recycling, recruitment, and/or heterogeneity of the effector cells, and/or because of heterogeneity or delayed death of the target cells

Residual blood processing by centrifugation, cell salvage or ultrafiltration in cardiac surgery: effects on clinical hemostatic and ex-vivo rheological parameters.

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Vonk, Alexander B; Muntajit, Warayouth; Bhagirath, Pranav; van Barneveld, Laurentius J; Romijn, Johannes W; de Vroege, Roel; Boer, Christa

2012-10-01

The study compared the effects of three blood concentration techniques after cardiopulmonary bypass on clinical hemostatic and ex-vivo rheological parameters. Residual blood of patients undergoing elective cardiac surgery was processed by centrifugation, cell salvage or ultrafiltration, and retransfused (n = 17 per group). Study parameters included blood loss, (free) hemoglobin, hematocrit, fibrinogen and erythrocyte aggregation, deformability and 2,3-diphosphoglycerate content. Patient characteristics were similar between groups. Ultrafiltration was associated with the highest weight of the transfusion bag [649 ± 261 vs. 320 ± 134 g (centrifugation) and 391 ± 158 g (cell salvage); P levels in the transfusion bag. Retransfusion of cell saver blood induced the largest gain in postoperative patient hemoglobin levels when compared to centrifugation and ultrafiltration, and was associated with the largest increase in 2,3-diphosphoglycerate when compared to ultrafiltration (Δ2,3-diphosphoglycerate 1.34 ± 1.92 vs. -0.77 ± 1.56 mmol/l; P = 0.03). Cell salvage is superior with respect to postoperative hemoglobin gain and washout of free hemoglobin when compared to centrifugation or ultrafiltration.

The effect of centrifugation at various g force levels on rheological properties of rat, dog, pig and human red blood cells.

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Kiss, Ferenc; Toth, Eniko; Miszti-Blasius, Kornel; Nemeth, Norbert

2016-01-01

Laboratory investigations often require centrifugation of blood samples for various erythrocyte tests. Although there is a lack of data about the effect of centrifugation at various g force levels on erythrocyte rheological properties. We aimed to investigate the effect of a 10-minute centrifugation at 500, 1000 or 1500 g at 15°C of rat, dog, pig and human venous (K3-EDTA, 1.5 mg/ml) blood samples. Hematological parameters, erythrocyte deformability, cell membrane stability, osmotic gradient ektacytometry (osmoscan) and erythrocyte aggregation were determined. Hematological and erythrocyte deformability parameters showed interspecies differences, centrifugation caused no significant alterations. Cell membrane stability for human erythrocytes centrifuged at higher g level showed less decrease in deformability. Osmoscan O min parameter showed slight elevation in dog centrifuged aliquots. Erythrocyte aggregation parameters changed unexpectedly. Rat and dog erythrocyte aggregation indices significantly dropped in centrifuged aliquots. Pig erythrocyte aggregation indices increased significantly after centrifugation. Human erythrocyte aggregation was the most stable one among the investigated species. The used centrifugation protocols caused the largest alterations in erythrocyte aggregation in a controversial way among the investigated species. On the other hand, erythrocyte deformability parameters were stable, cell membrane stability and osmoscan data show minor shifts.

Directly observed reversible shape changes and hemoglobin stratification during centrifugation of human and Amphiuma red blood cells.

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Hoffman, Joseph F; Inoué, Shinya

2006-02-21

This paper describes changes that occur in human and Amphiuma red blood cells observed during centrifugation with a special microscope. Dilute suspensions of cells were layered, in a centrifuge chamber, above an osmotically matched dense solution, containing Nycodenz, Ficoll, or Percoll (Pharmacia) that formed a density gradient that allowed the cells to slowly settle to an equilibrium position. Biconcave human red blood cells moved downward at low forces with minimum wobble. The cells oriented vertically when the force field was increased and Hb sedimented as the lower part of each cell became bulged and assumed a "bag-like" shape. The upper centripetal portion of the cell became thinner and remained biconcave. These changes occurred rapidly and were completely reversible upon lowering the centrifugal force. Bag-shaped cells, upon touching red cells in rouleau, immediately reverted to biconcave disks as they flipped onto a stack. Amphiuma red cells displayed a different type of reversible stratification and deformation at high force fields. Here the cells became stretched, with the nucleus now moving centrifugally, the Hb moving centripetally, and the bottom of the cells becoming thinner and clear. Nevertheless, the distribution of the marginal bands at the cells' rim was unchanged. We conclude that centrifugation, per se, while changing a red cell's shape and the distribution of its intracellular constituents, does so in a completely reversible manner. Centrifugation of red cells harboring altered or missing structural elements could provide information on shape determinants that are still unexplained.

A fast and highly sensitive blood culture PCR method for clinical detection of Salmonella enterica serovar Typhi

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Zhou Liqing

2010-04-01

Full Text Available Abstract Background Salmonella Typhi causes an estimated 21 million new cases of typhoid fever and 216,000 deaths every year. Blood culture is currently the gold standard for diagnosis of typhoid fever, but it is time-consuming and takes several days for isolation and identification of causative organisms. It is then too late to initiate proper antibiotic therapy. Serological tests have very low sensitivity and specificity, and no practical value in endemic areas. As early diagnosis of the disease and prompt treatment are essential for optimal management, especially in children, a rapid sensitive detection method for typhoid fever is urgently needed. Although PCR is sensitive and rapid, initial research indicated similar sensitivity to blood culture and lower specificity. We developed a fast and highly sensitive blood culture PCR method for detection of Salmonella Typhi, allowing same-day initiation of treatment after accurate diagnosis of typhoid. Methods An ox bile tryptone soy broth was optimized for blood culture, which allows the complete lysis of blood cells to release intracellular bacteria without inhibiting the growth of Salmonella Typhi. Using the optimised broth Salmonella Typhi bacteria in artificial blood samples were enriched in blood culture and then detected by a PCR targeting the fliC-d gene of Salmonella Typhi. Results Tests demonstrated that 2.4% ox bile in blood culture not only lyzes blood cells completely within 1.5 hours so that the intracellular bacteria could be released, but also has no inhibiting effect on the growth of Salmonella Typhi. Three hour enrichment of Salmonella Typhi in tryptone soya broth containing 2.4% ox bile could increase the bacterial number from 0.75 CFU per millilitre of blood which is similar to clinical typhoid samples to the level which regular PCR can detect. The whole blood culture PCR assay takes less than 8 hours to complete rather than several days for conventional blood culture

Direct Identification and Antimicrobial Susceptibility Testing of Bacteria From Positive Blood Culture Bottles by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry and the Vitek 2 System.

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Jo, Sung Jin; Park, Kang Gyun; Han, Kyungja; Park, Dong Jin; Park, Yeon-Joon

2016-03-01

We evaluated the reliability and accuracy of the combined use of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) bacterial identification and Vitek 2 antimicrobial susceptibility testing (AST) for bacteria from positive blood culture bottles. Direct identification and AST were performed in parallel to the standard methods in monomicrobial positive blood culture bottles. In total, 254 isolates grown on aerobic and/or anaerobic bottles were identified with MALDI-TOF Vitek MS (bioMérieux, France), and 1,978 microorganism/antimicrobial agent combinations were assessed. For isolates from anaerobic bottles, an aliquot of the culture broth was centrifuged, washed, and filtered through a nylon mesh. For isolates from aerobic/pediatric bottles, a lysis step using 9.26% ammonium chloride solution and 2% saponin solution was included. The overall correct identification rate was 81.8% (208/254) and that for gram-positive/gram-negative isolates was 73.9%/92.6%, respectively, and it was 81.8%, 87.6%, and 57.9% for isolates from aerobic, anaerobic, and pediatric bottles, respectively. Identification was not possible in 45 cases, and most of these isolates were streptococci (N=14) and coagulase-negative staphylococci (N=11). Misidentification occurred only in one case. Compared with standard methods, direct AST showed 97.9% (1,936/1,978) agreement with very major error of 0.25%, major error of 0.05%, and minor error of 1.8%. This simple and cost-effective sample preparation method gives reliable results for the direct identification and AST of bacteria. For the identification of streptococci and coagulase-negative staphylococci, the method should be further improved.

[Pre-analytical stability before centrifugation of 7 biochemical analytes in whole blood].

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Perrier-Cornet, Andreas; Moineau, Marie-Pierre; Narbonne, Valérie; Plee-Gautier, Emmanuelle; Le Saos, Fabienne; Carre, Jean-Luc

2015-01-01

The pre-analytical stability of 7 biochemical parameters (parathyroid hormone -PTH-, vitamins A, C E and D, 1,25-dihydroxyvitamin D and insulin) at +4 °C, was studied on whole blood samples before centrifugation. The impact of freezing at -20°C was also analyzed/performed for PTH and vitamin D. The differences in the results of assays for whole blood samples, being kept for different times between sampling time and analysis, from 9 healthy adults, were compaired by using a Student t test. The 7 analytes investigated remained stable up to 4 hours at +4°C in whole blood. This study showed that it is possible to accept uncentrifuged whole blood specimens kept at +4°C before analysis. PTH is affected by freezing whereas vitamin D is not.

Quantification of platelets obtained by different centrifugation protocols in SHR rats

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João Alberto Yazigi Junior

2015-12-01

Full Text Available ABSTRACT OBJECTIVE: To quantify the platelet concentration in the blood of SHR rats, by means of different centrifugation protocols, and to evaluate what the most effective method for obtaining platelets is. METHODS: We used 40 male rats of the isogenic SHR lineage. The animals were divided into three groups: control, using whole blood without centrifugation; single centrifugation, using whole blood subjected to a single centrifugation at 200 × gand 400 × g; and double centrifugation, using whole blood subjected one centrifugation at different rotations, followed by collection of whole plasma subjected to another centrifugation at different rotations: 200 × g+ 200 ×g; 200 × g+ 400 × g; 200 × g+ 800 × g; 400 ×g+ 400 × g; 400 × g+ 800 × g. Samples of 3 ml of blood were drawn from each animal by means of cardiac puncture. The blood was stored in Vacutainer collection tubes containing 3.2% sodium citrate. The blood from the control group animals was analyzed without being subjected to centrifugation. After the blood from the other groups of animals had been subjected to centrifugation, the whole plasma was collected and subjected to platelet counting in the lower third of the sample. RESULTS: We obtained greatest platelet enrichment in the subgroup with two centrifugations comprising 400 × gfor 10 min + 400 ×gfor 10 min, in which the mean platelet concentration was 11.30 times higher than that of the control group. CONCLUSION: It was possible to obtain a high platelet concentration using viable simple techniques, by means of centrifugation of whole blood and use of commonly used materials. The most effective method for obtaining platelet concentrate was found in samples subjected to two centrifugations.

Centrifugal Separation Device Based on Two-Layer Laminar Flow in Microchannels for High-Throughput and Continuous Blood Cell/Plasma Separation

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Taizo Kobayashi,; Taisuke Funamoto,; Makoto Hosaka,; Satoshi Konishi,

2010-07-01

This paper presents a novel type of centrifugation device that is based on the two-layer laminar flow in micro flow channels for continuous blood cell/plasma separation. We propose to rotate the flow channels which are arranged along the circumference around the rotational axis. Downsizing the channel width reduced both the cell sedimentation time and the required centrifugal force, because the channel width corresponds to the centrifugal sedimentation length. First, plasma and cells were continuously extracted from pig blood in each of the branch channels using a milled acrylic prototype device (channel width = 800 μm, volume = 150 μl). Next, the relationship between the channel width (125, 250, and 500 μm) and the sedimentation time taken for various centrifugal forces (2.3, 9, 36, and 145 G) was evaluated using the downsized microchannels fabricated by hot-embossing and thermal bonding technologies. Using downsized microchannels with a width of 125 μm successfully reduced the sedimentation time to 85 s as compared to the sedimentation time of 270 s for a channel of a width of 500 μm, when a centrifugal force of 2.3 G was applied. The use of the proposed device did not result in obvious hemolysis at the centrifugal forces lower than 335 G.

Quantification of platelets obtained by different centrifugation protocols in SHR rats

OpenAIRE

João Alberto Yazigi Junior; João Baptista Gomes dos Santos; Bruno Rodrigues Xavier; Marcela Fernandes; Sandra Gomes Valente; Vilnei Mattiolli Leite

2015-01-01

ABSTRACT OBJECTIVE: To quantify the platelet concentration in the blood of SHR rats, by means of different centrifugation protocols, and to evaluate what the most effective method for obtaining platelets is. METHODS: We used 40 male rats of the isogenic SHR lineage. The animals were divided into three groups: control, using whole blood without centrifugation; single centrifugation, using whole blood subjected to a single centrifugation at 200 × gand 400 × g; and double centrifugation, usin...

Influences of centrifugation on cells and tissues in liposuction aspirates: optimized centrifugation for lipotransfer and cell isolation.

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Kurita, Masakazu; Matsumoto, Daisuke; Shigeura, Tomokuni; Sato, Katsujiro; Gonda, Koichi; Harii, Kiyonori; Yoshimura, Kotaro

2008-03-01

Although injective autologous fat transplantation is one of the most attractive options for soft-tissue augmentation, problems such as unpredictability and fibrosis resulting from fat necrosis limit its universal acceptance. Centrifugation is one of most common methods for overcoming these difficulties. This study was performed to investigate quantitatively the effects of centrifugation on liposuction aspirates to optimize centrifugal conditions for fat transplantation and isolation of adipose-derived stem cells. Liposuction aspirates, obtained from eight healthy female donors, were either not centrifuged or centrifuged at 400, 700, 1200, 3000, or 4200 g for 3 minutes. The volumes of the oil, adipose, and fluid portions and numbers of blood cells and adipose-derived cells in each portion were examined. The processed adipose tissues (1 ml) were injected into athymic mice, and grafts were harvested and weighed at 4 weeks. Morphologic alterations were observed using light and scanning electron microscopy. Centrifugation concentrated adipose tissues and adipose-derived stem cells in the adipose portion and partly removed red blood cells from the adipose portion. Centrifugation at more than 3000 g significantly damaged adipose-derived stem cells. Centrifugation enhanced graft take per 1 ml centrifuged adipose but reduced calculated graft take per 1 ml adipose before centrifugation. Excessive centrifugation can destroy adipocytes and adipose-derived stem cells, but appropriate centrifugation concentrates them, resulting in enhanced graft take. The authors tentatively recommend 1200 g as an optimized centrifugal force for obtaining good short- and long-term results in adipose transplantation.

Direct identification and susceptibility testing of positive blood cultures using high speed cold centrifugation and Vitek II system.

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Bazzi, Ali M; Rabaan, Ali A; Fawarah, Mahmoud M; Al-Tawfiq, Jaffar A

Compared to routine isolated colony-based methods, direct testing of bacterial pellets from positive blood cultures reduces turnaround time for reporting of antibiotic susceptibility. The aim of this study was to compare the accuracy, and precision, of a rapid method for direct identification and susceptibility testing of blood cultures with the routine method used in our laboratory, using Vitek 2. A total of 60 isolates were evaluated using the candidate and the routine method. The candidate method had 100% accuracy for the identification of Gram negative bacteria, Staphylococcus and Enterococcus, 50% for Streptococcus and 33.3% for Corynebacterium species. Susceptibility testing of Gram negative isolates yielded 98-100% essential agreement. For Staphylococcus and Enterococcus isolates, essential agreement was 100% for 17 antibiotics except for moxifloxacin. Direct testing of blood culture samples with Vitek 2 produced reliable identification and susceptibility results 18-24h sooner for aerobic/anaerobic facultative Gram-negative bacteria and Gram-positive Staphylococcus and Enterococcus strains. Copyright © 2016 King Saud Bin Abdulaziz University for Health Sciences. Published by Elsevier Ltd. All rights reserved.

Outcomes and biochemical parameters following cardiac surgery: effects of transfusion of residual blood using centrifugation and multiple-pass hemoconcentration.

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McNair, Erick; McKay, William; Qureshi, Abdul Mohamed; Rosin, Mark; Gamble, Jon; Dalshaug, Greg; Mycyk, Taras; Prasad, Kailash

2013-12-01

To determine whether or not there was a significant difference between the methods of centrifugation (CF) and multiple-pass hemoconcentration (MPH) of the residual cardiopulmonary-bypass volume in relation to biochemical measurements and patient outcomes. Prospective, randomized, and controlled. Conducted at a western Canadian tertiary care hospital. Consisted of 61 consecutive male and female patients from ages 40 to 80 who were scheduled for cardiac surgery with cardiopulmonary bypass. Either the centrifugation or multiple-pass hemoconcentration method was used to process the residual blood from the cardiopulmonary bypass circuit. The 12-hour postoperative levels of serum hemoglobin were not significantly different in the centrifugation group as compared to the multiple-pass hemoconcentration group. However, the serum levels of total protein and albumin were significantly higher in the multiple-pass hemoconcentration group as compared to the centrifugation group. Additionally, after 12-hours postoperatively, the serum fibrinogen and platelet counts were significantly higher in the multiple-pass hemoconcentration group as compared to those of the centrifugation group. The allogeneic product transfusion index and the chest-tube blood drainage indices were lower in the multiple-pass hemoconcentration group as compared to the centrifugation group. Although the CF method provided a product in a shorter turnaround time, with consistent clearance of heparin, the MPH method trended towards enhanced biochemical and clinical patient outcomes over the 12-hour postoperative period. Copyright © 2013 Elsevier Inc. All rights reserved.

Quantification of platelets obtained by different centrifugation protocols in SHR rats.

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Yazigi Junior, João Alberto; Dos Santos, João Baptista Gomes; Xavier, Bruno Rodrigues; Fernandes, Marcela; Valente, Sandra Gomes; Leite, Vilnei Mattiolli

2015-01-01

To quantify the platelet concentration in the blood of SHR rats, by means of different centrifugation protocols, and to evaluate what the most effective method for obtaining platelets is. We used 40 male rats of the isogenic SHR lineage. The animals were divided into three groups: control, using whole blood without centrifugation; single centrifugation, using whole blood subjected to a single centrifugation at 200 × g and 400 × g; and double centrifugation, using whole blood subjected one centrifugation at different rotations, followed by collection of whole plasma subjected to another centrifugation at different rotations: 200 × g + 200 × g; 200 × g + 400 × g; 200 × g + 800 × g; 400 × g + 400 × g; 400 × g + 800 × g. Samples of 3 ml of blood were drawn from each animal by means of cardiac puncture. The blood was stored in Vacutainer collection tubes containing 3.2% sodium citrate. The blood from the control group animals was analyzed without being subjected to centrifugation. After the blood from the other groups of animals had been subjected to centrifugation, the whole plasma was collected and subjected to platelet counting in the lower third of the sample. We obtained greatest platelet enrichment in the subgroup with two centrifugations comprising 400 × g for 10 min + 400 × g for 10 min, in which the mean platelet concentration was 11.30 times higher than that of the control group. It was possible to obtain a high platelet concentration using viable simple techniques, by means of centrifugation of whole blood and use of commonly used materials. The most effective method for obtaining platelet concentrate was found in samples subjected to two centrifugations.

The euglobulin clot lysis time to assess the impact of nanoparticles on fibrinolysis

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Minet, Valentine, E-mail: valentine.minet@unamur.be; Alpan, Lutfiye; Mullier, François [University of Namur – UNamur, Department of Pharmacy, Namur Thrombosis and Hemostasis Center (NTHC), Namur Nanosafety Center (NNC), NAmur Research Institute for Life Sciences NARILIS (Belgium); Toussaint, Olivier [Laboratory of Cellular Biochemistry and Biology (URBC) (Belgium); Lucas, Stéphane [University of Namur (UNamur), Research Centre for the Physics of Matter and Radiation (PMR-LARN), Namur Nanosafety Center NNC, NAmur Research Institute for Life Sciences NARILIS (Belgium); Dogné, Jean-Michel; Laloy, Julie, E-mail: julie.laloy@unamur.be [University of Namur – UNamur, Department of Pharmacy, Namur Thrombosis and Hemostasis Center (NTHC), Namur Nanosafety Center (NNC), NAmur Research Institute for Life Sciences NARILIS (Belgium)

2015-07-15

Nanoparticles (NPs) are developed for many applications in various fields, including nanomedicine. The NPs used in nanomedicine may disturb homeostasis in blood. Secondary hemostasis (blood coagulation) and fibrinolysis are complex physiological processes regulated by activators and inhibitors. An imbalance of this system can either lead to the development of hemorrhages or thrombosis. No data are currently available on the impact of NPs on fibrinolysis. The objectives of this study are (1) to select a screening test to study ex vivo the impact of NPs on fibrinolysis and (2) to test NPs with different physicochemical properties. Euglobulin clot lysis time test was selected to screen the impact of some NPs on fibrinolysis using normal pooled plasma. A dose-dependent decrease in the lysis time was observed with silicon dioxide and silver NPs without disturbing the fibrin network. Carbon black, silicon carbide, and copper oxide did not affect the lysis time at the tested concentrations.

Comparison of PCR-Based Diagnosis with Centrifuged-Based Enrichment Method for Detection of Borrelia persica in Animal Blood Samples.

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Naddaf, S R; Kishdehi, M; Siavashi, Mr

2011-01-01

The mainstay of diagnosis of relapsing fever (RF) is demonstration of the spirochetes in Giemsa-stained thick blood smears, but during non fever periods the bacteria are very scanty and rarely detected in blood smears by microscopy. This study is aimed to evaluate the sensitivity of different methods developed for detection of low-grade spirochetemia. Animal blood samples with low degrees of spirochetemia were tested with two PCRs and a nested PCR targeting flaB, GlpQ, and rrs genes. Also, a centrifuged-based enrichment method and Giemsa staining were performed on blood samples with various degrees of spirochetemia. The flaB-PCR and nested rrs-PCR turned positive with various degrees of spirochetemia including the blood samples that turned negative with dark-field microscopy. The GlpQ-PCR was positive as far as at least one spirochete was seen in 5-10 microscopic fields. The sensitivity of GlpQ-PCR increased when DNA from Buffy Coat Layer (BCL) was used as template. The centrifuged-based enrichment method turned positive with as low concentration as 50 bacteria/ml blood, while Giemsa thick staining detected bacteria with concentrations ≥ 25000 bacteria/ml. Centrifuged-based enrichment method appeared as much as 500-fold more sensitive than thick smears, which makes it even superior to some PCR assays. Due to simplicity and minimal laboratory requirements, this method can be considered a valuable tool for diagnosis of RF in rural health centers.

Biochemical assessment of growth factors and circulation of blood components contained in the different fractions obtained by centrifugation of venous blood.

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Corigiano, M; Ciobanu, G; Baldoni, E; Pompa, G

2014-01-01

The aim of this study was to evaluate a biochemical marker with different elements of a normal blood serum and centrifuged blood serum after a different rotation system. For this technique, we used five fractions of a blood Concentrated Growth Factors system (bCGF) and a particular device for the different rotation program. Blood samples were collected from 10 volunteers aged between 35 and 55 in the Operative Unit of the “Sapienza” University of Rome with only a fraction of different biochemical elements. Through an individual blood phase separator tube of venous blood, active factions of serum and 4 fractions of red buffy coat were taken. The biochemical markers with 14 elements were examined at times: P1-11 minutes, P2-12minutes, P3-15 minutes. Exclusively biological materials which are normally applied in the regeneration techniques for different defects and lesions were used with this technique. After specific rotation programs, a different result was obtained for each cycle: P1, P2, P3. In test tubes obtained by separated blood, we observed a higher concentration of proteins, ions, and other antigens compared to normal blood plasma. Examining the biochemical results of different elements, we observed an increase (P≤0,01). Since each person’s DNA is different, we could not have the same results in 5 fractions of blood concentration, we did, however, find a good increase in only a fraction of proteins, immunoglobulin and different ions. We obtained five fractions after centrifugation, and we had an increase in different biochemical elements compared to normal blood (P≤0,01) which is significant at different times. These biochemical elements were stimulated by different growth factors, which are used by the immune system, and they induced the formation of hard and soft tissues and good regeneration.

Removal of cellular-type hemoglobin-based oxygen carrier (hemoglobin-vesicles) from blood using centrifugation and ultrafiltration.

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Sakai, Hiromi; Sou, Keitaro; Horinouchi, Hirohisa; Tsuchida, Eishun; Kobayashi, Koichi

2012-02-01

The hemoglobin-vesicle (HbV) is an artificial oxygen carrier encapsulating a concentrated hemoglobin solution in a phospholipid vesicle (liposome). During or after transporting oxygen, macrophages capture HbVs in the reticuloendothelial system (RES) with an approximate circulation half-life of 3 days. Animal studies show transient splenohepatomegaly after large doses, but HbVs were completely degraded, and the components were excreted in a few weeks. If a blood substitute is used for emergency use until red blood cell transfusion becomes available or for temporary use such as a priming fluid for an extracorporeal circuit, then one option would be to remove HbVs from the circulating blood without waiting a few weeks for removal by the RES. Using a mixture of beagle dog whole blood and HbV, we tested the separation of HbV using a centrifugal Fresenius cell separator and an ultrafiltration system. The cell separator system separated the layers of blood cell components from the HbV-containing plasma layer by centrifugal force, and then the HbV was removed from plasma phase by the ultrafiltration system. The HbVs (250-280 nm) are larger than plasma proteins (blood cell components (> 3 µm). The size of HbVs is advantageous to be separated from the original blood components, and the separated blood components can be returned to circulation. © 2011, Copyright the Authors. Artificial Organs © 2011, International Center for Artificial Organs and Transplantation and Wiley Periodicals, Inc.

Rapid identification of bacteria in positive blood culture by matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

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Schmidt, V; Jarosch, A; März, P; Sander, C; Vacata, V; Kalka-Moll, W

2012-03-01

Blood culture is probably the most significant specimen used for the diagnosis of bacterial infections, especially for bloodstream infections. In the present study, we compared the resin-containing BD BACTEC™ Plus-Aerobic (Becton Dickinson), non-charcoal-containing BacT/Alert(®) SA (bioMérieux), and charcoal-containing BacT/Alert(®) FA (bioMérieux) blood culture bottles with direct identification by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). A total of 103 bacterial isolates, from clinical blood cultures, representing the most frequent 13 genera and 24 species were examined. Bacteria were extracted from positive blood culture broth by density centrifugation and then subjected to identification by MALDI-TOF MS using two different volumes and chemical treatments. Overall, correct identification by MALDI-TOF MS was obtained for the BD BACTEC™ Plus-Aerobic, BacT/Alert(®) SA, and BacT/Alert(®) FA blood culture bottles in 72%, 45.6%, and 23%, respectively, for gram-negative bacteria in 86.6%, 69.2%, and 47.1%, respectively, and for gram-positive bacteria in 60.0%, 28.8%, and 5.4%, respectively. The lack of identification was observed mainly with viridans streptococci. Depending on the blood culture bottles used in routine diagnostic procedures and the protocol for bacterial preparation, the applied MALDI-TOF MS represents an efficient and rapid method for direct bacterial identification.

Centrifugation speed affects light transmission aggregometry.

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Merolla, M; Nardi, M A; Berger, J S

2012-02-01

Light transmission aggregometry (LTA) is considered the gold standard for investigating platelet activity ex vivo. However, LTA protocols are not standardized, and differences in LTA procedure are a potential source of variance in results. Centrifugation speed is an essential component of platelet preparation in LTA, has yet to be standardized, and may affect platelet aggregation results. We sought to investigate the effect of relative centrifugal force (RCF) intensity on LTA results. Ten healthy controls had venous blood drawn and centrifuged at 150, 200, 300, and 500 g for 10 min. Cell counts in whole blood and platelet-rich plasma (PRP) were measured using a hematology analyzer. LTA was performed using 1.0 μm adenosine diphosphate (ADP) and 0.4 μm epinephrine as an agonist. Aggregation (%) was compared at 60, 120, 180, and 300 s and at maximum aggregation. Centrifugation speed was associated with decreasing platelet count (P centrifuge RCF at 60, 120, 180, 300 s and at maximum aggregation (P centrifugation speed in the interpretation of LTA results, supporting the need for standardization of centrifugation RCF in LTA protocols. © 2011 Blackwell Publishing Ltd.

An evaluation of three processing methods and the effect of reduced culture times for faster direct identification of pathogens from BacT/ALERT blood cultures by MALDI-TOF MS.

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Loonen, A J M; Jansz, A R; Stalpers, J; Wolffs, P F G; van den Brule, A J C

2012-07-01

Matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) is a fast and reliable method for the identification of bacteria from agar media. Direct identification from positive blood cultures should decrease the time to obtaining the result. In this study, three different processing methods for the rapid direct identification of bacteria from positive blood culture bottles were compared. In total, 101 positive aerobe BacT/ALERT bottles were included in this study. Aliquots from all bottles were used for three bacterial processing methods, i.e. the commercially available Bruker's MALDI Sepsityper kit, the commercially available Molzym's MolYsis Basic5 kit and a centrifugation/washing method. In addition, the best method was used to evaluate the possibility of MALDI application after a reduced incubation time of 7 h of Staphylococcus aureus- and Escherichia coli-spiked (1,000, 100 and 10 colony-forming units [CFU]) aerobe BacT/ALERT blood cultures. Sixty-six (65%), 51 (50.5%) and 79 (78%) bottles were identified correctly at the species level when the centrifugation/washing method, MolYsis Basic 5 and Sepsityper were used, respectively. Incorrect identification was obtained in 35 (35%), 50 (49.5%) and 22 (22%) bottles, respectively. Gram-positive cocci were correctly identified in 33/52 (64%) of the cases. However, Gram-negative rods showed a correct identification in 45/47 (96%) of all bottles when the Sepsityper kit was used. Seven hours of pre-incubation of S. aureus- and E. coli-spiked aerobe BacT/ALERT blood cultures never resulted in reliable identification with MALDI-TOF MS. Sepsityper is superior for the direct identification of microorganisms from aerobe BacT/ALERT bottles. Gram-negative pathogens show better results compared to Gram-positive bacteria. Reduced incubation followed by MALDI-TOF MS did not result in faster reliable identification.

Prediction and verification of centrifugal dewatering of P. pastoris fermentation cultures using an ultra scale-down approach.

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Lopes, A G; Keshavarz-Moore, E

2012-08-01

Recent years have seen a dramatic rise in fermentation broth cell densities and a shift to extracellular product expression in microbial cells. As a result, dewatering characteristics during cell separation is of importance, as any liquor trapped in the sediment results in loss of product, and thus a decrease in product recovery. In this study, an ultra scale-down (USD) approach was developed to enable the rapid assessment of dewatering performance of pilot-scale centrifuges with intermittent solids discharge. The results were then verified at scale for two types of pilot-scale centrifuges: a tubular bowl equipment and a disk-stack centrifuge. Initial experiments showed that employing a laboratory-scale centrifugal mimic based on using a comparable feed concentration to that of the pilot-scale centrifuge, does not successfully predict the dewatering performance at scale (P-value centrifuge. Initial experiments used Baker's yeast feed suspensions followed by fresh Pichia pastoris fermentation cultures. This work presents a simple and novel USD approach to predict dewatering levels in two types of pilot-scale centrifuges using small quantities of feedstock (centrifuge needs to be operated, reducing the need for repeated pilot-scale runs during early stages of process development. Copyright © 2012 Wiley Periodicals, Inc.

Haemopoietic progenitor cells in human peripheral blood

International Nuclear Information System (INIS)

Zwaan, F.E.

1980-01-01

The purpose of the investigation reported is to purify haemopoietic progenitor cells from human peripheral blood using density gradient centrifugation in order to isolate a progenitor cell fraction without immunocompetent cells. The purification technique of peripheral blood flow colony forming unit culture (CFU-c) by means of density gradient centrifugation and a combined depletion of various rosettes is described. The results of several 'in vitro' characteristics of purified CFU-c suspensions and of the plasma clot diffusion chamber culture technique are presented. Irradiation studies revealed that for both human bone marrow and peripheral blood the CFU-c were less radioresistant than clusters. Elimination of monocytes (and granulocytes) from the test suspensions induced an alteration in radiosensitivity pararmeters. The results obtained with the different techniques are described by analysing peripheral progenitor cell activity in myeloproliferative disorders. (Auth.)

Comparison of PCR-Based Diagnosis with Centrifuged-Based Enrichment Method for Detection of Borrelia Persica in Animal Blood Samples

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SR Naddaf

2011-06-01

Full Text Available Background: The mainstay of diagnosis of relapsing fever (RF is demonstration of the spirochetes in Giemsa-stained thick blood smears, but during non fever periods the bacteria are very scanty and rarely detected in blood smears by mi­cros­copy. This study is aimed to evaluate the sensitivity of different methods developed for detection of low-grade spi­ro­chetemia. Methods: Animal blood samples with low degrees of spirochetemia were tested with two PCRs and a nested PCR tar­get­ing flaB, GlpQ, and rrs genes. Also, a centrifuged-based enrichment method and Giemsa staining were per­formed on blood samples with various degrees of spirochetemia. Results: The flaB-PCR and nested rrs-PCR turned positive with various degrees of spirochetemia including the blood samples that turned negative with dark-field microscopy. The GlpQ-PCR was positive as far as at least one spi­ro­chete was seen in 5-10 microscopic fields. The sensitivity of GlpQ-PCR increased when DNA from Buffy Coat Layer (BCL was used as template. The centrifuged-based enrichment method turned positive with as low concentra­tion as 50 bacteria/ml blood, while Giemsa thick staining detected bacteria with concentrations ≥ 25000 bacteria/ml. Conclusion: Centrifuged-based enrichment method appeared as much as 500-fold more sensitive than thick smears, which makes it even superior to some PCR assays. Due to simplicity and minimal laboratory requirements, this method can be considered a valuable tool for diagnosis of RF in rural health centers.

Comparison of PCR-Based Diagnosis with Centrifuged-Based Enrichment Method for Detection of Borrelia persica in Animal Blood Samples

Directory of Open Access Journals (Sweden)

SR Naddaf

2011-06-01

Background: The mainstay of diagnosis of relapsing fever (RF is demonstration of the spirochetes in Giemsa-stained thick blood smears, but during non fever periods the bacteria are very scanty and rarely detected in blood smears by mi­cros­copy. This study is aimed to evaluate the sensitivity of different methods developed for detection of low-grade spi­ro­chetemia. Methods: Animal blood samples with low degrees of spirochetemia were tested with two PCRs and a nested PCR tar­get­ing flaB, GlpQ, and rrs genes. Also, a centrifuged-based enrichment method and Giemsa staining were per­formed on blood samples with various degrees of spirochetemia. Results: The flaB-PCR and nested rrs-PCR turned positive with various degrees of spirochetemia including the blood samples that turned negative with dark-field microscopy. The GlpQ-PCR was positive as far as at least one spi­ro­chete was seen in 5-10 microscopic fields. The sensitivity of GlpQ-PCR increased when DNA from Buffy Coat Layer (BCL was used as template. The centrifuged-based enrichment method turned positive with as low concentra­tion as 50 bacteria/ml blood, while Giemsa thick staining detected bacteria with concentrations ≥ 25000 bacteria/ml.  Conclusion: Centrifuged-based enrichment method appeared as much as 500-fold more sensitive than thick smears, which makes it even superior to some PCR assays. Due to simplicity and minimal laboratory requirements, this method can be considered a valuable tool for diagnosis of RF in rural health centers.  

A Novel Centrifugation Method Using a Cell Salvage Device Offers an Alternative to the Use of Leukocyte-Depleting Filters for Autologous Blood Transfusions.

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Barchilon, Michael; Gaspar, Cristina; Mexas, Angela; Nieter, Don

2016-12-01

Autotransfusion protocols often use the use of costly filters, such as leukocyte-depleting filters (LDFs), to minimize reinfusion of activated leukocytes and inflammatory mediators associated with reperfusion injury (RI). LDFs are used extensively in hospital settings; however, they represent an additional capital expenditure for hospitals, as well as a constraint on the reinfusion rate of blood products for health-care providers. We compared a commonly used LDF to a novel centrifugation method employing a widely used cell salvage device. Complete blood counts and enzyme-linked immunosorbent assays (ELISAs) measuring tumor necrosis factor-α (TNF-α) and interleukin-2 (IL-2) were performed to compare the efficacy of these methodologies. The LDF removed, on average, 94% of all leukocytes, including 96% of neutrophils. The centrifugation method removed, on average, 89% of all leukocytes, including 91% of neutrophils and resulted in a highly concentrated red blood cell product. Our results suggest both methods offer equivalent leukocyte reduction. TNF-α was also comparably reduced following our novel centrifugation method and the LDF method and IL-2 levels were undetectable in all samples. These results indicate our novel centrifugation method may preclude the need for a LDF during select autotransfusion applications.

Development of a 3D circular microfluidic centrifuge for the separation of mixed particles by using their different centrifuge times

International Nuclear Information System (INIS)

Jeon, H J; Kim, D I; Kim, M J; Nguyen, X D; Park, D H; Go, J S

2015-01-01

This paper presents a circular microfluidic centrifuge with two inlets and two outlets to separate mixed microparticles with a specially designed sample injection hole. To separate the mixed particles, it uses a rotational flow, generated in a chamber by counter primary flows in the microchannels. The shape and sizes of the circular microfluidic centrifuge have been designed through numerical evaluation to have a large relative centrifugal force. The difference of centrifuge times of the mixed particles of 1 μm and 6 μm was determined to be 8.2 s at an inlet Reynolds number of 500 and a sample Reynolds number of 20. In the experiment, this was measured to be about 10 s. From the separation of the two polymer particles analogous to the representative sizes of platelets and red blood cells, the circular microfluidic centrifuge shows a potential to separate human blood cells size-selectively by using the difference of centrifuge times. (paper)

21 CFR 864.9285 - Automated cell-washing centrifuge for immuno-hematology.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Automated cell-washing centrifuge for immuno... Establishments That Manufacture Blood and Blood Products § 864.9285 Automated cell-washing centrifuge for immuno-hematology. (a) Identification. An automated cell-washing centrifuge for immuno-hematology is a device used...

Differential white cell count by centrifugal microfluidics.

Energy Technology Data Exchange (ETDEWEB)

Sommer, Gregory Jon; Tentori, Augusto M.; Schaff, Ulrich Y.

2010-07-01

We present a method for counting white blood cells that is uniquely compatible with centrifugation based microfluidics. Blood is deposited on top of one or more layers of density media within a microfluidic disk. Spinning the disk causes the cell populations within whole blood to settle through the media, reaching an equilibrium based on the density of each cell type. Separation and fluorescence measurement of cell types stained with a DNA dye is demonstrated using this technique. The integrated signal from bands of fluorescent microspheres is shown to be proportional to their initial concentration in suspension. Among the current generation of medical diagnostics are devices based on the principle of centrifuging a CD sized disk functionalized with microfluidics. These portable 'lab on a disk' devices are capable of conducting multiple assays directly from a blood sample, embodied by platforms developed by Gyros, Samsung, and Abaxis. [1,2] However, no centrifugal platform to date includes a differential white blood cell count, which is an important metric complimentary to diagnostic assays. Measuring the differential white blood cell count (the relative fraction of granulocytes, lymphocytes, and monocytes) is a standard medical diagnostic technique useful for identifying sepsis, leukemia, AIDS, radiation exposure, and a host of other conditions that affect the immune system. Several methods exist for measuring the relative white blood cell count including flow cytometry, electrical impedance, and visual identification from a stained drop of blood under a microscope. However, none of these methods is easily incorporated into a centrifugal microfluidic diagnostic platform.

Designing and modeling a centrifugal microfluidic device to separate target blood cells

Science.gov (United States)

Shamloo, Amir; Selahi, AmirAli; Madadelahi, Masoud

2016-03-01

The objective of this study is to design a novel and efficient portable lab-on-a-CD (LOCD) microfluidic device for separation of specific cells (target cells) using magnetic beads. In this study the results are shown for neutrophils as target cells. However, other kinds of target cells can be separated in a similar approach. The designed microfluidics can be utilized as a point of care system for neutrophil detection. This microfluidic system employs centrifugal and magnetic forces for separation. After model validation by the experimental data in the literature (that may be used as a design tool for developing centrifugo-magnetophoretic devices), two models are presented for separation of target cells using magnetic beads. The first model consists of one container in the inlet section and two containers in the outlets. Initially, the inlet container is filled with diluted blood sample which is a mixture of red blood cells (RBCs) plus neutrophils which are attached to Magnetic beads. It is shown that by using centrifugal and magnetic forces, this model can separate all neutrophils with recovery factor of ~100%. In the second model, due to excess of magnetic beads in usual experimental analysis (to ensure that all target cells are attached to them) the geometry is improved by adding a third outlet for these free magnetic beads. It is shown that at angular velocity of 45 rad s-1, recovery factor of 100% is achievable for RBCs, free magnetic beads and neutrophils as target cells.

Blood culture bottles are superior to conventional media for vitreous culture.

Science.gov (United States)

Thariya, Patsuda; Yospaiboon, Yosanan; Sinawat, Suthasinee; Sanguansak, Thuss; Bhoomibunchoo, Chavakij; Laovirojjanakul, Wipada

2016-08-01

To compare blood culture bottles and conventional media for the vitreous culture in patients with clinically suspected infectious endophthalmitis. Retrospective comparative study at KKU Eye Center, Khon Kaen University. There were 342 patients with clinically suspected infectious endophthalmitis participated in the study. The vitreous specimens were inoculated in both blood culture bottles and on conventional culture media (blood agar, MacConkey agar, chocolate agar, Sabouraud dextrose agar and thioglycolate broth). The number of positive culture yields in both blood culture bottles and conventional media. Positive culture yields in both methods were found in 151 eyes (49.5%). There were 136 of 151 eyes (90.1%) with positive culture in blood culture bottles, whereas 99 of 151 eyes (65.6%) yielded positive cultures in conventional media. These findings were different with a statistical significance (P culture bottles and conventional media improved the yield. Blood culture bottles are superior to conventional media for vitreous culture in clinically suspected infectious endophthalmitis. Vitreous culture using blood culture bottles should be recommended as the primary method for microbiological diagnosis. A combination of both methods further improves the positive culture yield. © 2016 Royal Australian and New Zealand College of Ophthalmologists.

Electrical lysis: dynamics revisited and advances in On-chip operation.

Science.gov (United States)

Morshed, Bashir; Shams, Maitham; Mussivand, Tofy

2013-01-01

Electrical lysis (EL) is the process of breaking the cell membrane to expose the internal contents under an applied high electric field. Lysis is an important phenomenon for cellular analysis, medical treatment, and biofouling control. This paper aims to review, summarize, and analyze recent advancements on EL. Major databases including PubMed, Ei Engineering Village, IEEE Xplore, and Scholars Portal were searched using relevant keywords. More than 50 articles published in English since 1997 are cited in this article. EL has several key advantages compared to other lysis techniques such as chemical, mechanical, sonication, or laser, including rapid speed of operation, ability to control, miniaturization, low cost, and low power requirement. A variety of cell types have been investigated for including protoplasts, E. coli, yeasts, blood cells, and cancer cells. EL has been developed and applied for decontamination, cytology, genetics, single-cell analysis, cancer treatment, and other applications. On-chip EL is a promising technology for multiplexed automated implementation of cell-sample preparation and processing with micro- or nanoliter reagents.

Influence of centrifugation conditions on the results of 77 routine clinical chemistry analytes using standard vacuum blood collection tubes and the new BD-Barricor tubes.

Science.gov (United States)

Cadamuro, Janne; Mrazek, Cornelia; Leichtle, Alexander B; Kipman, Ulrike; Felder, Thomas K; Wiedemann, Helmut; Oberkofler, Hannes; Fiedler, Georg M; Haschke-Becher, Elisabeth

2018-02-15

Although centrifugation is performed in almost every blood sample, recommendations on duration and g-force are heterogeneous and mostly based on expert opinions. In order to unify this step in a fully automated laboratory, we aimed to evaluate different centrifugation settings and their influence on the results of routine clinical chemistry analytes. We collected blood from 41 healthy volunteers into BD Vacutainer PST II-heparin-gel- (LiHepGel), BD Vacutainer SST II-serum-, and BD Vacutainer Barricor heparin-tubes with a mechanical separator (LiHepBar). Tubes were centrifuged at 2000xg for 10 minutes and 3000xg for 7 and 5 minutes, respectively. Subsequently 60 and 21 clinical chemistry analytes were measured in plasma and serum samples, respectively, using a Roche COBAS instrument. High sensitive Troponin T, pregnancy-associated plasma protein A, ß human chorionic gonadotropin and rheumatoid factor had to be excluded from statistical evaluation as many of the respective results were below the measuring range. Except of free haemoglobin (fHb) measurements, no analyte result was altered by the use of shorter centrifugation times at higher g-forces. Comparing LiHepBar to LiHepGel tubes at different centrifugation setting, we found higher lactate-dehydrogenase (LD) (P = 0.003 to centrifuged at higher speed (3000xg) for a shorter amount of time (5 minutes) without alteration of the analytes tested in this study. When using LiHepBar tubes for blood collection, a separate LD reference value might be needed.

Isolation of human monocytes by double gradient centrifugation and their differentiation to macrophages in teflon-coated cell culture bags.

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Menck, Kerstin; Behme, Daniel; Pantke, Mathias; Reiling, Norbert; Binder, Claudia; Pukrop, Tobias; Klemm, Florian

2014-09-09

Human macrophages are involved in a plethora of pathologic processes ranging from infectious diseases to cancer. Thus they pose a valuable tool to understand the underlying mechanisms of these diseases. We therefore present a straightforward protocol for the isolation of human monocytes from buffy coats, followed by a differentiation procedure which results in high macrophage yields. The technique relies mostly on commonly available lab equipment and thus provides a cost and time effective way to obtain large quantities of human macrophages. Briefly, buffy coats from healthy blood donors are subjected to a double density gradient centrifugation to harvest monocytes from the peripheral blood. These monocytes are then cultured in fluorinated ethylene propylene (FEP) Teflon-coated cell culture bags in the presence of macrophage colony-stimulating factor (M-CSF). The differentiated macrophages can be easily harvested and used for subsequent studies and functional assays. Important methods for quality control and validation of the isolation and differentiation steps will be highlighted within the protocol. In summary, the protocol described here enables scientists to routinely and reproducibly isolate human macrophages without the need for cost intensive tools. Furthermore, disease models can be studied in a syngeneic human system circumventing the use of murine macrophages.

Use of short-radius centrifugation to augment ankle-brachial indices.

Science.gov (United States)

Grenon, S Marlene; Mateus, Jaime; Hsiang, York; Sidhu, Ravi; Young, Laurence; Gagnon, Joel

2009-06-01

Peripheral arterial disease is mainly caused by atherosclerosis and is characterized by decreased circulation, lower blood pressure, and insufficient tissue perfusion in the lower extremities. The hemodynamics of standing and altered gravity environments have been well studied relative to arm blood pressures but are less well understood for ankle pressures. Because regional blood pressure depends, in part, on the gravitational pressure gradient, we hypothesized that artificial gravity exposure on a short-arm centrifuge with the center of rotation above the head would increase blood pressure in the lower extremities. Cardiovascular parameters for 12 healthy subjects were measured during exposure to supine short-arm centrifugation at 20, 25, and 30 revolutions per minute (rpm), corresponding to centripetal accelerations of 0.94, 1.47, and 2.11 Gz at the foot level, respectively. Systolic ankle blood pressure significantly increased at all levels of centrifugation. Ankle-brachial indices (the ratio of systolic ankle to arm blood pressures) increased significantly from 1.17 +/- 0.03 to 1.58 +/- 0.03 at 0.94 Gz (P blood pressure significantly increased at 2.11 Gz, but heart rate did not change significantly. All parameters returned to normal after cessation of centrifugation. We demonstrated that short-radius centrifugation leads to an increase in ankle-brachial indices. This could have potential implications for the treatment of peripheral arterial disease.

Lysis of typhus-group rickettsia-infected targets by lymphokine activated killers

International Nuclear Information System (INIS)

Carl, M.; Dasch, G.A.

1986-01-01

The authors recently described a subset of OKT8, OKT3-positive lymphocytes from typhus-group rickettsia immune individuals which were capable of lysing autologous PHA-blasts or Epstein-Barr virus transformed B cells (LCL) infected with typhus-group rickettsiae. In order to determine if killing by these effectors was HLA-restricted, they stimulated peripheral blood mononuclear cells (PBMC) from typhus-group rickettsia immune individuals in vitro with typhus-group rickettsia-derived antigen for one week and then measured lysis of autologous LCL or HLA-mismatched LCL in a 4-6 hour Cr 51 -release assay. There was significant lysis of both the autologous and the HLA-mismatched infected targets as compared to the corresponding uninfected targets. Since this suggested that the effectors were lymphokine activated killers (LAK) rather than cytotoxic T lymphocytes, they then tested this hypothesis by stimulating PBMC from both immune and non-immune individuals in vitro for one week with purified interleukin 2 and measuring lysis of infected, autologous LCL. PBMC thus treated, from both immune and non-immune individuals, were capable of significantly lysing autologous, infected LCL as compared to the non-infected control. They therefore conclude that targets infected with typhus-group rickettsiae are susceptible to lysis to LAK

Induced Pluripotent Stem Cell Generation from Blood Cells Using Sendai Virus and Centrifugation.

Science.gov (United States)

Rim, Yeri Alice; Nam, Yoojun; Ju, Ji Hyeon

2016-12-21

The recent development of human induced pluripotent stem cells (hiPSCs) proved that mature somatic cells can return to an undifferentiated, pluripotent state. Now, reprogramming is done with various types of adult somatic cells: keratinocytes, urine cells, fibroblasts, etc. Early experiments were usually done with dermal fibroblasts. However, this required an invasive surgical procedure to obtain fibroblasts from the patients. Therefore, suspension cells, such as blood and urine cells, were considered ideal for reprogramming because of the convenience of obtaining the primary cells. Here, we report an efficient protocol for iPSC generation from peripheral blood mononuclear cells (PBMCs). By plating the transduced PBMCs serially to a new, matrix-coated plate using centrifugation, this protocol can easily provide iPSC colonies. This method is also applicable to umbilical cord blood mononuclear cells (CBMCs). This study presents a simple and efficient protocol for the reprogramming of PBMCs and CBMCs.

Impact of age and diagnosis on viability during centrifugation and cryopreservation of peripheral blood stem cell products.

Science.gov (United States)

Civriz Bozdag, S; Bay, M; Ayyıldız, E; Topcuoglu, P; Ilhan, O

2012-08-01

The viability of the hematopoietic stem cells infused to the patient is important for transplant outcome. We evaluated 31 peripheral blood stem cell product collected from 15 patients. We aimed to check the viabilities of the cells from patients with different age and diagnosis, in different stages of the cryopreservation procedure. We showed a markedly decreased viability rate after centrifugation and addition of DMSO. Percentages of viabilities were similar between young and old patients in each step. Type of hematological malignancy did not make a significant influence on the viability. High speed centrifugation has a negative impact on the viability. Copyright © 2012 Elsevier Ltd. All rights reserved.

Time-to-positivity, type of culture media and oxidase test performed on positive blood culture vials to predict Pseudomonas aeruginosa in patients with Gram-negative bacilli bacteraemia.

Science.gov (United States)

Cobos-Triguero, N; Zboromyrska, Y; Morata, L; Alejo, I; De La Calle, C; Vergara, A; Cardozo, C; Arcas, M P; Soriano, A; Marco, F; Mensa, J; Almela, M; Martínez, J A

2017-02-01

The aim of this study was to determine the usefulness of oxidase test and time-to-positivity (TTP) in aerobic and anaerobic blood culture vials to detect the presence of Pseudomonas aeruginosa in patients with Gram-negative bacilli (GNB) bacteraemia. TTP was recorded for each aerobic and anaerobic blood culture vial of monomicrobial bacteraemia due to GNB. Oxidase test was performed in a pellet of the centrifuged content of the positive blood culture. An algorithm was developed in order to perform the oxidase test efficiently taking into account TTP and type of vial. A total of 341 episodes of GNB bacteraemia were analysed. Sensitivity, specificity, positive predictive value and negative predictive value of the oxidase test performed on positive vials with GNB to predict P. aeruginosa were 95%, 99%, 91%, and 99%, respectively. When growth was first or exclusively detected in anaerobic vials, P. aeruginosa was never identified hence the performance of the oxidase test could be avoided. When growth was only or first detected in aerobic vials, a TTP≥8h predicted P. aeruginosa in 37% or cases (63 of 169), therefore oxidase test is highly recommended. Oxidase test performed onto positive blood culture vials previously selected by TTP and type of vials is an easy and inexpensive way to predict P. aeruginosa. In most cases, this can lead to optimization of treatment in less than 24 hours.

Designing and modeling a centrifugal microfluidic device to separate target blood cells

International Nuclear Information System (INIS)

Shamloo, Amir; Selahi, AmirAli; Madadelahi, Masoud

2016-01-01

The objective of this study is to design a novel and efficient portable lab-on-a-CD (LOCD) microfluidic device for separation of specific cells (target cells) using magnetic beads. In this study the results are shown for neutrophils as target cells. However, other kinds of target cells can be separated in a similar approach. The designed microfluidics can be utilized as a point of care system for neutrophil detection. This microfluidic system employs centrifugal and magnetic forces for separation. After model validation by the experimental data in the literature (that may be used as a design tool for developing centrifugo-magnetophoretic devices), two models are presented for separation of target cells using magnetic beads. The first model consists of one container in the inlet section and two containers in the outlets. Initially, the inlet container is filled with diluted blood sample which is a mixture of red blood cells (RBCs) plus neutrophils which are attached to Magnetic beads. It is shown that by using centrifugal and magnetic forces, this model can separate all neutrophils with recovery factor of ∼100%. In the second model, due to excess of magnetic beads in usual experimental analysis (to ensure that all target cells are attached to them) the geometry is improved by adding a third outlet for these free magnetic beads. It is shown that at angular velocity of 45 rad s −1 , recovery factor of 100% is achievable for RBCs, free magnetic beads and neutrophils as target cells. (paper)

In vitro differentiation of human umbilical cord blood mesenchymal ...

African Journals Online (AJOL)

Mesenchymal stem cells (MSCs) were isolated by gradient density centrifugation from umbilical cord blood. Spindle-shaped adherent cells were permitted to grow to 70% confluence in primary culture media which was reached by day 12. Induction of differentiation started by culturing cells with differentiation medium ...

Density-Gradient Mediated Band Extraction of Leukocytes from Whole Blood Using Centrifugo-Pneumatic Siphon Valving on Centrifugal Microfluidic Discs

Science.gov (United States)

Kearney, Sinéad M.; Kilcawley, Niamh A.; Early, Philip L.; Glynn, Macdara T.; Ducrée, Jens

2016-01-01

Here we present retrieval of Peripheral Blood Mononuclear Cells by density-gradient medium based centrifugation for subsequent analysis of the leukocytes on an integrated microfluidic “Lab-on-a-Disc” cartridge. Isolation of white blood cells constitutes a critical sample preparation step for many bioassays. Centrifugo-pneumatic siphon valves are particularly suited for blood processing as they function without need of surface treatment and are ‘low-pass’, i.e., holding at high centrifugation speeds and opening upon reduction of the spin rate. Both ‘hydrostatically’ and ‘hydrodynamically’ triggered centrifugo-pneumatic siphon valving schemes are presented. Firstly, the geometry of the pneumatic chamber of hydrostatically primed centrifugo-pneumatic siphon valves is optimised to enable smooth and uniform layering of blood on top of the density-gradient medium; this feature proves to be key for efficient Peripheral Blood Mononuclear Cell extraction. A theoretical analysis of hydrostatically primed valves is also presented which determines the optimum priming pressure for the individual valves. Next, ‘dual siphon’ configurations for both hydrostatically and hydrodynamically primed centrifugo-pneumatic siphon valves are introduced; here plasma and Peripheral Blood Mononuclear Cells are extracted through a distinct siphon valve. This work represents a first step towards enabling on disc multi-parameter analysis. Finally, the efficiency of Peripheral Blood Mononuclear Cells extraction in these structures is characterised using a simplified design. A microfluidic mechanism, which we termed phase switching, is identified which affects the efficiency of Peripheral Blood Mononuclear Cell extraction. PMID:27167376

Optimisation of a double-centrifugation method for preparation of canine platelet-rich plasma.

Science.gov (United States)

Shin, Hyeok-Soo; Woo, Heung-Myong; Kang, Byung-Jae

2017-06-26

Platelet-rich plasma (PRP) has been expected for regenerative medicine because of its growth factors. However, there is considerable variability in the recovery and yield of platelets and the concentration of growth factors in PRP preparations. The aim of this study was to identify optimal relative centrifugal force and spin time for the preparation of PRP from canine blood using a double-centrifugation tube method. Whole blood samples were collected in citrate blood collection tubes from 12 healthy beagles. For the first centrifugation step, 10 different run conditions were compared to determine which condition produced optimal recovery of platelets. Once the optimal condition was identified, platelet-containing plasma prepared using that condition was subjected to a second centrifugation to pellet platelets. For the second centrifugation, 12 different run conditions were compared to identify the centrifugal force and spin time to produce maximal pellet recovery and concentration increase. Growth factor levels were estimated by using ELISA to measure platelet-derived growth factor-BB (PDGF-BB) concentrations in optimised CaCl 2 -activated platelet fractions. The highest platelet recovery rate and yield were obtained by first centrifuging whole blood at 1000 g for 5 min and then centrifuging the recovered platelet-enriched plasma at 1500 g for 15 min. This protocol recovered 80% of platelets from whole blood and increased platelet concentration six-fold and produced the highest concentration of PDGF-BB in activated fractions. We have described an optimised double-centrifugation tube method for the preparation of PRP from canine blood. This optimised method does not require particularly expensive equipment or high technical ability and can readily be carried out in a veterinary clinical setting.

Performance of Gram staining on blood cultures flagged negative by an automated blood culture system.

Science.gov (United States)

Peretz, A; Isakovich, N; Pastukh, N; Koifman, A; Glyatman, T; Brodsky, D

2015-08-01

Blood is one of the most important specimens sent to a microbiology laboratory for culture. Most blood cultures are incubated for 5-7 days, except in cases where there is a suspicion of infection caused by microorganisms that proliferate slowly, or infections expressed by a small number of bacteria in the bloodstream. Therefore, at the end of incubation, misidentification of positive cultures and false-negative results are a real possibility. The aim of this work was to perform a confirmation by Gram staining of the lack of any microorganisms in blood cultures that were identified as negative by the BACTEC™ FX system at the end of incubation. All bottles defined as negative by the BACTEC FX system were Gram-stained using an automatic device and inoculated on solid growth media. In our work, 15 cultures that were defined as negative by the BACTEC FX system at the end of the incubation were found to contain microorganisms when Gram-stained. The main characteristic of most bacteria and fungi growing in the culture bottles that were defined as negative was slow growth. This finding raises a problematic issue concerning the need to perform Gram staining of all blood cultures, which could overload the routine laboratory work, especially laboratories serving large medical centers and receiving a large number of blood cultures.

Factors influencing lysis time stochasticity in bacteriophage λ

Directory of Open Access Journals (Sweden)

Dennehy John J

2011-08-01

Full Text Available Abstract Background Despite identical genotypes and seemingly uniform environments, stochastic gene expression and other dynamic intracellular processes can produce considerable phenotypic diversity within clonal microbes. One trait that provides a good model to explore the molecular basis of stochastic variation is the timing of host lysis by bacteriophage (phage. Results Individual lysis events of thermally-inducible λ lysogens were observed using a temperature-controlled perfusion chamber mounted on an inverted microscope. Both mean lysis time (MLT and its associated standard deviation (SD were estimated. Using the SD as a measure of lysis time stochasticity, we showed that lysogenic cells in controlled environments varied widely in lysis times, and that the level of lysis time stochasticity depended on allelic variation in the holin sequence, late promoter (pR' activity, and host growth rate. In general, the MLT was positively correlated with the SD. Both lower pR' activities and lower host growth rates resulted in larger SDs. Results from premature lysis, induced by adding KCN at different time points after lysogen induction, showed a negative correlation between the timing of KCN addition and lysis time stochasticity. Conclusions Taken together with results published by others, we conclude that a large fraction of λ lysis time stochasticity is the result of random events following the expression and diffusion of the holin protein. Consequently, factors influencing the timing of reaching critical holin concentrations in the cell membrane, such as holin production rate, strongly influence the mean lysis time and the lysis time stochasticity.

Geometric Optimization for Non-Thrombogenicity of a Centrifugal Blood Pump through Flow Visualization

Science.gov (United States)

Toyoda, Masahiro; Nishida, Masahiro; Maruyama, Osamu; Yamane, Takashi; Tsutsui, Tatsuo; Sankai, Yoshiyuki

A monopivot centrifugal blood pump, whose impeller is supported with a pivot bearing and a passive magnetic bearing, is under development for implantable artificial heart. The hemolysis level is less than that of commercial centrifugal pumps and the pump size is as small as 160 mL in volume. To solve a problem of thrombus caused by fluid dynamics, flow visualization experiments and animal experiments have been undertaken. For flow visualization a three-fold scale-up model, high-speed video system, and particle tracking velocimetry software were used. To verify non-thrombogenicity one-week animal experiments were conducted with sheep. The initially observed thrombus around the pivot was removed through unifying the separate washout holes to a small centered hole to induce high shear around the pivot. It was found that the thrombus contours corresponded to the shear rate of 300s-1 for red thrombus and 1300-1700s-1 for white thrombus, respectively. Thus flow visualization technique was found to be a useful tool to predict thrombus location.

Direct identification of bacteria from positive BacT/ALERT blood culture bottles using matrix-assisted laser desorption ionization-time-of-flight mass spectrometry.

Science.gov (United States)

Mestas, Javier; Felsenstein, Susanna; Bard, Jennifer Dien

2014-11-01

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry is a fast and robust method for the identification of bacteria. In this study, we evaluate the performance of a laboratory-developed lysis method (LDT) for the rapid identification of bacteria from positive BacT/ALERT blood culture bottles. Of the 168 positive bottles tested, 159 were monomicrobial, the majority of which were Gram-positive organisms (61.0% versus 39.0%). Using a cut-off score of ≥1.7, 80.4% of the organisms were correctly identified to the species level, and the identification rate of Gram-negative organisms (90.3%) was found to be significantly greater than that of Gram-positive organisms (78.4%). The simplicity and cost-effectiveness of the LDT enable it to be fully integrated into the routine workflow of the clinical microbiology laboratory, allowing for rapid identification of Gram-positive and Gram-negative bacteria within an hour of blood culture positivity. Copyright © 2014 Elsevier Inc. All rights reserved.

Use of a centrifugation-based, point-of-care device for production of canine autologous bone marrow and platelet concentrates.

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Thoesen, Michael S; Berg-Foels, Wendy S Vanden; Stokol, Tracy; Rassnick, Kenneth M; Jacobson, May S; Kevy, Sherwin V; Todhunter, Rory J

2006-10-01

To analyze a centrifugation-based, point-of-care device that concentrates canine platelets and bone marrow-derived cells. 19 adult sexually intact dogs. Anticoagulated peripheral blood (60 mL) and 60 mL of anticoagulated bone marrow aspirate (BMA) were concentrated by centrifugation with the centrifugation-based, point-of-care device to form a platelet and a bone marrow concentrate (BMC) from 11 dogs. Blood samples were analyzed on the basis of hemograms, platelet count, and PCV. The BMA and BMC were analyzed to determine PCV, total nucleated cell count, RBC count, and differential cell counts. The BMC stromal cells were cultured in an osteoinductive medium. Eight additional dogs were used to compare the BMC yield with that in which heparin was infused into the bone marrow before aspiration. The centrifugation-based, point-of-care device concentrated platelets by 6-fold over baseline (median recovery, 63.1%) with a median of 1,336 x 10(3) platelets/microL in the 7-mL concentrate. The nucleated cells in BMCs increased 7-fold (median recovery, 42.9%) with a median of 720 x 10(3) cells/microL in the 4-mL concentrate. The myeloid nucleated cells and mononuclear cells increased significantly in BMCs with a significant decrease in PCV, compared with that of BMAs. Stromal cell cultures expressed an osteoblastic phenotype in culture. Infusion of heparin into the bone marrow eliminated clot formation and created less variation in the yield (median recovery, 61.9%). Bone marrow-derived cell and platelet-rich concentrates may form bone if delivered in an engineered graft, thus decreasing the need for cancellous bone grafts.

An implantable centrifugal blood pump for long term circulatory support.

Science.gov (United States)

Yamazaki, K; Litwak, P; Kormos, R L; Mori, T; Tagusari, O; Antaki, J F; Kameneva, M; Watach, M; Gordon, L; Umezu, M; Tomioka, J; Koyanagi, H; Griffith, B P

1997-01-01

A compact centrifugal blood pump was developed as an implantable left ventricular assist system. The impeller diameter is 40 mm and the pump dimensions are 55 x 64 mm. This first prototype was fabricated from titanium alloy, resulting in a pump weight of 400 g including a brushless DC motor. Weight of the second prototype pump was reduced to 280 g. The entire blood contacting surface is coated with diamond like carbon to improve blood compatibility. Flow rates of over 7 L/min against 100 mmHg pressure at 2,500 rpm with 9 W total power consumption have been measured. A newly designed mechanical seal with a recirculating purge system ("Cool-Seal") is used as a shaft seal. In this seal system, seal temperature is kept under 40 degrees C to prevent heat denaturation of blood proteins. Purge fluid also cools the pump motor coil and journal bearing. The purge fluid is continuously purified and sterilized by an ultrafiltration filter incorporated into the paracorporeal drive console. In vitro experiments with bovine blood demonstrated an acceptably low hemolysis rate (normalized index of hemolysis = 0.005 +/- 0.002 g/100 L). In vivo experiments are currently ongoing using calves. Via left thoracotomy, left ventricular apex-descending aorta bypass was performed utilizing a PTFE (Polytetrafluoroethylene) vascular graft, with the pump placed in the left thoracic cavity. In two in vivo experiments, pump flow rate was maintained at 5-8 L/min, and pump power consumption remained stable at 9-10 W. All plasma free hemoglobin levels were measured at < 15 mg/dl. The seal system has demonstrated good seal capability with negligible purge fluid consumption (< 0.5 ml/ day). Both animals remain under observation after 162 and 91 days of continuous pump function.

Spin doctors: new innovations for centrifugal apheresis.

Science.gov (United States)

Kambic, H E; Nosé, Y

1997-08-01

The preparation of plasma from blood has a long history dating back to the early 1900s when the concept of blood washing replaced the traditional blood letting. Over the next 57 years landmark discoveries such as centrifugal and membrane filtration systems led to different and rapid plasma, solute, and cell separation. These were not singular events but rather events influenced by the converging chemical, physiological, and engineering advances that have characterized the latter half of the 20th century. These events have led to entire new fields of biomedical research. The biotechnology for on-line plasma separation and plasma treatment has opened a new era, expanding the application of extracorporeal technology to modern therapeutic medicine. The association of biochemical or cellular abnormalities with various disease states provides the rationale for therapeutic plasma exchange (the removal of large amounts of patient's plasma, alone or with replacement with crystalloid) and therapeutic cytopheresis (removal of cellular elements). The purpose of this review is to provide a historical picture of the innovative ideas of the spin doctors and their devices, which predate the centrifugal blood and cell separators commonplace to any hospital or blood bank worldwide. The emphasis is to define the historical events and their impacts on the development of centrifugal devices and apheresis technologies.

Synthesis and functioning of the colicin E1 lysis protein: Comparison with the colicin A lysis protein

International Nuclear Information System (INIS)

Cavard, D.

1991-01-01

The colicin E1 lysis protein, CelA, was identified as a 3-kDa protein in induced cells of Escherichia coli K-12 carrying pColE1 by pulse-chase labeling with either [ 35 S]cysteine or [ 3 H]lysine. This 3-kDa protein was acylated, as shown by [2- 3 H]glycerol labeling, and seemed to correspond to the mature CelA protein. The rate of modification and processing of CelA was different from that observed for Cal, the colicin A lysis protein. In contrast to Cal, no intermediate form was detected for CelA, no signal peptide accumulated, and no modified precursor form was observed after globomycin treatment. Thus, the rate of synthesis would not be specific to lysis proteins. Solubilization in sodium dodecyl sulfate of the mature forms of both CelA and Cal varied similarly at the time of colicin release, indicating a change in lysis protein structure. This particular property would play a role in the mechanism of colicin export. The accumulation of the signal peptide seems to be a factor determining the toxicity of the lysis proteins since CelA provoked less cell damage than Cal. Quasi-lysis and killing due to CelA were higher in degP mutants than in wild-type cells. They were minimal in pldA mutants

Suitability of small diagnostic peripheral-blood samples for cell-therapy studies.

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Stephanou, Coralea; Papasavva, Panayiota; Zachariou, Myria; Patsali, Petros; Epitropou, Marilena; Ladas, Petros; Al-Abdulla, Ruba; Christou, Soteroulla; Antoniou, Michael N; Lederer, Carsten W; Kleanthous, Marina

2017-02-01

Primary hematopoietic stem and progenitor cells (HSPCs) are key components of cell-based therapies for blood disorders and are thus the authentic substrate for related research. We propose that ubiquitous small-volume diagnostic samples represent a readily available and as yet untapped resource of primary patient-derived cells for cell- and gene-therapy studies. In the present study we compare isolation and storage methods for HSPCs from normal and thalassemic small-volume blood samples, considering genotype, density-gradient versus lysis-based cell isolation and cryostorage media with different serum contents. Downstream analyses include viability, recovery, differentiation in semi-solid media and performance in liquid cultures and viral transductions. We demonstrate that HSPCs isolated either by ammonium-chloride potassium (ACK)-based lysis or by gradient isolation are suitable for functional analyses in clonogenic assays, high-level HSPC expansion and efficient lentiviral transduction. For cryostorage of cells, gradient isolation is superior to ACK lysis, and cryostorage in freezing media containing 50% fetal bovine serum demonstrated good results across all tested criteria. For assays on freshly isolated cells, ACK lysis performed similar to, and for thalassemic samples better than, gradient isolation, at a fraction of the cost and hands-on time. All isolation and storage methods show considerable variation within sample groups, but this is particularly acute for density gradient isolation of thalassemic samples. This study demonstrates the suitability of small-volume blood samples for storage and preclinical studies, opening up the research field of HSPC and gene therapy to any blood diagnostic laboratory with corresponding bioethics approval for experimental use of surplus material. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Laboratory Workflow Analysis of Culture of Periprosthetic Tissues in Blood Culture Bottles.

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Peel, Trisha N; Sedarski, John A; Dylla, Brenda L; Shannon, Samantha K; Amirahmadi, Fazlollaah; Hughes, John G; Cheng, Allen C; Patel, Robin

2017-09-01

Culture of periprosthetic tissue specimens in blood culture bottles is more sensitive than conventional techniques, but the impact on laboratory workflow has yet to be addressed. Herein, we examined the impact of culture of periprosthetic tissues in blood culture bottles on laboratory workflow and cost. The workflow was process mapped, decision tree models were constructed using probabilities of positive and negative cultures drawn from our published study (T. N. Peel, B. L. Dylla, J. G. Hughes, D. T. Lynch, K. E. Greenwood-Quaintance, A. C. Cheng, J. N. Mandrekar, and R. Patel, mBio 7:e01776-15, 2016, https://doi.org/10.1128/mBio.01776-15), and the processing times and resource costs from the laboratory staff time viewpoint were used to compare periprosthetic tissues culture processes using conventional techniques with culture in blood culture bottles. Sensitivity analysis was performed using various rates of positive cultures. Annualized labor savings were estimated based on salary costs from the U.S. Labor Bureau for Laboratory staff. The model demonstrated a 60.1% reduction in mean total staff time with the adoption of tissue inoculation into blood culture bottles compared to conventional techniques (mean ± standard deviation, 30.7 ± 27.6 versus 77.0 ± 35.3 h per month, respectively; P < 0.001). The estimated annualized labor cost savings of culture using blood culture bottles was $10,876.83 (±$337.16). Sensitivity analysis was performed using various rates of culture positivity (5 to 50%). Culture in blood culture bottles was cost-effective, based on the estimated labor cost savings of $2,132.71 for each percent increase in test accuracy. In conclusion, culture of periprosthetic tissue in blood culture bottles is not only more accurate than but is also cost-saving compared to conventional culture methods. Copyright © 2017 American Society for Microbiology.

A mechanism of acquired resistance to complement-mediated lysis by Entamoeba histolytica.

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Gutiérrez-Kobeh, L; Cabrera, N; Pérez-Montfort, R

1997-04-01

Some Entamoeba histolytica strains resist complement-mediated lysis by serum. Susceptible and resistant strains activate the complement system equivalently, but resistant amebas evade killing by membrane attack complexes. Our objective was to determine the mechanism by which trophozoites of E. histolytica resist lysis by human serum. Amebas were made resistant to lysis by incubation with increasing concentrations of normal human serum. The possibility that resistant cells ingest membrane attack complexes was explored by subcellular fractionation of susceptible and resistant trophozoites treated with sublytic concentrations of human serum containing radiolabeled C9. In both cases, most of the label was in the fractions containing plasma membrane. The susceptible strain consistently showed more label associated with these fractions than the resistant strain. Thus, the possibility that the membrane attack complexes were released to the medium was explored. Both resistant and susceptible trophozoites release to the medium similar amounts of material excluded by Sepharose CL-2B in the presence or absence of normal human serum. Labeled C9 elutes together with the main bulk of proteins from the medium: this indicates that it is not in vesicles or high molecular weight aggregates. Coincubation of susceptible amebas with lysates of resistant trophozoites confers resistance to susceptible cells within 30 min. Resistance to lysis by serum can also be acquired by susceptible amebas after coincubation with lysates from human erythrocytes or after feeding them with whole human red blood cells. Resistant but not susceptible trophozoites show intense immunofluorescent staining on their surface with anti-human erythrocytic membrane antibody. These results suggest that amebas acquire resistance to lysis by serum by incorporating into their membranes complement regulatory proteins.

Direct Cellular Lysis/Protein Extraction Protocol for Soil Metaproteomics

Energy Technology Data Exchange (ETDEWEB)

Chourey, Karuna [ORNL; Jansson, Janet [Lawrence Berkeley National Laboratory (LBNL); Verberkmoes, Nathan C [ORNL; Shah, Manesh B [ORNL; Chavarria, Krystle L. [Lawrence Berkeley National Laboratory (LBNL); Tom, Lauren M [Lawrence Berkeley National Laboratory (LBNL); Brodie, Eoin L. [Lawrence Berkeley National Laboratory (LBNL); Hettich, Robert {Bob} L [ORNL

2010-01-01

We present a novel direct protocol for deep proteome characterization of microorganisms in soil. The method employs thermally assisted detergent-based cellular lysis (SDS) of soil samples, followed by TCA precipitation for proteome extraction/cleanup prior to liquid chromatography-mass spectrometric characterization. This approach was developed and optimized using different soils inoculated with genome-sequenced bacteria (Gram-negative Pseudomonas putida or Gram-positive Arthrobacter chlorophenolicus). Direct soil protein extraction was compared to protein extraction from cells isolated from the soil matrix prior to lysis (indirect method). Each approach resulted in identification of greater than 500 unique proteins, with a wide range in molecular mass and functional categories. To our knowledge, this SDS-TCA approach enables the deepest proteome characterizations of microbes in soil to date, without significant biases in protein size, localization, or functional category compared to pure cultures. This protocol should provide a powerful tool for ecological studies of soil microbial communities.

Description of a double centrifugation tube method for concentrating canine platelets.

Science.gov (United States)

Perazzi, Anna; Busetto, Roberto; Martinello, Tiziana; Drigo, Michele; Pasotto, Daniela; Cian, Francesco; Patruno, Marco; Iacopetti, Ilaria

2013-07-22

To evaluate the efficiency of platelet-rich plasma preparations by means of a double centrifugation tube method to obtain platelet-rich canine plasma at a concentration at least 4 times higher than the baseline value and a concentration of white blood cells not exceeding twice the reference range. A complete blood count was carried out for each sample and each concentrate. Whole blood samples were collected from 12 clinically healthy dogs (consenting blood donors). Blood was processed by a double centrifugation tube method to obtain platelet concentrates, which were then analyzed by a flow cytometry haematology system for haemogram. Platelet concentration and white blood cell count were determined in all samples. Platelet concentration at least 4 times higher than the baseline value and a white blood cell count not exceeding twice the reference range were obtained respectively in 10 cases out of 12 (83.3%) and 11 cases out of 12 (91.6%). This double centrifugation tube method is a relatively simple and inexpensive method for obtaining platelet-rich canine plasma, potentially available for therapeutic use to improve the healing process.

Analysis of anaerobic blood cultures in burned patients.

Science.gov (United States)

Regules, Jason A; Carlson, Misty D; Wolf, Steven E; Murray, Clinton K

2007-08-01

The utility of anaerobic blood culturing is often debated in the general population, but there is limited data on the modern incidence, microbiology, and utility of obtaining routine anaerobic blood cultures for burned patients. We performed a retrospective review of the burned patients electronic medical records database for all blood cultures drawn between January 1997 and September 2005. We assessed blood cultures for positivity, organisms identified, and growth in aerobic or anaerobic media. 85,103 blood culture sets were drawn, with 4059 sets from burned patients. Three hundred and forty-five single species events (619 total blood culture isolates) were noted in 240 burned patients. For burned patients, four isolates were obligate anaerobic bacteria (all Propionibacterium acnes). Anaerobic versus aerobic culture growth was recorded in 310 of 619 (50.1%) burned patient blood culture sets. 46 (13.5%) of the identified organisms, most of which were not obligate anaerobic bacteria, were identified from solely anaerobic media. The results of our study suggest that the detection of significant anaerobic bacteremia in burned patients is very rare and that anaerobic bottles are not needed in this population for that indication. However anaerobic blood cultures systems are also able to detect facultative and obligate aerobic bacteria; therefore, the deletion of the anaerobic culture medium may have deleterious clinical impact.

Effect of solvent/detergent-treated pooled plasma on fibrinolysis in reconstituted whole blood.

Science.gov (United States)

Saadah, Nicholas H; van der Meer, Pieter F; Brinkman, Herm Jan M; de Korte, Dirk; Bontekoe, Ido J; Korsten, Herbert H; Middelburg, Rutger A; van der Bom, Johanna G; Schipperus, Martin R

2017-10-01

Hyperfibrinolysis has been observed in patients heavily transfused with solvent/detergent-treated pooled plasma (S/D plasma). We compared coagulation and fibrinolytic variables in blood containing S/D plasma with blood containing fresh-frozen plasma (FFP), with and without α2-antiplasmin or tranexamic acid (TXA) supplementation. Whole blood samples were reconstituted from red blood cells, platelet (PLT) concentrates, and varying mixtures of FFP and S/D plasma. Hematocrit and PLT count of reconstituted whole blood samples were varied. For a subset of runs, α2-antiplasmin or TXA was added to S/D plasma whole blood samples. Thromboelastography (TEG) analysis was performed to assess 50% clot lysis time (CLT 50% ), maximum amplitude (MA), and initial clotting time (R-time). The change in CLT 50% of whole blood as the plasma compartment transitions from FFP to S/D plasma was -52% (95% confidence interval [CI], -60% to -45%; p plasma in whole blood. α2-Antiplasmin and TXA restored clot lysis time in S/D plasma whole blood. Whole blood with S/D plasma has shorter clot lysis times in vitro compared to whole blood with FFP. α2-Antiplasmin and TXA restore clot lysis time of S/D plasma whole blood to that of FFP whole blood. Clinicians should be aware of the decreased clot lysis time associated with S/D plasma transfusion. © 2017 AABB.

Estimation of the radial force using a disturbance force observer for a magnetically levitated centrifugal blood pump.

Science.gov (United States)

Pai, C N; Shinshi, T; Shimokohbe, A

2010-01-01

Evaluation of the hydraulic forces in a magnetically levitated (maglev) centrifugal blood pump is important from the point of view of the magnetic bearing design. Direct measurement is difficult due to the absence of a rotor shaft, and computational fluid dynamic analysis demands considerable computational resource and time. To solve this problem, disturbance force observers were developed, using the radial controlled magnetic bearing of a centrifugal blood pump, to estimate the radial forces on the maglev impeller. In order to design the disturbance observer, the radial dynamic characteristics of a maglev impeller were evaluated under different working conditions. It was observed that the working fluid affects the additional mass and damping, while the rotational speed affects the damping and stiffness of the maglev system. Based on these results, disturbance force observers were designed and implemented. The designed disturbance force observers present a bandwidth of 45 Hz. In non-pulsatile conditions, the magnitude of the estimated radial thrust increases in proportion to the flowrate, and the rotational speed has little effect on the force direction. At 5 l/min against 100 mmHg, the estimated radial thrust is 0.95 N. In pulsatile conditions, this method was capable of estimating the pulsatile radial thrust with good response.

Blood Culture Test

Science.gov (United States)

... Blood Cultures. Medscape from American Journal of Clinical Pathology [On-line information]. Available online at http://www. ... August 2013. Fisher, M. et. al. (Updated 2013 March 20). Sepsis. ARUP Consult [On-line information]. Available ...

Platelet concentration of plateletrich plasma from dogs, obtained through three centrifugation speeds

Directory of Open Access Journals (Sweden)

Vanessa Couto de Magalhães Ferraz

2007-12-01

Full Text Available The platelets release at least 4 growth factors (Platelet Derived Growth Factor. ²1 and ²2 Transforming Growth Factors and Insulin-like Growth Factor which are responsible for the migration and activation of cells that will start the reparation of soft tissues and bones. The Platelet Rich Plasma is an autogenous source for Growth Factors, obtained by platelet concentration by centrifuging total blood. This study aimed the comparison of platelet concentrations in plasma centrifuged in three different centrifugation speeds (1300, 1600 e 3200rpm, for the production of platelet rich plasma. Blood was drowned from 15 dogs, 40ml of each, and these were divided into four groups and centrifuged at 800rpm. Then the first group was centrifuged at 1300rpm, the second at 1600rpm, the third at 3200rpm and the last was used as control, named plasma. The mean percentage increase in the platelet concentration for each technique was: 1300 - 183%, 1600 - 210% and 3200 - 222%. But in centrifugation at 3200 rpm, platelets presented altered morphology and different sizes in every sample studied, which was understood as severe cell damage. It was concluded that the best technique for the preparation of the platelet rich plasma in dogs consisted of the previous centrifugation of the blood at 800rpm for ten minutes, and then the plasma should be separated. This plasma is then submitted to a second centrifugation of 1600rpm for 10 minutes, and the platelet poor plasma is separated and discharged.

A microfluidic chip for direct and rapid trapping of white blood cells from whole blood

Science.gov (United States)

Chen, Jingdong; Chen, Di; Yuan, Tao; Xie, Yao; Chen, Xiang

2013-01-01

Blood analysis plays a major role in medical and science applications and white blood cells (WBCs) are an important target of analysis. We proposed an integrated microfluidic chip for direct and rapid trapping WBCs from whole blood. The microfluidic chip consists of two basic functional units: a winding channel to mix and arrays of two-layer trapping structures to trap WBCs. Red blood cells (RBCs) were eliminated through moving the winding channel and then WBCs were trapped by the arrays of trapping structures. We fabricated the PDMS (polydimethylsiloxane) chip using soft lithography and determined the critical flow velocities of tartrazine and brilliant blue water mixing and whole blood and red blood cell lysis buffer mixing in the winding channel. They are 0.25 μl/min and 0.05 μl/min, respectively. The critical flow velocity of the whole blood and red blood cell lysis buffer is lower due to larger volume of the RBCs and higher kinematic viscosity of the whole blood. The time taken for complete lysis of whole blood was about 85 s under the flow velocity 0.05 μl/min. The RBCs were lysed completely by mixing and the WBCs were trapped by the trapping structures. The chip trapped about 2.0 × 103 from 3.3 × 103 WBCs. PMID:24404026

[Role of anaerobic blood culture in the simultaneous blood culture taking for the diagnosis of bacteremia].

Science.gov (United States)

Guajardo-Lara, Claudia Elena; Saldaña-Ramírez, Martha Idalia; Ayala-Gaytán, Juan Jacobo; Valdovinos-Chávez, Salvador Bruno

2016-01-01

Harboring a high mortality, the incidence of sepsis is increasing; thus detection, identification and susceptibility tests of the involved microorganisms become urgent. We reviewed the records from January 2013 until July 2014 of a total of 4110 blood culture bottles taken from adult patients in a private tertiary hospital. Growth of microorganisms was observed in 559 bottles (12.6%). We emphasize that 2648 blood cultures (60%) were taken in two paired aerobic and anaerobic bottles drawn at the same time (1324 pairs); from these, growth was observed in 182 inoculated bottles drawn from two different sites at the same time from 135 patients (13.7%). In 86 pairs of bottles with samples from 54 patients (40%), growth occurred only in the aerobic blood culture bottles. Also, growth of microorganisms was observed only in anaerobic bottles in 24 pairs (13.19%), corresponding to 21 patients (15.5%, panaerobic bottle. The usefulness of blood cultures for anaerobes for the identification of obligate anaerobic bacteremia which rarely occur is low (2.2% of patients with bacteremia); however, in 15.55% of the patients the risk of completely overlook bacteremia was present, and in 53% of patients with positive cultures, bacteremia was established earlier, and thus permitted earlier and accurate decision making.

A centrifuge-less plasma separation method from whole blood anticoagulated with EDTA-2K for the use of clinical laboratory tests.

Science.gov (United States)

Tatsumi, N; Tsuda, I; Fukumori, T; Hino, M; Takubo, T; Kondo, H

2001-12-01

In the modern medical laboratory system, simple and rapid processing of specimens are required. In the current system with the transportation line, its centrifugation part would disturb smooth flow of the testing because it needs much time for the centrifugation. To solve the problems, a serum separation method was tried for the whole blood specimen using poly-L-lysine, concanavalin A and phyto-hemoagglutinin. Ploy-L-lysine with molecular weight 130,000 to 210,000 in a final concentration of 0.1% could accelerate blood sedimentation, although its supernatant contaminated platelets. Concanavalin and phytohemoagulutinin could accelerate the sedimentation and obtained plasma, but the method could yield enough amount of supernatant by 1 hour standing. As the purpose of this study is to develop a centrifugeless method, a sieve method using a steel mesh and a magnet was applied to the mixture of EDTA blood, red-cell adhesives and thrombin. The method was unique to separate plasma, but the yield was not so high and chemistry data were not fitted with serum data in some of tests. Thus, the trial would be a new technology, but it was judged that some further improvement will be needed technically.

Solubilization of proteins: the importance of lysis buffer choice.

Science.gov (United States)

Peach, Mandy; Marsh, Noelle; Miskiewicz, Ewa I; MacPhee, Daniel J

2015-01-01

The efficient extraction of proteins of interest from cells and tissues is not always straightforward. Here we demonstrate the differences in extraction of the focal adhesion protein Kindlin-2 from choriocarcinoma cells using NP-40 and RIPA lysis buffer. Furthermore, we demonstrate the use of a more denaturing urea/thiourea lysis buffer for solubilization, by comparing its effectiveness for solubilization of small heat-shock proteins from smooth muscle with the often utilized RIPA lysis buffer. Overall, the results demonstrate the importance of establishing the optimal lysis buffer for specific protein solubilization within the experimental workflow.

STUDIES ON THE BACTERIOPHAGE OF D'HERELLE : IX. EVIDENCE OF HYDROLYSIS OF BACTERIAL PROTEIN DURING LYSIS.

Science.gov (United States)

Hetler, D M; Bronfenbrenner, J

1928-07-31

1. During the process of lysis by bacteriophage, there is an appreciable increase in the amount of free amino acid present in the culture. 2. The increase of free amino acid is due to hydrolysis of bacterial protein.

Pressure-mediated reduction of ultrasonically induced cell lysis

International Nuclear Information System (INIS)

Ciaravino, V.E.; Miller, M.W.; Carstensen, E.L.

1981-01-01

Chinese hamster V-79 cells, exposed in polystyrene tubes for 5 min to 1-MHz continuous-wave ultrasound, were lysed more by a 10 than a 5 W/cm 2 intensity. Higher atmospheric pressure was needed to eliminate lysis with the former relative to the latter intensity, but lysis by 10 W/cm 2 was completely climinated with 2 atm of hydrostatic pressure. The reduction in lysis per unit increase in atmospheric pressure was comparable for both ultrasound intensities

Influence of the centrifuge time of primary plasma tubes on routine coagulation testing.

Science.gov (United States)

Lippi, Giuseppe; Salvagno, Gian Luca; Montagnana, Martina; Manzato, Franco; Guidi, Gian Cesare

2007-07-01

Preparation of blood specimens is a major bottleneck in the laboratory throughput. Reliable strategies for reducing the time required for specimen processing without affecting quality should be acknowledged, especially for laboratories performing stat analyses. The present investigation was planned to establish a minimal suitable centrifuge time for primary samples collected for routine coagulation testing. Five sequential primary vacuum tubes containing 0.109 mol/l buffered trisodium citrate were collected from 10 volunteers and were immediately centrifuged on a conventional centrifuge at 1500 x g, at room temperature for 1, 2, 5, 10 and 15 min, respectively. Hematological and routine coagulation testing, including prothrombin time, activated partial thromboplastin time and fibrinogen, were performed. The centrifugation time was inversely associated with residual blood cell elements in plasma, especially platelets. Statistically significant variations from the reference 15-min centrifuge specimens were observed for fibrinogen in samples centrifuged for 5 min at most and for the activated partial thromboplastin time in samples centrifuged for 2 min at most. Meaningful biases related to the desirable bias were observed for fibrinogen in samples centrifuged for 2 min at most, and for the activated partial thromboplastin time in samples centrifuged for 1 min at most. According to our experimental conditions, a 5-10 min centrifuge time at 1500 x g may be suitable for primary tubes collected for routine coagulation testing.

Rapid identification of bacteria from positive blood culture bottles by use of matrix-assisted laser desorption-ionization time of flight mass spectrometry fingerprinting.

Science.gov (United States)

Christner, Martin; Rohde, Holger; Wolters, Manuel; Sobottka, Ingo; Wegscheider, Karl; Aepfelbacher, Martin

2010-05-01

Early and adequate antimicrobial therapy has been shown to improve the clinical outcome in bloodstream infections (BSI). To provide rapid pathogen identification for targeted treatment, we applied matrix-assisted laser desorption-ionization time of flight (MALDI-TOF) mass spectrometry fingerprinting to bacteria directly recovered from blood culture bottles. A total of 304 aerobic and anaerobic blood cultures, reported positive by a Bactec 9240 system, were subjected in parallel to differential centrifugation with subsequent mass spectrometry fingerprinting and reference identification using established microbiological methods. A representative spectrum of bloodstream pathogens was recovered from 277 samples that grew a single bacterial isolate. Species identification by direct mass spectrometry fingerprinting matched reference identification in 95% of these samples and worked equally well for aerobic and anaerobic culture bottles. Application of commonly used score cutoffs to classify the fingerprinting results led to an identification rate of 87%. Mismatching mostly resulted from insufficient bacterial numbers and preferentially occurred with Gram-positive samples. The respective spectra showed low concordance to database references and were effectively rejected by score thresholds. Spiking experiments and examination of the respective study samples even suggested applicability of the method to mixed cultures. With turnaround times around 100 min, the approach allowed for reliable pathogen identification at the day of blood culture positivity, providing treatment-relevant information within the critical phase of septic illness.

The effect of centrifugation speed and time on pre-analytical platelet activation.

Science.gov (United States)

Söderström, Anna C; Nybo, Mads; Nielsen, Christian; Vinholt, Pernille J

2016-12-01

The results of laboratory analyses are affected by pre-analytical variables, and in particular can platelets be activated by shear handling stress and secrete granular substances. We therefore evaluated the effect of centrifugation speed and time on pre-analytical platelet activation. Citrate- and EDTA-anticoagulated blood from healthy volunteers were centrifuged at 80-10,000 g for 5-15 min to prepare plasma and platelet-rich plasma. Pre-analytical platelet activation was assessed by flow cytometric measurement of platelet P-selectin (CD62p) expression. Blood cell counts, mean platelet volume (MPV), immature platelet fraction (IPF), and platelet distribution width (PDW) were measured. Platelet aggregation in platelet-rich plasma induced by arachidonic acid (AA), ADP or thrombin receptor activator peptide-6 (TRAP) was tested by 96-well aggregometry. The median percentage of platelets expressing P-selectin in citrate- and EDTA-plasma centrifuged at 2000 g for 10 min were 43% [interquartile range (IQR), 38%-53%] and 56% (IQR, 31%-78%), respectively (p=0.82). Platelet-rich plasma prepared at 100-250 g for 10 min had significantly lower platelet P-selectin expression (11%-15%), pcentrifuged. In platelet-rich plasma, increasing centrifugation speed significantly increased platelet yield but decreased contamination from other blood cells, platelet composition was altered as platelet parameters (MPV, IPF, and PDW) was lowered. Platelet aggregation was not affected by the centrifugation speed platelet-rich plasma was prepared. Proportional to centrifugation speed, platelets in plasma and platelet-rich plasma were activated with centrifugation speed, cell content and composition changed while platelet aggregation was unaltered.

Evaluation of the impact of density gradient centrifugation on fetal cell loss during enrichment from maternal peripheral blood.

Science.gov (United States)

Emad, Ahmed; Drouin, Régen

2014-09-01

Physical separation by density gradient centrifugation (DGC) is usually used as an initial step of multistep enrichment protocols for purification of fetal cells (FCs) from maternal blood. Many protocols were designed but no single approach was efficient enough to provide noninvasive prenatal diagnosis. Procedures and methods were difficult to compare because of the nonuniformity of protocols among different groups. Recovery of FCs is jeopardized by their loss during the process of enrichment. Any loss of FCs must be minimized because of the multiplicative effect of each step of the enrichment process. The main objective of this study was to evaluate FC loss caused by DGC. Fetal cells were quantified in peripheral blood samples obtained from both euploid and aneuploid pregnancies before and after enrichment by buoyant DGC using Histopaque 1.119 g/mL. Density gradient centrifugation results in major loss of 60% to 80% of rare FCs, which may further complicate subsequent enrichment procedures. Eliminating aggressive manipulations can significantly minimize FC loss. Data obtained raise questions about the appropriateness of the DGC step for the enrichment of rare FCs and argues for the use of the alternative nonaggressive version of the procedure presented here or prioritizing other methods of enrichments. © 2014 John Wiley & Sons, Ltd.

Rapid fabrication of detachable three-dimensional tissues by layering of cell sheets with heating centrifuge.

Science.gov (United States)

Haraguchi, Yuji; Kagawa, Yuki; Hasegawa, Akiyuki; Kubo, Hirotsugu; Shimizu, Tatsuya

2018-01-18

Confluent cultured cells on a temperature-responsive culture dish can be harvested as an intact cell sheet by decreasing temperature below 32°C. A three-dimensional (3-D) tissue can be fabricated by the layering of cell sheets. A resulting 3-D multilayered cell sheet-tissue on a temperature-responsive culture dish can be also harvested without any damage by only temperature decreasing. For shortening the fabrication time of the 3-D multilayered constructs, we attempted to layer cell sheets on a temperature-responsive culture dish with centrifugation. However, when a cell sheet was attached to the culture surface with a conventional centrifuge at 22-23°C, the cell sheet hardly adhere to the surface due to its noncell adhesiveness. Therefore, in this study, we have developed a heating centrifuge. In centrifugation (55g) at 36-37°C, the cell sheet adhered tightly within 5 min to the dish without significant cell damage. Additionally, centrifugation accelerated the cell sheet-layering process. The heating centrifugation shortened the fabrication time by one-fifth compared to a multilayer tissue fabrication without centrifugation. Furthermore, the multilayered constructs were finally detached from the dishes by decreasing temperature. This rapid tissue-fabrication method will be used as a valuable tool in the field of tissue engineering and regenerative therapy. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 2018. © 2018 American Institute of Chemical Engineers.

Increased Resistance to osmotic lysis of sickled erythrocytes ...

African Journals Online (AJOL)

treated with CNw had significantly reduced osmotic lysis when compared with the untreated set (P<0.05, respectively) at various hypotonic NaCl concentrations. Various Hb genotypes exhibited a graded increase in osmotic pressure lysis in ...

Closed continuous-flow centrifuge rotor

Science.gov (United States)

Breillatt, Jr., Julian P.; Remenyik, Carl J.; Sartory, Walter K.; Thacker, Louis H.; Penland, William Z.

1976-01-01

A blood separation centrifuge rotor having a generally parabolic core disposed concentrically and spaced apart within a housing having a similarly shaped cavity. Blood is introduced through a central inlet and into a central passageway enlarged downwardly to decrease the velocity of the entrant blood. Septa are disposed inside the central passageway to induce rotation of the entrant blood. A separation chamber is defined between the core and the housing wherein the whole blood is separated into red cell, white cell, and plasma zones. The zones are separated by annular splitter blades disposed within the separation chamber. The separated components are continuously removed through conduits communicating through a face seal to the outside of the rotor.

Tumor lysis syndrome in children

International Nuclear Information System (INIS)

Suarez, Amaranto

2004-01-01

Tumor lysis syndrome is a metabolic emergency characterized by electrolyte alteration with or without acute renal failure. It occurs mainly in patients with malignant tumors that have a high growth fraction, or after cytotoxic therapy, as a result of the massive degradation of malignant cells and the release of high amounts of intracellular elements that exceed the capacity of renal excretion. The objective of the treatment is the prevention of nephropathy due to uric acid deposits, and the correction of metabolic acidosis and electrolyte alterations. This paper reviews the incidence, the physiopathology, and the treatment of tumor lysis syndrome in children

Hematocrit analysis through the use of an inexpensive centrifugal polyester-toner device with finger-to-chip blood loading capability

International Nuclear Information System (INIS)

Thompson, Brandon L.; Gilbert, Rachel J.; Mejia, Maximo; Shukla, Nishant; Haverstick, Doris M.; Garner, Gavin T.; Landers, James P.

2016-01-01

Hematocrit (HCT) measurements are important clinical diagnostic variables that help physicians diagnose and treat various medical conditions, ailments, and diseases. In this work, we present the HCT Disc, a centrifugal microdevice fabricated by a Print, Cut and Laminate (PCL) method to generate a 12-sample HCT device from materials costing <0.5 USD (polyester and toner or PeT). Following introduction from a drop of blood (finger stick), whole blood metering and cell sedimentation are controlled by centrifugal force, only requiring a CD player motor as external hardware and, ultimately, a cell phone for detection. The sedimented volume from patient blood in the HCT Disc was analyzed using a conventional scanner/custom algorithm for analysis of the image to determine a hematocrit value, and these were compared to values generated in a clinical laboratory, which correlated well. To enhance portability and assure simplicity of the HCT measurement, values from image analysis by a cell phone using a custom application was compared to the scanner. Fifteen samples were analyzed with cell phone image analysis system and were found to be within 4% of the HCT values determined in the clinical lab. We demonstrate the feasibility of the PeT device for HCT measurement, and highlight its uniquely low cost (<0.5 USD), speed (sample-to-answer <8 min), multiplexability (12 samples), low volume whole blood requirement (<3 μL), rotation speeds (<4000 rpm) needed for effective measurement as well as the direct finger-to-chip sample loading capability. - Highlights: • A 12-sample hematocrit device was developed from polyester-toner materials. • The device can analyze a patient's hematocrit within 8 min from 3 μL of blood. • Cell phone image analysis is used to correctly determine clinical hematocrits.

Hematocrit analysis through the use of an inexpensive centrifugal polyester-toner device with finger-to-chip blood loading capability

Energy Technology Data Exchange (ETDEWEB)

Thompson, Brandon L.; Gilbert, Rachel J. [Department of Chemistry, McCormick Road, University of Virginia, Charlottesville, VA 22904 (United States); Mejia, Maximo [Department of Mechanical Engineering, Engineer' s Way, University of Virginia, Charlottesville, VA 22904 (United States); Shukla, Nishant [Department of Computer Science, Engineer' s Way, University of Virginia, Charlottesville, VA 22904 (United States); Haverstick, Doris M. [Department of Pathology, University of Virginia Health Science Center, Charlottesville, VA 22908 (United States); Garner, Gavin T. [Department of Mechanical Engineering, Engineer' s Way, University of Virginia, Charlottesville, VA 22904 (United States); Landers, James P., E-mail: landers@virginia.edu [Department of Chemistry, McCormick Road, University of Virginia, Charlottesville, VA 22904 (United States); Department of Mechanical Engineering, Engineer' s Way, University of Virginia, Charlottesville, VA 22904 (United States); Department of Pathology, University of Virginia Health Science Center, Charlottesville, VA 22908 (United States)

2016-06-14

Hematocrit (HCT) measurements are important clinical diagnostic variables that help physicians diagnose and treat various medical conditions, ailments, and diseases. In this work, we present the HCT Disc, a centrifugal microdevice fabricated by a Print, Cut and Laminate (PCL) method to generate a 12-sample HCT device from materials costing <0.5 USD (polyester and toner or PeT). Following introduction from a drop of blood (finger stick), whole blood metering and cell sedimentation are controlled by centrifugal force, only requiring a CD player motor as external hardware and, ultimately, a cell phone for detection. The sedimented volume from patient blood in the HCT Disc was analyzed using a conventional scanner/custom algorithm for analysis of the image to determine a hematocrit value, and these were compared to values generated in a clinical laboratory, which correlated well. To enhance portability and assure simplicity of the HCT measurement, values from image analysis by a cell phone using a custom application was compared to the scanner. Fifteen samples were analyzed with cell phone image analysis system and were found to be within 4% of the HCT values determined in the clinical lab. We demonstrate the feasibility of the PeT device for HCT measurement, and highlight its uniquely low cost (<0.5 USD), speed (sample-to-answer <8 min), multiplexability (12 samples), low volume whole blood requirement (<3 μL), rotation speeds (<4000 rpm) needed for effective measurement as well as the direct finger-to-chip sample loading capability. - Highlights: • A 12-sample hematocrit device was developed from polyester-toner materials. • The device can analyze a patient's hematocrit within 8 min from 3 μL of blood. • Cell phone image analysis is used to correctly determine clinical hematocrits.

Lysis of fresh human solid tumors by autologous lymphocytes activated in vitro with lectins

International Nuclear Information System (INIS)

Mazumder, A.; Grimm, E.A.; Zhang, H.Z.; Rosenberg, S.A.

1982-01-01

Human peripheral blood lymphocytes (PBL), obtained from patients with a variety of cancers, were incubated in vitro with phytohemagglutinin, concanavalin A, and crude or lectin-free T-cell growth factors. The lectin-activated PBL of nine patients were capable of lysing fresh autologous tumor during a 4-hr 51Cr release assay. Multiple metastases from the same patient were equivalently lysed by these activated autologous PBL. No lysis of fresh PBL or lectin-induced lymphoblast cell targets was seen, although tumor, PBL, and lymphoblast cells were shown to be equally lysable using allosensitized cells. The activated cells could be expanded without loss of cytotoxicity in crude or lectin-free T-cell growth factors. The generation of cells lytic to fresh autologous tumor was dependent on the presence of adherent cells, although the lytic cell itself was not adherent. Proliferation was not involved in the induction of lytic cells since equal lysis was induced in irradiated and nonirradiated lymphocytes. Lectin was not required in the lytic assay, and the addition of alpha-methyl-D-mannoside to concanavalin A-activated lymphoid cells did not increase the lysis of fresh tumor cells. Activation by lectin for 3 days appears to be an efficient and convenient method for generating human cells lytic to fresh autologous tumor. These lytic cells may be of value for studies of the cell-mediated lysis of human tumor and possibly for tumor immunotherapy as well

Theoretical prediction and validation of cell recovery rates in preparing platelet-rich plasma through a centrifugation.

Science.gov (United States)

Piao, Linfeng; Park, Hyungmin; Jo, Chris Hyunchul

2017-01-01

In the present study, we propose a theoretical framework to predict the recovery rates of platelets and white blood cells in the process of centrifugal separation of whole blood contained in a tube for the preparation of platelet-rich plasma. Compared to previous efforts to optimize or standardize the protocols of centrifugation, we try to further the physical background (i.e., based on the multiphase flow phenomena) of analysis to develop a universal approach that can be applied to widely different conditions. That is, one-dimensional quasi-linear partial differential equation to describe the centrifugal sedimentation of dispersed phase (red and white blood cells) in continuous phase (plasma) is derived based on the kinematic-wave theory. With the information of whole blood volume and tube geometry considered, it is possible to determine the positions of interfaces between supernatant/suspension and suspension/sediment, i.e., the particle concentration gradient in a tube, for a wide range of centrifugation parameters (time and acceleration). While establishing a theory to predict the recovery rates of the platelet and white blood cell from the pre-determined interface positions, we also propose a new correlation model between the recovery rates of plasma and platelets, which is found to be a function of the whole blood volume, centrifugal time and acceleration, and tube geometry. The present predictions for optimal condition show good agreements with available human clinical data, obtained from different conditions, indicating the universal applicability of our method. Furthermore, the dependence of recovery rates on centrifugal conditions reveals that there exist a different critical acceleration and time for the maximum recovery rate of platelets and white blood cells, respectively. The other parameters such as hematocrit, whole blood volume and tube geometry are also found to strongly affect the maximum recovery rates of blood cells, and finally, as a strategy

Chemical Cell Lysis System Applicable to Lab-on-a-Disc.

Science.gov (United States)

Lim, Dayeseul; Yoo, Jae Chern

2017-09-01

The design and fabrication of a heating system has been a significant challenge in implementing chemical lysis on a lab-on-a-disc (LOD). The proposed system contains a sample inlet, phase change material (PCM) array, heating chamber, and valve in a single disc, providing cost-effective, rapid, and fully automated chemical cell lysis. Compared to the conventional cell lysis system, our cell lysis system has many advantages, such as a compact structure that is easily integrated into the LOD and reduced processing time and labor. The experiments are conducted with Salmonella typhimurium strains to demonstrate the performance. The experimental results show that the proposed approach is greatly effective in realizing a chemical cell lysis system on an LOD with higher throughput in terms of purity and yield of DNA.

Urea enhances cell lysis of Schizosaccharomyces pombe ura4 mutants.

Science.gov (United States)

Nishino, Kohei; Kushima, Misaki; Kaino, Tomohiro; Matsuo, Yasuhiro; Kawamukai, Makoto

2017-07-01

Cell lysis is induced in Schizosaccharomyces pombe ∆ura4 cells grown in YPD medium, which contains yeast extract, polypeptone, and glucose. To identify the medium components that induce cell lysis, we first tested various kinds of yeast extracts from different suppliers. Cell lysis of ∆ura4 cells on YE medium was observed when yeast extracts from OXOID, BD, Oriental, and Difco were used, but not when using yeast extract from Kyokuto. To determine which compounds induced cell lysis, we subjected yeast extract and polypeptone to GC-MS analysis. Ten kinds of compounds were detected in OXOID and BD yeast extracts, but not in Kyokuto yeast extract. Among them was urea, which was also present in polypeptone, and it clearly induced cell lysis. Deletion of the ure2 gene, which is responsible for utilizing urea, abolished the lytic effect of urea. The effect of urea was suppressed by deletion of pub1, and a similar phenotype was observed in the presence of polypeptone. Thus, urea is an inducer of cell lysis in S. pombe ∆ura4 cells.

Observations on centrifugation: application to centrifuge development.

Science.gov (United States)

Roberts, T; Smith, M; Roberts, B

1999-11-01

This report outlines the background to the development of an automated, serial, discrete centrifuge, reporting on the criteria considered essential in such an instrument. We established the criteria by examining the detailed logistics of centrifuge operation in a hospital laboratory. The mean sample load per run, using six centrifuges, was 13.6 samples, and the user-selectable cycle time ranged from 00:01:10 to 00:12:33 (hours:minutes:seconds) with a fixed g value of 1050. During the laboratory working window, (0900-1700), only 50% of the centrifuge capacity was utilized and more than one-third of the sample workload was delayed for >5 min because the centrifuges were not emptied promptly. In addition, 35% of the sample workload was centrifuged for less than the time prescribed in the operational specifications. Based on these findings, we designed a new continuous, serial centrifuge to overcome some of the deficiencies noted in the logistics study. The centrifuge operates continuously, nominally treating 150 samples/h, with a cycle time of 5 min at 1,000 g. The cycle time and g value are variable between limits, and their selection governs the throughput rate. Each sample is centrifuged separately in individual rotors mounted in a sturdy carousel with a periphery that traverses a load/unload station. There is no sample delay because of operator absence, and the capacity is fully utilized. The centrifuge can operate in a stand-alone capacity or has the capability of being integrated into a sample preparation system or as a direct front end for high-throughput analyzers.

Isolating peripheral lymphocytes by density gradient centrifugation and magnetic cell sorting.

Science.gov (United States)

Brosseron, Frederic; Marcus, Katrin; May, Caroline

2015-01-01

Combining density gradient centrifugation with magnetic cell sorting provides a powerful tool to isolate blood cells with high reproducibility, yield, and purity. It also allows for subsequent separation of multiple cell types, resulting in the possibility to analyze different purified fractions from one donor's sample. The centrifugation step divides whole blood into peripheral blood mononuclear cells (PBMC), erythrocytes, and platelet-rich plasma. In the following, lymphocyte subtypes can be consecutively isolated from the PBMC fraction. This chapter describes enrichment of erythrocytes, CD14-positive monocytes and CD3-positive T lymphocytes. Alternatively, other cell types can be targeted by using magnetic beads specific for the desired subpopulation.

Effect of Centrifuge Temperature on Routine Coagulation Tests.

Science.gov (United States)

Yazar, Hayrullah; Özdemir, Fatma; Köse, Elif

2018-01-01

This study investigated the effects of cooled and standard centrifuges on the results of coagulation tests to examine the effects of centrifugation temperature. Equal-volume blood samples from each patient were collected at the same time intervals and subjected to standard (25°C) and cooled centrifugation (2-4°C). Subsequently, the prothrombin time (PT), international normalized ratio (INR), activated partial thromboplastin time (aPTT), fibrinogen, and D-dimer values were determined in runs with the same lot numbers in the same coagulation device using the Dia-PT R (PT and INR), Dia-PTT-liquid (aPTT), Dia-FIB (fibrinogen), and Dia-D-dimer kits, respectively. The study enrolled 771 participants. The PT was significantly (p centrifuges were as follows: PT 10.30 versus 10.50 s; PT (INR) 1.04 versus 1.09 s; APTT 28.90 versus 29.40 s; fibrinogen 321.5 versus 322.1 mg/dL; and D-dimer 179.5 versus 168.7 µg FEU/mL. There were significant differences (p centrifuges. Centrifuge temperature can have a significant effect on the results of coagulation tests. However, broad and specific disease-based studies are needed. © 2018 S. Karger AG, Basel.

Design optimization of flow channel and performance analysis for a new-type centrifugal blood pump

Science.gov (United States)

Ji, J. J.; Luo, X. W.; Y Wu, Q.

2013-12-01

In this paper, a new-type centrifugal blood pump, whose impeller is suspended inside a pump chamber with hydraulic bearings, is presented. In order to improve the hydraulic performance of the pump, an internal flow simulation is conducted to compare the effects of different geometrical parameters of pump flow passage. Based on the numerical results, the pumps can satisfy the operation parameters and be free of hemolysis. It is noted that for the pump with a column-type supporter at its inlet, the pump head and hydraulic efficiency decreases compared to the pump with a step-type support structure. The performance drop is caused by the disturbed flow upstream impeller inlet. Further, the unfavorable flow features such as reverse flow and low velocity in the pump may increases the possibility of thrombus. It is also confirmed that the casing shape can little influence pump performance. Those results are helpful for design optimization in blood pump development.

Reagentless mechanical cell lysis by nanoscale barbs in microchannels for sample preparation.

Science.gov (United States)

Di Carlo, Dino; Jeong, Ki-Hun; Lee, Luke P

2003-11-01

A highly effective, reagentless, mechanical cell lysis device integrated in microfluidic channels is reported. Sample preparation, specifically cell lysis, is a critical element in 'lab-on-chip' applications. However, traditional methods of cell lysis require purification steps or complicated fabrication steps that a simple mechanical method of lysis may avoid. A simple and effective mechanical cell lysis system is designed, microfabricated, and characterized to quantify the efficiency of cell lysis and biomolecule accessibility. The device functionality is based on a microfluidic filter region with nanostructured barbs created using a modified deep reactive ion etching process. Mechanical lysis is characterized by using a membrane impermeable dye. Three main mechanisms of micro-mechanical lysis are described. Quantitative measurements of accessible protein as compared to a chemically lysed sample are acquired with optical absorption measurements at 280 and 414 nm. At a flow rate of 300 microL min(-1) within the filter region total protein and hemoglobin accessibilities of 4.8% and 7.5% are observed respectively as compared to 1.9% and 3.2% for a filter without nanostructured barbs.

Purification of Proteins From Cell-Culture Medium or Cell-Lysate by High-Speed Counter-Current Chromatography Using Cross-Axis Coil Planet Centrifuge

Science.gov (United States)

Shibusawa, Yoichi; Ito, Yoichiro

2014-01-01

This review describes protein purifications from cell culture medium or cell-lysate by high speed counter-current chromatography using the cross-axis coil planet centrifuge. Purifications were performed using aqueous two phase systems composed of polyethylene glycols and dextrans. PMID:25360182

Simulated stand tests and centrifuge training to prevent orthostatic intolerance on Earth, moon, and Mars.

Science.gov (United States)

Coats, Brandon W; Sharp, M Keith

2010-03-01

One proposed method to overcome postflight orthostatic intolerance is for astronauts to undergo inflight centrifugation. Cardiovascular responses were compared between centrifuge and gravitational conditions using a seven-compartment cardiovascular model. Vascular resistance, heart rate, and stroke volume values were adopted from literature, while compartmental volumes and compliances were derived from impedance plethysmography of subjects (n=8) riding on a centrifuge. Three different models were developed to represent the typical male subject who completed a 10-min postflight stand test ("male finisher"), "non-finishing male" and "female" (all non-finishers). A sensitivity analysis found that both cardiac output and arterial pressure were most sensitive to total blood volume. Simulated stand tests showed that female astronauts were more susceptible to orthostatic intolerance due to lower initial blood pressure and higher pressure threshold for presyncope. Rates of blood volume loss by capillary filtration were found to be equivalent in female and male non-finishers, but four times smaller in male finishers. For equivalent times to presyncope during centrifugation as those during constant gravity, lower G forces at the level of the heart were required. Centrifuge G levels to match other cardiovascular parameters varied depending on the parameter, centrifuge arm length, and the gravity level being matched.

Optimization of Power Consumption for Centrifugation Process Based on Attenuation Measurements

Science.gov (United States)

Salim, M. S.; Abd Malek, M. F.; Sabri, Naseer; Omar, M. Iqbal bin; Mohamed, Latifah; Juni, K. M.

2013-04-01

The main objective of this research is to produce a mathematical model that allows decreasing the electrical power consumption of centrifugation process based on attenuation measurements. The centrifugation time for desired separation efficiency may be measured to determine the power consumed of laboratory centrifuge device. The power consumption is one of several parameters that affect the system reliability and productivity. Attenuation measurements of wave propagated through blood sample during centrifugation process were used indirectly to measure the power consumption of device. A mathematical model for power consumption was derived and used to modify the speed profile of centrifuge controller. The power consumption model derived based on attenuation measurements has successfully save the power consumption of centrifugation process keeping high separation efficiency. 18kW.h monthly for 100 daily time device operation had been saved using the proposed model.

Optimization of Power Consumption for Centrifugation Process Based on Attenuation Measurements

International Nuclear Information System (INIS)

Salim, M S; Iqbal bin Omar, M; Malek, M F Abd; Mohamed, Latifah; Sabri, Naseer; Juni, K M

2013-01-01

The main objective of this research is to produce a mathematical model that allows decreasing the electrical power consumption of centrifugation process based on attenuation measurements. The centrifugation time for desired separation efficiency may be measured to determine the power consumed of laboratory centrifuge device. The power consumption is one of several parameters that affect the system reliability and productivity. Attenuation measurements of wave propagated through blood sample during centrifugation process were used indirectly to measure the power consumption of device. A mathematical model for power consumption was derived and used to modify the speed profile of centrifuge controller. The power consumption model derived based on attenuation measurements has successfully save the power consumption of centrifugation process keeping high separation efficiency. 18kW.h monthly for 100 daily time device operation had been saved using the proposed model.

21 CFR 864.7275 - Euglobulin lysis time tests.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Euglobulin lysis time tests. 864.7275 Section 864.7275 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7275 Euglobulin lysis...

An implantable centrifugal blood pump with a recirculating purge system (Cool-Seal system).

Science.gov (United States)

Yamazaki, K; Litwak, P; Tagusari, O; Mori, T; Kono, K; Kameneva, M; Watach, M; Gordon, L; Miyagishima, M; Tomioka, J; Umezu, M; Outa, E; Antaki, J F; Kormos, R L; Koyanagi, H; Griffith, B P

1998-06-01

A compact centrifugal blood pump has been developed as an implantable left ventricular assist system. The impeller diameter is 40 mm, and pump dimensions are 55 x 64 mm. This first prototype, fabricated from titanium alloy, resulted in a pump weight of 400 g including a brushless DC motor. The weight of a second prototype pump was reduced to 280 g. The entire blood contacting surface is coated with diamond like carbon (DLC) to improve blood compatibility. Flow rates of over 7 L/min against 100 mm Hg pressure at 2,500 rpm with 9 W total power consumption have been measured. A newly designed mechanical seal with a recirculating purge system (Cool-Seal) is used for the shaft seal. In this seal system, the seal temperature is kept under 40 degrees C to prevent heat denaturation of blood proteins. Purge fluid also cools the pump motor coil and journal bearing. Purge fluid is continuously purified and sterilized by an ultrafiltration unit which is incorporated in the paracorporeal drive console. In vitro experiments with bovine blood demonstrated an acceptably low hemolysis rate (normalized index of hemolysis = 0.005 +/- 0.002 g/100 L). In vivo experiments are currently ongoing using calves. Via left thoracotomy, left ventricular (LV) apex descending aorta bypass was performed utilizing an expanded polytetrafluoroethylene (ePTFE) vascular graft with the pump placed in the left thoracic cavity. In 2 in vivo experiments, the pump flow rate was maintained at 5-9 L/min, and pump power consumption remained stable at 9-10 W. All plasma free Hb levels were measured at less than 15 mg/dl. The seal system has demonstrated good seal capability with negligible purge fluid consumption (<0.5 ml/day). In both calves, the pumps demonstrated trouble free continuous function over 6 month (200 days and 222 days).

Erythrocyte sedimentation rate and fibrinogen concentration of whole blood influences the cellular composition of platelet-rich plasma obtained from centrifugation methods.

Science.gov (United States)

Yin, Wenjing; Xu, Zhengliang; Sheng, Jiagen; Xie, Xuetao; Zhang, Changqing

2017-09-01

Erythrocyte sedimentation rate (ESR), which reflects the sedimentation rate of platelets, leukocytes and erythrocytes in response to centrifugal force, may influence the cellular composition of platelet-rich plasma (PRP) obtained via centrifugation methods. However, no relevant studies have substantiated this. In the present study, blood was collected from 40 healthy volunteers and used to prepare PRP with two plasma-based preparation systems [YinPRP and Plasma Rich in Growth Factor (PRGF) systems] and two buffy coat-based systems (RegenPRP and WEGOPRP systems) in a single-donor model. Volumes of PRP and platelet-poor plasma (PPP) that were removed in the preparation process were recorded. Analyses of ESR, haematocrit, C-reaction protein, coagulation, serum glucose and serum lipid of the whole blood used for PRP preparation were performed to evaluate the levels of ESR and the factors known to influence it. Whole blood analysis was performed to evaluate the cellular composition of PRP. Results demonstrated that there were marked positive correlations between the ESR of the whole blood used for PRP preparation and PPP removal efficiencies, platelet concentrations, platelet capture efficiencies and platelet enrichment factors of PRP formulations obtained from plasma-based systems, and PRP yield efficiency of RegenPRP and PPP removal efficiency of WEGOPRP. Furthermore, there were marked negative correlations between ESR and concentrations and enrichment factors of platelets, leukocytes and erythrocytes of RegenPRP. Fibrinogen concentration of the whole blood, which had a marked positive correlation with ESR, also influenced the cellular composition of PRP. These findings may increase the understanding of PRP preparation and provide substantial evidence for the individualised optimisation of PRP preparation systems used in clinical practice.

Assessment of centrifugation using for accelerated immunological microarray analysis for blood cells investigation

Directory of Open Access Journals (Sweden)

A. V. Shishkin

2011-01-01

Full Text Available Phase of incubation microarray with cell suspension is prolonged when cells are investigated. It takes from 20 to 60 min if cell sedimentation on the surface of microarray is the result of gravity . Decrease of this stage duration is possible due to centrifugation. In th is article influence of centrifugation on results of analysis is considered. Changes of morphological description of cells are estimated when they a re precipitatedwith different acceleration. Also availability of centrifugation using when it is necessary to obtain the high density of cell binding in test regions of microarray if cells concentration in sample is small is demonstrated.

CENTRIFUGE APPARATUS

Science.gov (United States)

Skarstrom, C.; Urey, H.C.; Cohen, K.

1960-08-01

A high-speed centrifuge for the separation of gaseous isotopes is designed comprising a centrifugal pump mounted on the outlet of a centrifuge bowl and arranged to pump the heavy and light fractions out of the centrifuge bowl in two separate streams.

Radiation-induced enhancement of enzymatic cell lysis of Micrococcus radiodurans

International Nuclear Information System (INIS)

Watanabe, H.; Takehisa, M.; Iizuka, H.

1981-01-01

The intact cells of M. radiodurans were rendered sensitive to the action of lytic enzyme (P2-2 enzyme) by irradiation. The radiation-induced enhancement of cell lysis with P2-2 enzyme was completely prevented by the addition of t-butanol and irradiation at liquid nitrogen temperature. These results indicate that the enhancement is due to indirect action resulting from OH radicals. Cell lysis by lysozyme was enhanced only when the cells were irradiated under N 2 O. The enhancement of cell lysis with lysozyme was also prevented by adding alcohols. On the other hand, when lipid components in cells were removed by extraction with n-butanol, the radiation-induced enhancement of cell lysis with P2-2 enzyme and lysozyme was not observed. From these results it is concluded that the enhancement of enzymatic cell lysis by irradiation is attributable to alteration in the lipid-rich layer of the cell wall caused by OH radicals

Radiation-induced enhancement of enzymatic cell lysis of Micrococcus radiodurans

Energy Technology Data Exchange (ETDEWEB)

Watanabe, H.; Takehisa, M. [Japan Atomic Energy Research Inst., Takasaki, Gunma, Takasaki Radiation Chemistry Research Establishment (Japan); Iizuka, H.

1981-10-15

The intact cells of M. radiodurans were rendered sensitive to the action of lytic enzyme (P2-2 enzyme) by irradiation. The radiation-induced enhancement of cell lysis with P2-2 enzyme was completely prevented by the addition of t-butanol and irradiation at liquid nitrogen temperature. These results indicate that the enhancement is due to indirect action resulting from OH radicals. Cell lysis by lysozyme was enhanced only when the cells were irradiated under N{sub 2}O. The enhancement of cell lysis with lysozyme was also prevented by adding alcohols. On the other hand, when lipid components in cells were removed by extraction with n-butanol, the radiation-induced enhancement of cell lysis with P2-2 enzyme and lysozyme was not observed. From these results it is concluded that the enhancement of enzymatic cell lysis by irradiation is attributable to alteration in the lipid-rich layer of the cell wall caused by OH radicals.

Microvascular pressure responses of second-generation rats chronically exposed to 2 g centrifugation

Science.gov (United States)

Richardson, D. R.; Knapp, C. F.

1977-01-01

Preliminary results are presented for a study aimed at a quantitative comparison of microvascular dynamics in second-generation rats reared in a 2-g force field produced by centrifugation with similar data from animals reared in a centrifuge that produced only a 1-g force. It is shown that the pressure distribution in the mesenteric microvasculature of the second generation of rats reared in a 2-g environment, as well as the animals' blood pressure response to epinephrine, are significantly different compared to their 1-g counterparts. In particular, 1-g and 2-g chronic centrifugation enhances the arterial blood pressure, and the 2-g force field attenuates the pressor effects of norepinephrine.

Two small lymphocyte subpopulations in human peripheral blood. I. Purification and surface marker profiles

DEFF Research Database (Denmark)

Hokland, M; Hokland, P; Heron, I

1978-01-01

By means of simple rosette sedimentation methods two subsets from human peripheral blood lymphocytes have been isolated: (1) (E, Fc)- and (2) (E, Ig)-. The first subset was obtained by centrifuging suspensions of macrophage-depleted PBL in which E and EA rosettes had been allowed to form simultan......By means of simple rosette sedimentation methods two subsets from human peripheral blood lymphocytes have been isolated: (1) (E, Fc)- and (2) (E, Ig)-. The first subset was obtained by centrifuging suspensions of macrophage-depleted PBL in which E and EA rosettes had been allowed to form...... simultaneously. The dominant marker of these E- Fc- cells was surface Ig, and during 4 days of culture this population did not alter its surface markers. Subset 2 was obtained in two ways following rosette centrifugation with AET-treated SRBC and rabbit anti-human Ig-coated autologous RBC. This 'Null cell...

The ameliorative effects of Moringa oleifera leaf extract on ...

African Journals Online (AJOL)

ADEYEYE

2017-03-13

Mar 13, 2017 ... aromatic hydrocarbons. It is commonly used as fuel for ... resistance of red blood cells to lysis by osmotic stress. (Oyewale et al., 2011; ... electrode clips for right arm, left arm, right leg, left leg and the heart ... rpm for 10 minutes using a bench centrifuge (model .... Review on candesartan: Pharmacological.

Blood culture results from healthy captive and free-ranging elasmobranchs.

Science.gov (United States)

Mylniczenko, Natalie D; Harris, Brigita; Wilborn, Rachel E; Young, Forrest A

2007-09-01

Blood culture is a diagnostic tool used in confirming bacterial disease in teleostean and elasmobranch fishes. Unlike teleosts, elasmobranchs have a normal microflora in multiple organs, but their blood has generally been considered to be sterile. In regular exams of elasmobranchs conducted at a public aquarium, occasional blood samples have tested positive on culture. This finding prompted a blood culture survey of healthy captive and wild elasmobranchs (sharks and stingrays), which showed that 26.7% of all animals were positive. Stingrays alone showed a 50% occurrence of positive blood cultures, although the total number of animals was low and freshwater species were included in this number. When elasmobranchs other than stingrays were evaluated according to metabolic category, pelagic animals had a higher percentage of positive cultures than nonpelagic animals (38.7% versus 13.9%). These results indicate that a single positive blood culture without other corroborating diagnostics is not sufficient to confirm septicemia in elasmobranchs.

The Generation of Human Induced Pluripotent Stem Cells from Blood Cells: An Efficient Protocol Using Serial Plating of Reprogrammed Cells by Centrifugation

Directory of Open Access Journals (Sweden)

Youngkyun Kim

2016-01-01

Full Text Available Human induced pluripotent stem cells (hiPSCs have demonstrated great potential for differentiation into diverse tissues. We report a straightforward and highly efficient method for the generation of iPSCs from PBMCs. By plating the cells serially to a newly coated plate by centrifugation, this protocol provides multiple healthy iPSC colonies even from a small number of PBMCs. The generated iPSCs expressed pluripotent markers and differentiated into all three germ layer lineages. The protocol can also be used with umbilical cord blood mononuclear cells (CBMCs. In this study, we present a simple and efficient protocol that improved the yield of iPSCs from floating cells such as PBMCs and CBMCs by serial plating and centrifugation.

A simple and novel modification of comet assay for determination of bacteriophage mediated bacterial cell lysis.

Science.gov (United States)

Khairnar, Krishna; Sanmukh, Swapnil; Chandekar, Rajshree; Paunikar, Waman

2014-07-01

The comet assay is the widely used method for in vitro toxicity testing which is also an alternative to the use of animal models for in vivo testing. Since, its inception in 1984 by Ostling and Johansson, it is being modified frequently for a wide range of application. In spite of its wide applicability, unfortunately there is no report of its application in bacteriophages research. In this study, a novel application of comet assay for the detection of bacteriophage mediated bacterial cell lysis was described. The conventional methods in bacteriophage research for studying bacterial lysis by bacteriophages are plaque assay method. It is time consuming, laborious and costly. The lytic activity of bacteriophage devours the bacterial cell which results in the release of bacterial genomic material that gets detected by ethidium bromide staining method by the comet assay protocol. The objective of this study was to compare efficacy of comet assay with different assay used to study phage mediated bacterial lysis. The assay was performed on culture isolates (N=80 studies), modified comet assay appear to have relatively higher sensitivity and specificity than other assay. The results of the study showed that the application of comet assay can be an economical, time saving and less laborious alternative to conventional plaque assay for the detection of bacteriophage mediated bacterial cell lysis. Copyright © 2014 Elsevier B.V. All rights reserved.

Shear Evaluation by Quantitative Flow Visualization Near the Casing Surface of a Centrifugal Blood Pump

Science.gov (United States)

Nishida, Masahiro; Yamane, Takashi; Tsukamoto, Yuki; Ito, Kazuyuki; Konishi, Yoshiaki; Masuzawa, Toru; Tsukiya, Tomonori; Endo, Seiko; Taenaka, Yoshiyuki

To clarify the correlation between high-shear flow and hemolysis in blood pumps, detail shear velocity distribution was quantified by an experimental method with a model centrifugal blood pump that has a series data of hemolysis tests and computational fluid dynamic analyses. Particular attention was paid to the shear velocity near the casing surface in the volute where the high shear causes in circumferentially wide region that is considerable to cause high hemolysis. Three pump models were compared concern with the radial gap width between the impeller and casing (the radial volute width) also with the outlet position whereas the impeller geometry was identical. These casing geometries were as follows: model 1-the gap width is standard 3mm and the outlet locates to make a smooth geometrical connection with the volute, model 2-the gap width is small 0.5mm and the outlet locates to make the smooth geometrical connection with the volute, and model 3-the gap width is small 0.5mm and the outlet locates to hardly make the smooth geometrical connection with the volute but be similar radial position with that of model 1. Velocity was quantified with a particle tracking velocimetry that is one of the quantitative flow visualization techniques, and the shear velocity was calculated. Results showed that all large shear velocity existed within the layers of about 0.1mm from the casing surface and that those layers were hardly affected by a vane passage even if the gap width is 0.5mm. They also showed that the maximum shear velocity appeared on the casing surface, and the shear velocities of models 2 and 3 were almost twice as large as that of model 1. This finding is in full corresponding with the results of hemolysis tests which showed that the hemolysis levels of both models 2 and 3 were 1.5 times higher than that of model 1. These results suggest that detailed high-shear evaluation near the casing surface in the volute is one of the most important keys in estimating the

Squamous cell carcinoma complicating an hereditary epidermo-lysis bullosa

International Nuclear Information System (INIS)

Mseddi, M.; Turki, H.; Marrekchi, S.; Abdelmaksoud, W.; Masmoudi, A.; Bouassida, S.; Zahaf, A.

2004-01-01

The dystrophic form of hereditary epidermo-lysis bullosa is associated with an increased frequency of squamous cell carcinoma. We report a new case. An 18-year-old patient, carrying a Hallopeau Siemens hereditary epidermo-lysis bullosa, presented a subcutaneous nodular lesion, for 1 year that ulcerated and budded with inguinal lymphadenopathy. The histological study ted to the conclusion of a well differentiated squamous cell carcinoma. The patient was treated surgically. Tumor and metastatic lymph nodes were excised. A radiotherapy was decided but the postoperative course was fatal due to an infection and to a deterioration of her general condition. Squamous cell carcinoma frequently occurs on the cicatricial lesion of hereditary epidermo-lysis bullosa and usually affects males with recessive hereditary epidermo-lysis bullosa. Metastases are frequent, precocious and multiple. The treatment may be surgical. The particularities of our observation are the young age of patient and the localization. (author)

Evaluation of a direct method for the identification and antibiotic susceptibility assessment of microrganisms isolated from blood cultures by automatic systems

Directory of Open Access Journals (Sweden)

Sergio Frugoni

2008-03-01

Full Text Available The purpose of blood cultures in the septic patient is to address a correct therapeutic approach. Identification and antibiotic susceptibility test carried out directly from the bottle may give important information in short time.The introduction of the automatic instrumentation has improved the discovering of pathogens in the blood, however the elapsing time between the positive detection and the microbiological report is still along. Is the evaluation of this study a fast, easy, cheap method to be applied to the routine, which could reduce the response time in the bacteraemia diagnosis.The automatic systems Vitek Senior (bioMérieux, and Vitek 2 (bioMérieux were used at Pio Albergo Trivulzio (Centre1 and at Istituto dei Tumori (Centre2 respectivetly.To remove blood cells, 7 ml. of the culture has been moved by vacuum sampling in a test tube and centrifuged for 10 minutes at 1000 rpm the supernatant has been further centrifuged for 10 minutes at 3000 rpm.0.5 ml. of BHI has been added to the pellet o sediment.The concentration of bacterial suspension has been fit for the inoculation. At the same time has been prepared standard cultures in suitable culture media were carried out for comparison. In the centro1 and centro2 have been isolated and identify respectively 63 and 31 Gram negative, and, 32 and 40 gram positive microorganisms have been isolated and identify in the Centre1 and Centre2 respectively.The identification Gram-negative and Gram positive microorganisms showed an agreement of 100% and 86.2% and 93.3% and 65.78% respectively between the direct and the standard method. For antibiotic susceptibility tests, 903 (Centre1 and 491 (Centre2 and 396 and 509 compounds were totally assessed in Gram negative and Gram positive bacteria respectively.The analysis has highlighted that: Centre1 has reported 0.30% very major errors (GE, 0.92% major errors (EM, 1.23% minor errors (Em. Centre 2 showed 0.57% very major errors (GE, 0.09% major errors

Lysis from without

Science.gov (United States)

2011-01-01

In this commentary I consider use of the term “lysis from without” (LO) along with the phenomenon's biological relevance. LO originally described an early bacterial lysis induced by high-multiplicity virion adsorption and that occurs without phage production (here indicated as LOV). Notably, this is more than just high phage multiplicities of adsorption leading to bacterial killing. The action on bacteria of exogenously supplied phage lysin, too, has been described as a form of LO (here, LOL). LOV has been somewhat worked out mechanistically for T4 phages, has been used to elucidate various phage-associated phenomena including discovery of the phage eclipse, may be relevant to phage ecology, and, with resistance to LO (LOR), is blocked by certain phage gene products. Speculation as to the impact of LOV on phage therapy also is fairly common. Since LOV assays are relatively easily performed and not all phages are able to induce LOV, a phage's potential to lyse bacteria without first infecting should be subject to at least in vitro experimental confirmation before the LOV label is applied. The term “abortive infection” may be used more generally to describe non-productive phage infections that kill bacteria. PMID:21687534

Centrifugation assay for measuring adhesion of serially passaged bovine chondrocytes to polystyrene surfaces.

Science.gov (United States)

Kaplan, David S; Hitchins, Victoria M; Vegella, Thomas J; Malinauskas, Richard A; Ferlin, Kimberly M; Fisher, John P; Frondoza, Carmelita G

2012-07-01

A major obstacle in chondrocyte-based therapy for cartilage repair is the limited availability of cells that maintain their original phenotype. Propagation of chondrocytes as monolayer cultures on polystyrene surfaces is used extensively for amplifying cell numbers. However, chondrocytes undergo a phenotypic shift when propagated in this manner and display characteristics of more adherent fibroblastic cells. Little information is available about the effect of this phenotypic shift on cellular adhesion properties. We evaluated changes in adhesion property as bovine chondrocytes were serially propagated up to five passages in monolayer culture using a centrifugation cell adhesion assay, which was based on counting of cells before and after being exposed to centrifugal dislodgement forces of 120 and 350 g. Chondrocytes proliferated well in a monolayer culture with doubling times of 2-3 days, but they appeared more fibroblastic and exhibited elongated cell morphology with continued passage. The centrifugation cell adhesion assay showed that chondrocytes became more adhesive with passage as the percentage of adherent cells after centrifugation increased and was not statistically different from the adhesion of the fibroblast cell line, L929, starting at passage 3. This increased adhesiveness correlated with a shift to a fibroblastic morphology and increased collagen I mRNA expression starting at passage 2. Our findings indicate that the centrifugation cell adhesion assay may serve as a reproducible tool to track alterations in chondrocyte phenotype during their extended propagation in culture.

Plasma centrifuge

International Nuclear Information System (INIS)

Ikehata, Takashi; Mase, Hiroshi

1998-01-01

The plasma centrifuge is one of statistical isotope separation processes which uses the centrifugal force of a J x B driven rotating plasma in a magnetic field to give rise to the mass-dependent radial transport of isotopic ions. The system has been developed as an alternative to the gas centrifuge because a much higher rotational velocity and separation factor have been achieved. In this review, the physical aspects of the plasma centrifuge followed by the recent experimental achievements are described, especially in comparison with the gas centrifuge. (author)

Novel, improved sample preparation for rapid, direct identification from positive blood cultures using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry.

Science.gov (United States)

Schubert, Sören; Weinert, Kirsten; Wagner, Chris; Gunzl, Beatrix; Wieser, Andreas; Maier, Thomas; Kostrzewa, Markus

2011-11-01

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) is widely used for rapid and reliable identification of bacteria and yeast grown on agar plates. Moreover, MALDI-TOF MS also holds promise for bacterial identification from blood culture (BC) broths in hospital laboratories. The most important technical step for the identification of bacteria from positive BCs by MALDI-TOF MS is sample preparation to remove blood cells and host proteins. We present a method for novel, rapid sample preparation using differential lysis of blood cells. We demonstrate the efficacy and ease of use of this sample preparation and subsequent MALDI-TOF MS identification, applying it to a total of 500 aerobic and anaerobic BCs reported to be positive by a Bactec 9240 system. In 86.5% of all BCs, the microorganism species were correctly identified. Moreover, in 18/27 mixed cultures at least one isolate was correctly identified. A novel method that adjusts the score value for MALDI-TOF MS results is proposed, further improving the proportion of correctly identified samples. The results of the present study show that the MALDI-TOF MS-based method allows rapid (directly from positive BCs and with high accuracy. Copyright © 2011 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

Optimized processing of growth factor mobilized peripheral blood CD34+ products by counterflow centrifugal elutriation.

Science.gov (United States)

Tran, Chy-Anh; Torres-Coronado, Monica; Gardner, Agnes; Gu, Angel; Vu, Hieu; Rao, Anitha; Cao, Lan-Feng; Ahmed, Amira; Digiusto, David

2012-05-01

Cell separation by counterflow centrifugal elutriation has been described for the preparation of monocytes for vaccine applications, but its use in other current good manufacturing practice (cGMP) operations has been limited. In this study, growth factor-mobilized peripheral blood progenitor cell products were collected from healthy donors and processed by elutriation using a commercial cell washing device. Fractions were collected for each product as per the manufacturer's instructions or using a modified protocol developed in our laboratory. Each fraction was analyzed for cell count, viability, and blood cell differential. Our data demonstrate that, using standard elutriation procedures, >99% of red blood cells and platelets were removed from apheresis products with high recoveries of total white blood cells and enrichment of CD34+ cells in two of five fractions. With modification of the basic protocol, we were able to collect all of the CD34+ cells in a single fraction. The CD34-enriched fractions were formulated, labeled with a ferromagnetic antibody to CD34, washed using the Elutra device, and transferred directly to a magnetic bead selection device for further purification. CD34+ cell purities from the column were extremely high (98.7 ± 0.9%), and yields were typical for the device (55.7 ± 12.3%). The processes were highly automated and closed from receipt of the apheresis product through formulation of target-enriched cell fractions. Thus, elutriation is a feasible method for the initial manipulations associated with primary blood cell therapy products and supports cGMP and current good tissue practice-compliant cell processing.

Automated cellular sample preparation using a Centrifuge-on-a-Chip.

Science.gov (United States)

Mach, Albert J; Kim, Jae Hyun; Arshi, Armin; Hur, Soojung Claire; Di Carlo, Dino

2011-09-07

The standard centrifuge is a laboratory instrument widely used by biologists and medical technicians for preparing cell samples. Efforts to automate the operations of concentration, cell separation, and solution exchange that a centrifuge performs in a simpler and smaller platform have had limited success. Here, we present a microfluidic chip that replicates the functions of a centrifuge without moving parts or external forces. The device operates using a purely fluid dynamic phenomenon in which cells selectively enter and are maintained in microscale vortices. Continuous and sequential operation allows enrichment of cancer cells from spiked blood samples at the mL min(-1) scale, followed by fluorescent labeling of intra- and extra-cellular antigens on the cells without the need for manual pipetting and washing steps. A versatile centrifuge-analogue may open opportunities in automated, low-cost and high-throughput sample preparation as an alternative to the standard benchtop centrifuge in standardized clinical diagnostics or resource poor settings.

Classification of positive blood cultures

DEFF Research Database (Denmark)

Gradel, Kim Oren; Knudsen, Jenny Dahl; Arpi, Magnus

2012-01-01

. For each classification, we tabulated episodes derived by the physicians assessment and the computer algorithm and compared 30-day mortality between concordant and discrepant groups with adjustment for age, gender, and comorbidity. RESULTS: Physicians derived 9,482 reference episodes from 21,705 positive......- vs. hospitalonset, whereas there were no material differences within the other comparison groups. CONCLUSIONS: Using data from health administrative registries, we found high agreement between the computer algorithms and the physicians assessments as regards contamination vs. bloodstream infection......ABSTRACT: BACKGROUND: Information from blood cultures is utilized for infection control, public health surveillance, and clinical outcome research. This information can be enriched by physicians assessments of positive blood cultures, which are, however, often available from selected patient groups...

Centrifugal Force Based Magnetic Micro-Pump Driven by Rotating Magnetic Fields

International Nuclear Information System (INIS)

Kim, S H; Hashi, S; Ishiyama, K

2011-01-01

This paper presents a centrifugal force based magnetic micro-pump for the pumping of blood. Most blood pumps are driven by an electrical motor with wired control. To develop a wireless and battery-free blood pump, the proposed pump is controlled by external rotating magnetic fields with a synchronized impeller. Synchronization occurs because the rotor is divided into multi-stage impeller parts and NdFeB permanent magnet. Finally, liquid is discharged by the centrifugal force of multi-stage impeller. The proposed pump length is 30 mm long and 19 mm in diameter which much smaller than currently pumps; however, its pumping ability satisfies the requirement for a blood pump. The maximum pressure is 120 mmHg and the maximum flow rate is 5000ml/min at 100 Hz. The advantage of the proposed pump is that the general mechanical problems of a normal blood pump are eliminated by the proposed driving mechanism.

Centrifugal Force Based Magnetic Micro-Pump Driven by Rotating Magnetic Fields

Science.gov (United States)

Kim, S. H.; Hashi, S.; Ishiyama, K.

2011-01-01

This paper presents a centrifugal force based magnetic micro-pump for the pumping of blood. Most blood pumps are driven by an electrical motor with wired control. To develop a wireless and battery-free blood pump, the proposed pump is controlled by external rotating magnetic fields with a synchronized impeller. Synchronization occurs because the rotor is divided into multi-stage impeller parts and NdFeB permanent magnet. Finally, liquid is discharged by the centrifugal force of multi-stage impeller. The proposed pump length is 30 mm long and19 mm in diameter which much smaller than currently pumps; however, its pumping ability satisfies the requirement for a blood pump. The maximum pressure is 120 mmHg and the maximum flow rate is 5000ml/min at 100 Hz. The advantage of the proposed pump is that the general mechanical problems of a normal blood pump are eliminated by the proposed driving mechanism.

Microfluidic device for acoustic cell lysis

Science.gov (United States)

Branch, Darren W.; Cooley, Erika Jane; Smith, Gennifer Tanabe; James, Conrad D.; McClain, Jaime L.

2015-08-04

A microfluidic acoustic-based cell lysing device that can be integrated with on-chip nucleic acid extraction. Using a bulk acoustic wave (BAW) transducer array, acoustic waves can be coupled into microfluidic cartridges resulting in the lysis of cells contained therein by localized acoustic pressure. Cellular materials can then be extracted from the lysed cells. For example, nucleic acids can be extracted from the lysate using silica-based sol-gel filled microchannels, nucleic acid binding magnetic beads, or Nafion-coated electrodes. Integration of cell lysis and nucleic acid extraction on-chip enables a small, portable system that allows for rapid analysis in the field.

Microfluidic size separation of cells and particles using a swinging bucket centrifuge.

Science.gov (United States)

Yeo, Joo Chuan; Wang, Zhiping; Lim, Chwee Teck

2015-09-01

Biomolecular separation is crucial for downstream analysis. Separation technique mainly relies on centrifugal sedimentation. However, minuscule sample volume separation and extraction is difficult with conventional centrifuge. Furthermore, conventional centrifuge requires density gradient centrifugation which is laborious and time-consuming. To overcome this challenge, we present a novel size-selective bioparticles separation microfluidic chip on a swinging bucket minifuge. Size separation is achieved using passive pressure driven centrifugal fluid flows coupled with centrifugal force acting on the particles within the microfluidic chip. By adopting centrifugal microfluidics on a swinging bucket rotor, we achieved over 95% efficiency in separating mixed 20 μm and 2 μm colloidal dispersions from its liquid medium. Furthermore, by manipulating the hydrodynamic resistance, we performed size separation of mixed microbeads, achieving size efficiency of up to 90%. To further validate our device utility, we loaded spiked whole blood with MCF-7 cells into our microfluidic device and subjected it to centrifugal force for a mere duration of 10 s, thereby achieving a separation efficiency of over 75%. Overall, our centrifugal microfluidic device enables extremely rapid and label-free enrichment of different sized cells and particles with high efficiency.

Determining Human Clot Lysis Time (in vitro with Plasminogen/Plasmin from Four Species (Human, Bovine, Goat, and Swine

Directory of Open Access Journals (Sweden)

Omaira Cañas Bermúdez

2015-05-01

Full Text Available Cardiovascular disease is the leading cause of death worldwide, including failures in the plasminogen/plasmin system which is an important factor in poor lysis of blood clots. This article studies the fibrinolytic system in four species of mammals, and it identifies human plasminogen with highest thrombolysis efficiency. It examines plasminogen from four species (human, bovine, goat, and swine and identifies the most efficient one in human clot lysis in vitro. All plasminogens were identically purified by affinity chromatography. Human fibrinogen was purified by fractionation with ethanol. The purification of both plasminogen and fibrinogen was characterized by one-dimensional SDS-PAGE (10%. Human clot formation in vitro and its dissolution by plasminogen/plasmin consisted of determining lysis time from clot formation to its dilution. Purification of proteins showed greater than 95% purity, human plasminogen showed greater ability to lyse clot than animal plasminogen. The article concludes that human plasminogen/plasmin has the greatest catalysis and efficiency, as it dissolves human clot up to three times faster than that of irrational species.

Maintenance, endogeneous, respiration, lysis, decay and predation

DEFF Research Database (Denmark)

loosdrecht, Marc C. M. Van; Henze, Mogens

1999-01-01

mechanism is microbiologically correct. The lysis/decay model mechanism is a strongly simplified representation of reality. This paper tries to review the processes grouped under endogenous respiration in activated sludge models. Mechanisms and processes such as maintenance, lysis, internal and external...... decay, predation and death-regeneration are discussed. From recent microbial research it has become evident that cells do not die by themselves. Bacteria are however subject to predation by protozoa. Bacteria store reserve polymers that in absence of external substrate are used for growth...

Micro Corona Ionizer as an Ozone Source for Bacterial Cell Lysis

Science.gov (United States)

Lee, Eun-Hee; Lim, Hyun Jeong; Chua, Beelee; Son, Ahjeong

2015-04-01

DNA extraction is a critical process of DNA assays including polymerase chain reaction (PCR), microarrays, molecular cloning, and DNA hybridization which has been well established and can be implemented by commercial kits. DNA extraction involves cell lysis, precipitation, and purification through the combination of physical and chemical processes. Cell lysis is essential to high DNA recovery yield which can be achieved via a variety of physical, chemical, and enzymatic methods. However, these methods were originally developed for bioassays that were labor intensive, time consuming, and vulnerable to contamination and inhibition. Here, we proposed to employ a micro corona ionizer as an ozone source to lyse bacterial cells. Ozone has been well known and used as a disinfectant which allows cell lysis and DNA extraction. Previously, we have shown that a micro corona ionizer is capable of generating a significant amount of ozone. In this study, we employed the micro corona ionizer for the bacterial cell lysis which consists of a 50 μm diameter cantilever wire as the discharge cathode and a 50 μm thick copper foil as anode. Applied voltages varied from 1900 to 2200 V with corresponding corona currents from 16 to 28 μA. The resultant ozone (concentration > 0.14 ppm) generated from the micro corona ionizer was bubbled into the sample via a miniature pump. We demonstrated the cell lysis of Pseudomonas putida as the target bacterium using the micro corona ionizer. At a flow rate of 38 ml/min and applied corona voltage of 2000 V, 98.5 ± 0.2% lysis (normalized to sonication result) was achieved after 10 min. In comparison, untreated and air-treated samples showed normalized % lysis of 11.9 ± 2.4 and 36.1 ± 1.7%, respectively. We also showed that the cell lysis efficiency could be significantly increased by increasing the flow rate and the applied corona voltage. By comparing the experimental results for continuous and pulsed treatment, we verified that the percentage of

Evaluation of single and double centrifugation tube methods for concentrating equine platelets.

Science.gov (United States)

Argüelles, D; Carmona, J U; Pastor, J; Iborra, A; Viñals, L; Martínez, P; Bach, E; Prades, M

2006-10-01

The aim of this study was to evaluate single and double centrifugation tube methods for concentrating equine platelets. Whole blood samples were collected from clinically normal horses and processed by use of single and double centrifugation tube methods to obtain four platelet concentrates (PCs): PC-A, PC-B, PC-C, and PC-D, which were analyzed using a flow cytometry hematology system for hemogram and additional platelet parameters (mean platelet volume, platelet distribution width, mean platelet component concentration, mean platelet component distribution width). Concentrations of transforming growth factor beta 1 (TGF-beta(1)) were determined in all the samples. Platelet concentrations for PC-A, PC-B, PC-C, and PC-D were 45%, 44%, 71%, and 21% higher, respectively, compared to the same values for citrated whole blood samples. TGF-beta(1) concentrations for PC-A, PC-B, PC-C, and PC-D were 38%, 44%, 44%, and 37% higher, respectively, compared to citrated whole blood sample values. In conclusion, the single and double centrifugation tube methods are reliable methods for concentrating equine platelets and for obtaining potentially therapeutic TGF-beta(1) levels.

Generation of Human Induced Pluripotent Stem Cells from Peripheral Blood Mononuclear Cells Using Sendai Virus.

Science.gov (United States)

Soares, Filipa A C; Pedersen, Roger A; Vallier, Ludovic

2016-01-01

This protocol describes the efficient isolation of peripheral blood mononuclear cells from circulating blood via density gradient centrifugation and subsequent generation of integration-free human induced pluripotent stem cells. Peripheral blood mononuclear cells are cultured for 9 days to allow expansion of the erythroblast population. The erythroblasts are then used to derive human induced pluripotent stem cells using Sendai viral vectors, each expressing one of the four reprogramming factors Oct4, Sox2, Klf4, and c-Myc.

Anaplasma Marginale isolation from infected bovine erythrocytes or from its floating culture

International Nuclear Information System (INIS)

Canon Q, Y.

1986-01-01

Isolation of Anaplasma Marginale is of great importance because this is the cause of anaplasmosis in cattle. Anaplasmosis is a mortal disease and spreads easily. To isolate the anaplasm, four different experiments were developed; in the first experiment, the parasite was isolated from parasitic blood, by means of three methods; osmotic shock, sonic vibration and treatment with hemolisine. In the second experiment, the parasite source was the top of the bacteriologic culture of anaplasma marginale obtained by means of slow centrifugation. In the third experiment, it was used parasitic blood diluted in PBS and liquid nitrogen criopreserved. In the fourth experiment.it was used parasitic blood which was separated by means of sonic oscillation. This method was more adequate to free the parasite from the host cell. Differential centrifugation was the best method to separate parasite of the stroma and ghost cells

Direct identification of bacteria from BacT/ALERT anaerobic positive blood cultures by MALDI-TOF MS: MALDI Sepsityper kit versus an in-house saponin method for bacterial extraction.

Science.gov (United States)

Meex, Cécile; Neuville, Florence; Descy, Julie; Huynen, Pascale; Hayette, Marie-Pierre; De Mol, Patrick; Melin, Pierrette

2012-11-01

In cases of bacteraemia, a rapid species identification of the causal agent directly from positive blood culture broths could assist clinicians in the timely targeting of empirical antimicrobial therapy. For this purpose, we evaluated the direct identification of micro-organisms from BacT/ALERT (bioMérieux) anaerobic positive blood cultures without charcoal using the Microflex matrix-assisted laser desorption/ionization (MALDI) time of flight MS (Bruker), after bacterial extraction by using two different methods: the MALDI Sepsityper kit (Bruker) and an in-house saponin lysis method. Bruker's recommended criteria for identification were expanded in this study, with acceptance of the species identification when the first three results with the best matches with the MALDI Biotyper database were identical, whatever the scores were. In total, 107 monobacterial cultures and six polymicrobial cultures from 77 different patients were included in this study. Among monomicrobial cultures, we identified up to the species level 67 and 66 % of bacteria with the MALDI Sepsityper kit and the saponin method, respectively. There was no significant difference between the two extraction methods. The direct species identification was particularly inconclusive for Gram-positive bacteria, as only 58 and 52 % of them were identified to the species level with the MALDI Sepsityper kit and the saponin method, respectively. Results for Gram-negative bacilli were better, with 82.5 and 90 % of correct identification to the species level with the MALDI Sepsityper kit and the saponin method, respectively. No misidentifications were given by the direct procedures when compared with identifications provided by the conventional method. Concerning the six polymicrobial blood cultures, whatever the extraction method used, a correct direct identification was only provided for one of the isolated bacteria on solid medium in all cases. The analysis of the time-to-result demonstrated a reduction

Calculations of the Acceleration of Centrifugal Loading on Adherent Cells

Science.gov (United States)

Chen, Kang; Song, Yang; Liu, Qing; Zhang, Chunqiu

2017-07-01

Studies have shown that the morphology and function of living cells are greatly affected by the state of different high acceleration. Based on the centrifuge, we designed a centrifugal cell loading machine for the mechanical biology of cells under high acceleration loading. For the machine, the feasibility of the experiment was studied by means of constant acceleration or variable acceleration loading in the Petri dish fixture and/or culture flask. Here we analyzed the distribution of the acceleration of the cells with the change of position and size of the culturing device quantitatively. It is obtained that Petri dish fixture and/or culture flask can be used for constant acceleration loading by experiments; the centripetal acceleration of the adherent cells increases with the increase of the distance between the rotor center of the centrifuge and the fixture of the Petri dish and the size of the fixture. It achieves the idea that the general biology laboratory can conduct the study of mechanical biology at high acceleration. It also provides a basis for more accurate study of the law of high acceleration on mechanobiology of cells.

Is anaerobic blood culture necessary? If so, who needs it?

Science.gov (United States)

Iwata, Kentaro; Takahashi, Miwa

2008-07-01

The role of anaerobic blood cultures is not validated, although they are drawn routinely. We performed a retrospective chart review at a private hospital in Japan for patients admitted between July 1, 2004 to June 30, 2005 to determine patient characteristics resulting in anaerobic blood culture. During the study period, 17,775 blood culture bottles were sent for the analysis, and 2132 bottles (12.0%) were positive for microbial growth. Bacteria were grown from 958 anaerobic bottles (44.7%), and 719 (33.7%) of those were judged to represent real infections, which accounted for 410 cases of bacteremia. Only 47 cases (11.5%) were detected by anaerobic cultures alone. Among those 47, obligate anaerobes represented 12 cases. Clinical evaluation could have predicted 7 of 12 cases of obligate anaerobic bacteremia. In the remaining 5 cases, the source of bacteremia was unclear. There were 2.7 cases of anaerobic bacteremia per 1000 blood cultures. The mortality attributable to anaerobic bacteremia was 50%. Among bacteremic cases not caused by obligate anaerobes yet diagnosed solely by anaerobic bottles, either the standard 2 sets of blood were not taken or their clinical outcomes were favorable. Anaerobic blood culture can be avoided in most cases. Anaerobic blood culture may be most helpful when (1) bacteremia because of obligate anaerobes is clinically suspected, (2) patients are severely immunocompromised, and (3) source of bacteremia is not identified by clinical evaluation.

Evaluation of a Spiral Groove Geometry for Improvement of Hemolysis Level in a Hydrodynamically Levitated Centrifugal Blood Pump.

Science.gov (United States)

Murashige, Tomotaka; Kosaka, Ryo; Sakota, Daisuke; Nishida, Masahiro; Kawaguchi, Yasuo; Yamane, Takashi; Maruyama, Osamu

2015-08-01

The purpose of this study is to evaluate a spiral groove geometry for a thrust bearing to improve the hemolysis level in a hydrodynamically levitated centrifugal blood pump. We compared three geometric models: (i) the groove width is the same as the ridge width at any given polar coordinate (conventional model); (ii) the groove width contracts inward from 9.7 to 0.5 mm (contraction model); and (iii) the groove width expands inward from 0.5 to 4.2 mm (expansion model). To evaluate the hemolysis level, an impeller levitation performance test and in vitro hemolysis test were conducted using a mock circulation loop. In these tests, the driving conditions were set at a pressure head of 200 mm Hg and a flow rate of 4.0 L/min. As a result of the impeller levitation performance test, the bottom bearing gaps of the contraction and conventional models were 88 and 25 μm, respectively. The impeller of the expansion model touched the bottom housing. In the hemolysis test, the relative normalized index of hemolysis (NIH) ratios of the contraction model in comparison with BPX-80 and HPM-15 were 0.6 and 0.9, respectively. In contrast, the relative NIH ratios of the conventional model in comparison with BPX-80 and HPM-15 were 9.6 and 13.7, respectively. We confirmed that the contraction model achieved a large bearing gap and improved the hemolysis level in a hydrodynamically levitated centrifugal blood pump. Copyright © 2015 International Center for Artificial Organs and Transplantation and Wiley Periodicals, Inc.

Development of a Compact Maglev Centrifugal Blood Pump Enclosed in a Titanium Housing

Science.gov (United States)

Pai, Chi Nan; Shinshi, Tadahiko; Asama, Junichi; Takatani, Setsuo; Shimokohbe, Akira

A compact centrifugal blood pump consisting of a controlled two-degrees-of-freedom radial magnetic bearing and a brushless DC motor enclosed in a titanium housing has been developed for use as an implantable ventricular assist device. The magnetic bearing also supports axial and angular motions of the impeller via a magnetic coupling. The top housing is made of pure titanium, while the impeller and the stator are coated with pure titanium and Ti-6Al-7Nb, respectively, to improve the biocompatibility of the pump. The combination of pure titanium and titanium alloy was chosen because of the sensitivity of eddy current type displacement sensors through the intervening conducting wall. The dimensions of the pump are 69.0 mm in diameter and 28.5 mm in height. During a pump performance test, axial shifting of the impeller due to hydraulic forces led to variations in the rotational positioning signal, causing loss of control of the rotational speed. This problem was solved by conditioning the rotational positioning signal. With a flow rate of 5 l/min against a head pressure of 100 mmHg, the power consumption and efficiency of the pump were 5.5 W and 20%, respectively. Furthermore, the hemolysis of the blood pump was 43.6% lower when compared to that of a commercially available pump.

Does this adult patient with suspected bacteremia require blood cultures?

Science.gov (United States)

Coburn, Bryan; Morris, Andrew M; Tomlinson, George; Detsky, Allan S

2012-08-01

Clinicians order blood cultures liberally among patients in whom bacteremia is suspected, though a small proportion of blood cultures yield true-positive results. Ordering blood cultures inappropriately may be both wasteful and harmful. To review the accuracy of easily obtained clinical and laboratory findings to inform the decision to obtain blood cultures in suspected bacteremia. A MEDLINE and EMBASE search (inception to April 2012) yielded 35 studies that met inclusion criteria for evaluating the accuracy of clinical variables for bacteremia in adult immunocompetent patients, representing 4566 bacteremia and 25,946 negative blood culture episodes. Data were extracted to determine the prevalence and likelihood ratios (LRs) of findings for bacteremia. The pretest probability of bacteremia varies depending on the clinical context, from low (eg, cellulitis: 2%) to high (eg, septic shock: 69%). Elevated temperatures alone do not accurately predict bacteremia (for ≥38°C [>100.3°F], LR, 1.9 [95% CI, 1.4-2.4]; for ≥38.5°C [>101.2°F], LR, 1.4 [95% CI, 1.1-2.0]), nor does isolated leukocytosis (LR, cultures should not be ordered for adult patients with isolated fever or leukocytosis without considering the pretest probability. SIRS and the decision rule may be helpful in identifying patients who do not need blood cultures. These conclusions do not apply to immunocompromised patients or when endocarditis is suspected.

Optimizing platelet-rich plasma gel formation by varying time and gravitational forces during centrifugation.

Science.gov (United States)

Jo, Chris H; Roh, Young Hak; Kim, Ji Eun; Shin, Sue; Yoon, Kang Sup

2013-10-01

Despite the increasing clinical use of topical platelet-rich plasma (PRP) to enhance tissue healing and regeneration, there is no properly standardized method of autologous PRP gel preparation. This study examined the effect of the centrifugation time and gravitational force (g) on the platelet recovery ratio of PRP and determined the most effective centrifugation conditions for preparing PRP. Two-step centrifugations for preparing PRP were used in 39 subjects who had consented prior to the study's start. The separating centrifugation (Step 1, used to separate whole blood into its two main components: red blood cells and plasma) was tested from 500g to 1900g at 200g increments for 5 minutes (min), and from 100g to 1300g at 200g increments for 10 minutes. After separating centrifugation, upper plasma layer was transferred to another plain tube for the condensation centrifugation and remaining lower cell layer was discarded. The condensation centrifugation (Step 2, used to condense the platelets in the separated plasma) was tested at 1000g for 15 min, 1500g for 15 min, 2000g for 5 min and 3000g for 5 min, additionally at 1000g for 10 min and 1500g for 10 min. Platelet gelation was induced by adding 10% calcium gluconate to final PRP with volume ratio of 1:10. The optimal separating centrifugation conditions were followed by 900g for 5 minutes and the condensation conditions were followed by 1500g for 15 minutes, of which recovery ratios were 92.0 ± 3.1% and 84.3 ± 10.0%, respectively.

Effect of postmortem sampling technique on the clinical significance of autopsy blood cultures.

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Hove, M; Pencil, S D

1998-02-01

Our objective was to investigate the value of postmortem autopsy blood cultures performed with an iodine-subclavian technique relative to the classical method of atrial heat searing and antemortem blood cultures. The study consisted of a prospective autopsy series with each case serving as its own control relative to subsequent testing, and a retrospective survey of patients coming to autopsy who had both autopsy blood cultures and premortem blood cultures. A busy academic autopsy service (600 cases per year) at University of Texas Medical Branch Hospitals, Galveston, Texas, served as the setting for this work. The incidence of non-clinically relevant (false-positive) culture results were compared using different methods for collecting blood samples in a prospective series of 38 adult autopsy specimens. One hundred eleven adult autopsy specimens in which both postmortem and antemortem blood cultures were obtained were studied retrospectively. For both studies, positive culture results were scored as either clinically relevant or false positives based on analysis of the autopsy findings and the clinical summary. The rate of false-positive culture results obtained by an iodine-subclavian technique from blood drawn soon after death were statistically significantly lower (13%) than using the classical method of obtaining blood through the atrium after heat searing at the time of the autopsy (34%) in the same set of autopsy subjects. When autopsy results were compared with subjects' antemortem blood culture results, there was no significant difference in the rate of non-clinically relevant culture results in a paired retrospective series of antemortem blood cultures and postmortem blood cultures using the iodine-subclavian postmortem method (11.7% v 13.5%). The results indicate that autopsy blood cultures obtained using the iodine-subclavian technique have reliability equivalent to that of antemortem blood cultures.

The effect of centrifugation speed and time on pre-analytical platelet activation

DEFF Research Database (Denmark)

Söderström, Anna Cecilia; Nybo, Mads; Nielsen, Christian

2016-01-01

BACKGROUND: The results of laboratory analyses are affected by pre-analytical variables, and in particular can platelets be activated by shear handling stress and secrete granular substances. We therefore evaluated the effect of centrifugation speed and time on pre-analytical platelet activation....... METHODS: Citrate- and EDTA-anticoagulated blood from healthy volunteers were centrifuged at 80-10,000 g for 5-15 min to prepare plasma and platelet-rich plasma. Pre-analytical platelet activation was assessed by flow cytometric measurement of platelet P-selectin (CD62p) expression. Blood cell counts, mean...... of platelets expressing P-selectin in citrate- and EDTA-plasma centrifuged at 2000 g for 10 min were 43% [interquartile range (IQR), 38%-53%] and 56% (IQR, 31%-78%), respectively (p=0.82). Platelet-rich plasma prepared at 100-250 g for 10 min had significantly lower platelet P-selectin expression (11%-15%), p...

CENTRIFUGAL SEPARATORS

Science.gov (United States)

Skarstrom, C.

1959-03-10

A centrifugal separator is described for separating gaseous mixtures where the temperature gradients both longitudinally and radially of the centrifuge may be controlled effectively to produce a maximum separation of the process gases flowing through. Tbe invention provides for the balancing of increases and decreases in temperature in various zones of the centrifuge chamber as the result of compression and expansions respectively, of process gases and may be employed effectively both to neutralize harmful temperature gradients and to utilize beneficial temperaturc gradients within the centrifuge.

Centrifugal pumps

CERN Document Server

Anderson, HH

1981-01-01

Centrifugal Pumps describes the whole range of the centrifugal pump (mixed flow and axial flow pumps are dealt with more briefly), with emphasis on the development of the boiler feed pump. Organized into 46 chapters, this book discusses the general hydrodynamic principles, performance, dimensions, type number, flow, and efficiency of centrifugal pumps. This text also explains the pumps performance; entry conditions and cavitation; speed and dimensions for a given duty; and losses. Some chapters further describe centrifugal pump mechanical design, installation, monitoring, and maintenance. The

Cytokinesis-block micronucleus method in micro-blood cultures

International Nuclear Information System (INIS)

Liu Jinwen; Wang Lianzhi; Yang Cangzhen; Yao Yanyu

1991-01-01

This paper reports the cytokinesis-block micronucleus method in micro-blood cultures. The observations on detection induced micronuclei of different doses of 60 Co γ-rays irradiation and spontaneous micronucleus of different ages were performed with CB method in comporison with conventional micronucleus (CM) method. The results showed that with direct peripheral micro-blood cultures the cytoknesis-block micronuclei is also obtained. Using CB method, the micronuclei fequency of different ages was linear relationship, Y = 1.62 + 0.74 D, the spontaneous micronuclei frequency of different ages was 4.14%, the induced micronuclei also was a linear relationship, Y = 6.01 + 0.692 D. Using CM method, it showed that the induced micronuclei was a linear relationship, Y = 0.486 D - 1.968, but there is no significant difference between the micronuclei frequency of different ages. Comparison with CM and direct blood smear methods confirmed that the cytokinesis-block method of micro-blood cultures is more sensitive and precise

Blood glucose, lactate, pyruvate, glycerol, 3-hydroxybutyrate and acetoacetate measurements in man using a centrifugal analyser with a fluorimetric attachment.

Science.gov (United States)

Harrison, J; Hodson, A W; Skillen, A W; Stappenbeck, R; Agius, L; Alberti, K G

1988-03-01

Methods are described for the analysis of glucose, lactate, pyruvate, alanine, glycerol, 3-hydroxybutyrate and acetoacetate in perchloric acid extracts of human blood, using the Cobas Bio centrifugal analyser fitted with a fluorimetric attachment. Intra-assay and inter-assay coefficients of variation ranged from 1.9 to 7.9% and from 1.0 to 7.2% respectively. Correlation coefficients ranged from 0.96 to 0.99 against established continuous-flow and manual spectrophotometric methods. All seven metabolites can be measured using a single perchloric acid extract of 20 microliter of blood. The versatility of the assays is such that as little as 100 pmol pyruvate, 3-hydroxybutyrate or as much as 15 nmol glucose can be measured in the same 20 microliter extract.

Evaluation of a reduced centrifugation time and higher centrifugal force on various general chemistry and immunochemistry analytes in plasma and serum.

Science.gov (United States)

Møller, Mette F; Søndergaard, Tove R; Kristensen, Helle T; Münster, Anna-Marie B

2017-09-01

Background Centrifugation of blood samples is an essential preanalytical step in the clinical biochemistry laboratory. Centrifugation settings are often altered to optimize sample flow and turnaround time. Few studies have addressed the effect of altering centrifugation settings on analytical quality, and almost all studies have been done using collection tubes with gel separator. Methods In this study, we compared a centrifugation time of 5 min at 3000 ×  g to a standard protocol of 10 min at 2200 ×  g. Nine selected general chemistry and immunochemistry analytes and interference indices were studied in lithium heparin plasma tubes and serum tubes without gel separator. Results were evaluated using mean bias, difference plots and coefficient of variation, compared with maximum allowable bias and coefficient of variation used in laboratory routine quality control. Results For all analytes except lactate dehydrogenase, the results were within the predefined acceptance criteria, indicating that the analytical quality was not compromised. Lactate dehydrogenase showed higher values after centrifugation for 5 min at 3000 ×  g, mean bias was 6.3 ± 2.2% and the coefficient of variation was 5%. Conclusions We found that a centrifugation protocol of 5 min at 3000 ×  g can be used for the general chemistry and immunochemistry analytes studied, with the possible exception of lactate dehydrogenase, which requires further assessment.

Separations by centrifugal phenomena

International Nuclear Information System (INIS)

Hsu, H.W.

1981-01-01

The technical information presented herein emphasizes the uniqueness of the centrifugal separations methodology and pertinent theory for various kinds of centrifugation. The topics are arranged according to gas, liquid, and solid phases, in the order of increasing densities. Much space is devoted to liquid centrifugation because of the importance of this technique in chemical and biological laboratories. Many separational and characterizational examples are illustrated in detail. The material has been divided into 7 chapters entitled: 1) Introduction, 2) Basic Theory of Centrifugation, 3) Gas Centrifuges, 4) Preparative Liquid Centrifuges, 5) Analytical Liquid Centrifuges, 6) Liquid Centrifuges in Practice, and 7) Mechanical Separations by Centrifuges. Separate abstracts have been prepared for each chapter except the introduction

Comparison between MALDI-TOF MS and FilmArray Blood Culture Identification panel for rapid identification of yeast from positive blood culture.

Science.gov (United States)

Paolucci, M; Foschi, C; Tamburini, M V; Ambretti, S; Lazzarotto, T; Landini, M P

2014-09-01

In this study we evaluated MALDI-TOF MS and FilmArray methods for the rapid identification of yeast from positive blood cultures. FilmArray correctly identified 20/22 of yeast species, while MALDI-TOF MS identified 9/22. FilmArray is a reliable and rapid identification system for the direct identification of yeasts from positive blood cultures. Copyright © 2014 Elsevier B.V. All rights reserved.

Description of a double centrifugation tube method for concentrating canine platelets

OpenAIRE

Perazzi, Anna; Busetto, Roberto; Martinello, Tiziana; Drigo, Michele; Pasotto, Daniela; Cian, Francesco; Patruno, Marco; Iacopetti, Ilaria

2013-01-01

Background To evaluate the efficiency of platelet-rich plasma preparations by means of a double centrifugation tube method to obtain platelet-rich canine plasma at a concentration at least 4 times higher than the baseline value and a concentration of white blood cells not exceeding twice the reference range. A complete blood count was carried out for each sample and each concentrate. Whole blood samples were collected from 12 clinically healthy dogs (consenting blood donors). Blood was proces...

effect of natural blue-green algal cells lysis on freshwater quality

African Journals Online (AJOL)

Compaq

released into water due to algal cells lysis was performed by placing samples in two ... Keywords; Algae, cells lysis, Fatty acids, gas chromatography time-of-flight mass spectrometry, water quality ... Factors such as municipal and industrial.

Demonstration of NK cell-mediated lysis of varicella-zoster virus (VZV)-infected cells: characterization of the effector cells

Energy Technology Data Exchange (ETDEWEB)

Tilden, A.B.; Cauda, R.; Grossi, C.E.; Balch, C.M.; Lakeman, A.D.; Whitley, R.J.

1986-06-01

Infection with varicella-zoster virus (VZV) rendered RAJI cells more susceptible to lysis by non-adherent blood lymphocytes. At an effector to target ratio of 80:1 the mean percentage of /sup 51/Cr release of VZV-infected RAJI cells was 41 +/- 12%, whereas that of uninfected RAJI cells was 15 +/- 6%. The increased susceptibility to lysis was associated with increased effector to target conjugate formation in immunofluorescence binding assays. The effector cells cytotoxic for VZV-infected RAJI cells were predominantly Leu-11a/sup +/ Leu-4/sup -/ granular lymphocytes as demonstrated by fluorescence-activated cell sorting. The effector cell active against VZV-infected RAJI cells appeared similar to those active against herpes simplex virus (HSV)-infected cells, because in cold target competition experiments the lysis of /sup 51/Cr-labeled VZV-infected RAJI cells was efficiently inhibited by either unlabeled VZV-infected RAJI cells (mean 71% inhibition, 2:1 ratio unlabeled to labeled target) or HSV-infected RAJI cells (mean 69% inhibition) but not by uninfected RAJI cells (mean 10% inhibition). In contrast, competition experiments revealed donor heterogeneity in the overlap between effector cells for VZV- or HSV-infected RAJI vs K-562 cells.

Centrifugal dewatering of acid casein curd: effect of casein manufacturing and centrifugation variables on curd compression in a laboratory centrifuge.

Science.gov (United States)

Munro, P A; Van Til, H J

1988-10-20

Data relevant to curd compression in a horizontal, solid bowl decanter centrifuge have been obtained by studying the dewatering of acid casein curd in a batch laboratory centrifuge. Analysis of curd compression under centrifugal force predicts a moisture content gradient in the dewatered curd from a maximum at the curd-liquid interface to a minimum at the centrifuge bowl wall. This moisture content gradient was also measured experimentally, and its practical implications are discussed. Increases in centrifugal force, centrifugation time, and centrifugation temperature all caused a marked de crease in dewatered curd moisture content, whereas in creases in precipitation pH and maximum washing temperature caused a smaller decrease in dewatered curd moisture content.

Enhancement of recovery of Neisseria meningitidis by gelatin in blood culture media.

OpenAIRE

Pai, C H; Sorger, S

1981-01-01

The efficacy of gelatin for the recovery of Neisseria meningitidis from blood cultures was evaluated in a clinical setting. The organism was isolated from seven patients with meningococcal infections in blood culture media containing 1% gelatin. In contrast, only two blood cultures from these patients were positive in media without gelatin (P less than 0.05). Gelatin did not influence the recovery of other organisms isolated during this study. Conventional blood culture media may be supplemen...

Catheter placement for lysis of spontaneous intracerebral hematomas: does a catheter position in the core of the hematoma allow more effective and faster hematoma lysis?

Science.gov (United States)

Malinova, Vesna; Schlegel, Anna; Rohde, Veit; Mielke, Dorothee

2017-07-01

For the fibrinolytic therapy of intracerebral hematomas (ICH) using recombinant tissue plasminogen activator (rtPA), a catheter position in the core of the hematoma along the largest clot diameter was assumed to be optimal for an effective clot lysis. However, it never had been proven that core position indeed enhances clot lysis if compared with less optimal catheter positions. In this study, the impact of the catheter position on the effectiveness and on the time course of clot lysis was evaluated. We analyzed the catheter position using a relative error calculating the distance perpendicular to the catheter's center in relation to hematoma's diameter and evaluated the relative hematoma volume reduction (RVR). The correlation of the RVR with the catheter position was evaluated. Additionally, we tried to identify patterns of clot lysis with different catheter positions. The patient's outcome at discharge was evaluated using the Glasgow outcome score. A total of 105 patients were included in the study. The mean hematoma volume was 56 ml. The overall RVR was 62.7 %. In 69 patients, a catheter position in the core of the clot was achieved. We found no significant correlation between catheter position and hematoma RVR (linear regression, p = 0.14). Core catheter position leads to more symmetrical hematoma RVR. Faster clot lysis happens in the vicinity of the catheter openings. We found no significant difference in the patient's outcome dependent on the catheter position (linear regression, p = 0.90). The catheter position in the core of the hematoma along its largest diameter does not significantly influence the effectiveness of clot lysis after rtPA application.

Reduction in Blood Culture Contamination Through Use of Initial Specimen Diversion Device.

Science.gov (United States)

Rupp, Mark E; Cavalieri, R Jennifer; Marolf, Cole; Lyden, Elizabeth

2017-07-15

Blood culture contamination is a clinically significant problem that results in patient harm and excess cost. In a prospective, controlled trial at an academic center Emergency Department, a device that diverts and sequesters the initial 1.5-2 mL portion of blood (which presumably carries contaminating skin cells and microbes) was tested against standard phlebotomy procedures in patients requiring blood cultures due to clinical suspicion of serious infection. In sum, 971 subjects granted informed consent and were enrolled resulting in 904 nonduplicative subjects with 1808 blood cultures. Blood culture contamination was significantly reduced through use of the initial specimen diversion device™ (ISDD) compared to standard procedure: (2/904 [0.22%] ISDD vs 16/904 [1.78%] standard practice, P = .001). Sensitivity was not compromised: true bacteremia was noted in 65/904 (7.2%) ISDD vs 69/904 (7.6%) standard procedure, P = .41. No needlestick injuries or potential bloodborne pathogen exposures were reported. The monthly rate of blood culture contamination for all nurse-drawn and phlebotomist-drawn blood cultures was modeled using Poisson regression to compare the 12-month intervention period to the 6 month before and after periods. Phlebotomists (used the ISDD) experienced a significant decrease in blood culture contamination while the nurses (did not use the ISDD) did not. In sum, 73% of phlebotomists completed a post-study anonymous survey and widespread user satisfaction was noted. Use of the ISDD was associated with a significant decrease in blood culture contamination in patients undergoing blood cultures in an Emergency Department setting. NCT02102087. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America.

Effect of centrifugation time on growth factor and MMP release of an experimental platelet-rich fibrin-type product.

Science.gov (United States)

Eren, Gülnihal; Gürkan, Ali; Atmaca, Harika; Dönmez, Ayhan; Atilla, Gül

2016-07-01

Platelet-rich fibrin (PRF) has a controlled release of growth factors due to the fibrin matrix structure. Different centrifugation protocols were suggested for PRF preparation. Since the derivation method of PRF can alter its contents, in the present study it is aimed to investigate the cell contents and transforming growth factor beta-1 (TGF-β1), platelet-derived growth factor (PDGF-AB), vascular endothelial growth factor (VEGF), matrix metalloproteinase (MMP)-1 and-8 release from experimental PRF-type membranes obtained with different centrifugation times at 400 gravity. Three blood samples were collected from 20 healthy non-smoker volunteers. One tube was used for whole blood analyses. The other two tubes were centrifuged at 400 g for 10 minutes (group A) or 12 minutes (group B). Each experimental PRF-type membrane was placed in Dulbecco's Modified Eagle's Medium (DMEM)and at 1, 24 and 72 hours, TGF-β1, PDGF-AB, VEGF, MMP-1 and -8 release amounts were analysed by enzyme-linked immunosorbent assay (ELISA). The blood cell count of membranes was determined by subtracting plasma supernatant and red blood cell (RBC) mixture from the whole blood cell counts. At 72 hours, the VEGF level of group B was statistically higher than that of group A (p = 0.040). The centrifugation time was not found to influence the release of other growth factors, enzymes and cell counts. Within the limits of the present study, it might be suggested that centrifugation time at a constant gravity has a significant effect on the VEGF levels released from experimental PRF-type membrane. It can be concluded that due to the importance of VEGF in the tissue healing process, membranes obtained at 12-minute centrifugation time may show a superior potential in wound healing.

Centrifugation

International Nuclear Information System (INIS)

Subbaramajer.

1983-01-01

The theoretical analysis of the processes taking place at centrifugal method of isotope separation taking into account the latest investigations, in particular, investigation of velocity field applying the theory of boundary layers in rotating gas is conducted. As a result of using power computers for the solution of hydrodynamics equations by numerical methods sufficiently exact solutions of main hydrodynamic equations, reflecting the real centrifuge construction are derived. The increase of calculation accuracy of the flow field reflected also on the accuracy of the diffusion equation solution. Three parameters of similarity (height of transfer unit, flow, mass transfer coefficient) and their connection with the flow field, elementary separation coefficient in a cetrifugal field and molecular diffusion coefficient is determined. Modified formulas for the separation coefficient and separation centrifuge power taking into account similarity parameter changes over the axis are derived. The possibility of determining the system of controlled parameters optimizing the separation centrifuge power is shown

Platelet adhesiveness: the effect of centrifugation on the measurement of adhesiveness in platelet-rich plasma

Science.gov (United States)

McBride, J. A.

1968-01-01

Platelet adhesiveness has been measured in citrated whole blood and in platelet-rich plasma obtained from normal subjects, splenectomized patients, and from patients in whom the diagnosis of recurrent venous thrombosis had been made. The duration of centrifugation used in the preparation of platelet-rich plasma was found to have a profound effect on the measurement of platelet adhesiveness because the figure for platelet adhesiveness measured in platelet-rich plasma obtained by centrifugation was considerably lower than that found in citrated whole blood. This effect was particularly marked when platelet-rich plasma was obtained from subjects in whom platelet adhesiveness measured in whole blood was increased. PMID:5699080

Multidrug resistant Salmonellae isolated from blood culture samples ...

African Journals Online (AJOL)

This study investigates the prevalence of R-plasmids in Salmonella sp. isolated from blood samples of suspected typhoid patients in Warri, Nigeria. A total of 136 blood samples were collected between May and December,2009 and screened for the presence of Salmonellae using standard blood culture techniques of which ...

Leukocyte Lysis and Cytokine Induction by the Human Sexually Transmitted Parasite Trichomonas vaginalis.

Directory of Open Access Journals (Sweden)

Frances Mercer

2016-08-01

Full Text Available Trichomonas vaginalis (Tv is an extracellular protozoan parasite that causes the most common non-viral sexually transmitted infection: trichomoniasis. While acute symptoms in women may include vaginitis, infections are often asymptomatic, but can persist and are associated with medical complications including increased HIV susceptibility, infertility, pre-term labor, and higher incidence of cervical cancer. Heightened inflammation resulting from Tv infection could account for these complications. Effective cellular immune responses to Tv have not been characterized, and re-infection is common, suggesting a dysfunctional adaptive immune response. Using primary human leukocyte components, we have established an in vitro co-culture system to assess the interaction between Tv and the cells of the human immune system. We determined that in vitro, Tv is able to lyse T-cells and B-cells, showing a preference for B-cells. We also found that Tv lysis of lymphocytes was mediated by contact-dependent and soluble factors. Tv lysis of monocytes is far less efficient, and almost entirely contact-dependent. Interestingly, a common symbiont of Tv, Mycoplasma hominis, did not affect cytolytic activity of the parasite, but had a major impact on cytokine responses. M. hominis enabled more diverse inflammatory cytokine secretion in response to Tv and, of the cytokines tested, Tv strains cleared of M. hominis induced only IL-8 secretion from monocytes. The quality of the adaptive immune response to Tv is therefore likely influenced by Tv symbionts, commensals, and concomitant infections, and may be further complicated by direct parasite lysis of effector immune cells.

Leukocyte Lysis and Cytokine Induction by the Human Sexually Transmitted Parasite Trichomonas vaginalis

Science.gov (United States)

Mercer, Frances; Diala, Fitz Gerald I.; Chen, Yi-Pei; Molgora, Brenda M.; Ng, Shek Hang; Johnson, Patricia J.

2016-01-01

Trichomonas vaginalis (Tv) is an extracellular protozoan parasite that causes the most common non-viral sexually transmitted infection: trichomoniasis. While acute symptoms in women may include vaginitis, infections are often asymptomatic, but can persist and are associated with medical complications including increased HIV susceptibility, infertility, pre-term labor, and higher incidence of cervical cancer. Heightened inflammation resulting from Tv infection could account for these complications. Effective cellular immune responses to Tv have not been characterized, and re-infection is common, suggesting a dysfunctional adaptive immune response. Using primary human leukocyte components, we have established an in vitro co-culture system to assess the interaction between Tv and the cells of the human immune system. We determined that in vitro, Tv is able to lyse T-cells and B-cells, showing a preference for B-cells. We also found that Tv lysis of lymphocytes was mediated by contact-dependent and soluble factors. Tv lysis of monocytes is far less efficient, and almost entirely contact-dependent. Interestingly, a common symbiont of Tv, Mycoplasma hominis, did not affect cytolytic activity of the parasite, but had a major impact on cytokine responses. M. hominis enabled more diverse inflammatory cytokine secretion in response to Tv and, of the cytokines tested, Tv strains cleared of M. hominis induced only IL-8 secretion from monocytes. The quality of the adaptive immune response to Tv is therefore likely influenced by Tv symbionts, commensals, and concomitant infections, and may be further complicated by direct parasite lysis of effector immune cells. PMID:27529696

Isolation of mesenchymal stem cells from equine umbilical cord blood

DEFF Research Database (Denmark)

Koch, Thomas Gadegaard; Heerkens, Tammy; Thomsen, Preben Dybdahl

2007-01-01

. The hypothesis of this study was that equine MSCs could be isolated from fresh whole equine cord blood. Results: Cord blood was collected from 7 foals immediately after foaling. The mononuclear cell fraction was isolated by Ficoll density centrifugation and cultured in a DMEM low glucose based media at 38.5o......Background: There are no published studies on stem cells from equine cord blood although commercial storage of equine cord blood for future autologous stem cell transplantations is available. Mesenchymal stem cells (MSC) have been isolated from fresh umbilical cord blood of humans collected non......-invasively at the time of birth and from sheep cord blood collected invasively by a surgical intrauterine approach. Mesenchymal stem cells isolation percentage from frozen-thawed human cord blood is low and the future isolation percentage of MSCs from cryopreserved equine cord blood is therefore expectedly low...

Resistance of some leukemic blasts to lysis by lymphokine activated killer (LAK) cells

Energy Technology Data Exchange (ETDEWEB)

Panayotides, P; Sjoegren, A -M; Reizenstein, P; Porwit, A. Immunopathology Lab., Dept. of Pathology, Karolinska Hospital, Stockholm; Wasserman, J

1988-01-01

Peripheral blood mononuclear cells (PBMC) from healthy donors and AML patients in remission were stimulated with phytohemagglutinin (PHA) and recombinant interleukin-2 (IL-2). These stimulated cells (lymphokine activated killer (LAK) cells) showed increased DNA synthesis as measured by /sup 3/H-Thymidine uptake. A synergistic effect of PHA and IL-2 was found. LAK cells' ability to kill acute myeloid leukemia (AML) blasts was investigated by the /sup 51/Cr release assay. LAK cells showed a cytotoxicity (over 10% specific /sup 51/Cr release) against 9/12 leukemic blasts, even at effector/target (E/T) ratios as low as 5:1. However, on average only 22.2% (SD 11.8) and 36.5% (SD 12.5) /sup 51/Cr release were obtained in 4- and 18-hour cytotoxicity assays, respectively, at an E/T ratio of 20:1. Leukemic blasts in 3/12 AML cases and normal PBMC were entirely resistant to lysis, even at an E/T ratio of 80:1. Susceptibility to lysis was not correlated to peanut-agglutinin receptor expression. LAK cells were more cytotoxic towards the K-562 cell line (natural killer activity) than unstimulated PBMC.

Shaft/shaft-seal interface characteristics of a multiple disk centrifugal blood pump.

Science.gov (United States)

Manning, K B; Miller, G E

1999-06-01

A multiple disk centrifugal pump (MDCP) is under investigation as a potential left ventricular assist device. As is the case with most shaft driven pumps, leakage problems around the shaft/shaft seal interface are of major interest. If leakage were to occur during or after implantation, potential events such as blood loss, clotting, blood damage, and/or infections might result in adverse effects for the patient. Because these effects could be quite disastrous, potential shaft and shaft seal materials have been investigated to determine the most appropriate course to limit these effects. Teflon and nylon shaft seals were analyzed as potential candidates along with a stainless steel shaft and a Melonite coated shaft. The materials and shafts were evaluated under various time durations (15, 30, 45, and 60 min), motor speeds (800, 1,000, 1,200, and 1,400 rpm), and outer diameters (1/2 and 3/4 inches). The motor speed and geometrical configurations were typical for the MDCP under normal physiologic conditions. An air and water study was conducted to analyze the inner diameter wear, the inner temperature values, and the outer temperature values. Statistical comparisons were computed for the shaft seal materials, the shafts, and the outer diameters along with the inner and outer temperatures. The conclusions made from the results indicate that both the tested shaft seal materials and shaft materials are not ideal candidates to be used for the MDCP. Teflon experienced a significant amount of wear in air and water studies. Nylon did experience little wear, but heat generation was an evident problem. A water study on nylon was not conducted because of its molecular structure.

Aggregate Formation During the Viral Lysis of a Marine Diatom

Directory of Open Access Journals (Sweden)

Yosuke Yamada

2018-05-01

Full Text Available Recent studies have suggested that the viral lysis of microbes not only facilitates the conversion of particulate organic matter into dissolved organic matter, but also promotes the formation of organic aggregates, which enhance the export of organic carbon from the surface ocean to the deep sea. However, experimental data supporting this proposition are limited. Here, we tested the hypothesis that the viral infection of marine diatoms enhances aggregate formation. We used a model system consisting of Chaetoceros tenuissimus, a bloom-forming diatom with an approximate cell size of 3–10 μm, and a DNA virus, CtenDNAV type II, which replicates in the nucleus of C. tenuissimus. The volume of large particles (50–400 μm in equivalent spherical diameters, determined from photographic images was measured over time (up to 15 days in the diatom-alone control and a virus-added diatom culture. We also determined the concentrations of Coomassie-stainable particles (CSP, proteinaceous particles and transparent exopolymeric particles (TEP, acid-polysaccharide-rich particles with colorimetric methods. The total volume of large particles was significantly higher (5–59 fold in the virus-added diatoms than in the diatom-alone control during the period in which the viral lysis of the diatoms proceeded. One class of large particles produced in the virus-added diatoms was flake-shaped. The flakes were tightly packed and dense, and sank rapidly, possibly playing an important role in the vertical delivery of materials from the surface to the deep sea. The bulk CSP concentrations tended to be higher in the virus-added diatoms than in the diatom-alone control, whereas the reverse was true for the TEP. These results suggest that proteinaceous polymers are involved in aggregate formation. Our data support the emerging notion that the viral lysis of microbes facilitates aggregate formation and the export of organic carbon in the ocean.

Centrifugation protocols: tests to determine optimal lithium heparin and citrate plasma sample quality.

Science.gov (United States)

Dimeski, Goce; Solano, Connie; Petroff, Mark K; Hynd, Matthew

2011-05-01

Currently, no clear guidelines exist for the most appropriate tests to determine sample quality from centrifugation protocols for plasma sample types with both lithium heparin in gel barrier tubes for biochemistry testing and citrate tubes for coagulation testing. Blood was collected from 14 participants in four lithium heparin and one serum tube with gel barrier. The plasma tubes were centrifuged at four different centrifuge settings and analysed for potassium (K(+)), lactate dehydrogenase (LD), glucose and phosphorus (Pi) at zero time, poststorage at six hours at 21 °C and six days at 2-8°C. At the same time, three citrate tubes were collected and centrifuged at three different centrifuge settings and analysed immediately for prothrombin time/international normalized ratio, activated partial thromboplastin time, derived fibrinogen and surface-activated clotting time (SACT). The biochemistry analytes indicate plasma is less stable than serum. Plasma sample quality is higher with longer centrifugation time, and much higher g force. Blood cells present in the plasma lyse with time or are damaged when transferred in the reaction vessels, causing an increase in the K(+), LD and Pi above outlined limits. The cells remain active and consume glucose even in cold storage. The SACT is the only coagulation parameter that was affected by platelets >10 × 10(9)/L in the citrate plasma. In addition to the platelet count, a limited but sensitive number of assays (K(+), LD, glucose and Pi for biochemistry, and SACT for coagulation) can be used to determine appropriate centrifuge settings to consistently obtain the highest quality lithium heparin and citrate plasma samples. The findings will aid laboratories to balance the need to provide the most accurate results in the best turnaround time.

Centrifuge modeling of monopiles

DEFF Research Database (Denmark)

Klinkvort, Rasmus Tofte

2010-01-01

To gain a larger knowledge of the monopile foundation concept, centrifuge modeling is used by the geotechnical group at DTU. The centrifuge operated at DTU is a beam centrifuge and was built in 1976. In the recent years it has been upgraded with onboard data acquisition and control systems....... The capabilities of the centrifuge at DTU makes it possible to obtain a scale factor of 85 in experiments which equals a soil volume in prototype scale of a diameter of 40 meters and a depth of 40 meters. This paper describes centrifuge modeling theory, the centrifuge setup at DTU and as an example show results...... from centrifuge tests performed on large diameter piles installed in dry sand....

Effects of different centrifugation conditions on clinical chemistry and Immunology test results

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Nesic Predrag

2011-05-01

Full Text Available Abstract Background The effect of centrifugation time of heparinized blood samples on clinical chemistry and immunology results has rarely been studied. WHO guideline proposed a 15 min centrifugation time without citing any scientific publications. The centrifugation time has a considerable impact on the turn-around-time. Methods We investigated 74 parameters in samples from 44 patients on a Roche Cobas 6000 system, to see whether there was a statistical significant difference in the test results among specimens centrifuged at 2180 g for 15 min, at 2180 g for 10 min or at 1870 g for 7 min, respectively. Two tubes with different plasma separators (both Greiner Bio-One were used for each centrifugation condition. Statistical comparisons were made by Deming fit. Results Tubes with different separators showed identical results in all parameters. Likewise, excellent correlations were found among tubes to which different centrifugation conditions were applied. Fifty percent of the slopes lay between 0.99 and 1.01. Only 3.6 percent of the statistical tests results fell outside the significance level of p Conclusion A centrifugation time of either 7 or 10 min provided identical test results compared to the time of 15 min as proposed by WHO under the conditions used in our study.

Distribution, recovery and concentration of platelets and leukocytes in L-PRP prepared by centrifugation.

Science.gov (United States)

de Melo, Bruna Alice Gomes; Martins Shimojo, Andréa Arruda; Marcelino Perez, Amanda Gomes; Duarte Lana, José Fabio Santos; Andrade Santana, Maria Helena

2018-01-01

Platelet-rich plasma (PRP) is an autologous product prepared from whole blood (WB) that is widely used in regenerative medicine. In clinical practice, discontinuous centrifugation is used for both hand- and machine-prepared PRP. However, separation of WB fractions via centrifugation is a complex process, and the lack of clear mechanisms limits the understanding and evaluation of PRP preparation methods This paper focuses on the distribution, recovery and concentration factor of platelets and leukocytes in L-PRP (leukocyte and platelet-rich plasma) to define a concentration pattern for these blood components due to centrifugation conditions. WB collected from three healthy donors was centrifuged for 10min at 50-800 xg in a first step and then at 400 xg in a second step. The results from the first centrifugation step showed most platelets to be distributed in the upper layer (UL) and the buffy coat (BC), with approximately 14.5±5.2% retained in the bottom layer (BL). Most leukocytes were present in the BL. The greatest platelet recoveries from L-PRP were obtained at up to 150 xg (88.5±16.9%). The cumulative concentration factors with respect to the WB from the second centrifugation step were 6 and 1.2 for platelets and leukocytes, respectively. Thus, the concentration patterns delineated three centrifugation ranges with platelet/leukocyte ratios of 205±18, 325±15 and 107±4 and lymphocyte/granulocyte ratios of 1.54±0.74, 0.90±0.08 and 0.42±0.07. These findings contribute to a scientifically based standardization of L-PRP preparations. Copyright © 2017 Elsevier B.V. All rights reserved.

CENTRIFUGE END CAP

Science.gov (United States)

Beams, J.W.; Snoddy, L.B.

1960-08-01

An end cap for ultra-gas centrifuges is designed to impart or remove angular momentum to or from the gas and to bring the entering gas to the temperature of the gas inside the centrifuge. The end cap is provided with slots or fins for adjusting the temperature and the angular momentum of the entering gas to the temperature and momentum of the gas in the centrifuge and is constructed to introduce both the inner and the peripheral stream into the centrifuge.

Routine blood cultures in the management of pyelonephritis in pregnancy for improving outcomes.

Science.gov (United States)

Gomi, Harumi; Goto, Yoshihito; Laopaiboon, Malinee; Usui, Rie; Mori, Rintaro

2015-02-13

Pyelonephritis is a type of urinary tract infection (UTI) that affects the upper urinary tract and kidneys, and is one of the most common conditions for hospitalisation among pregnant women, aside from delivery. Samples of urine and blood are obtained and used for cultures as part of the diagnosis and management of the condition. Acute pyelonephritis requires hospitalisation with intravenous administration of antimicrobial agents. Several studies have questioned the necessity of obtaining blood cultures in addition to urine cultures, citing cost and questioning whether blood testing is superfluous. Pregnant women with bacteraemia require a change in the initial empirical treatment based on the blood culture. However, these cases are not common, and represent approximately 15% to 20% of cases. It is unclear whether blood cultures are essential for the effective management of the condition. To assess the effectiveness of routine blood cultures to improve health outcomes in the management of pyelonephritis in pregnant women. We searched the Cochrane Pregnancy and Childbirth Group's Trials Register without language or date restrictions (31 December 2014). Randomised controlled trials and quasi-randomised trials comparing outcomes among pregnant women with pyelonephritis who received initial management with or without blood cultures. Cluster-randomised trials were eligible for inclusion in this review but none were identified. Clinical trials using a cross-over design were not eligible for inclusion. Two review authors independently assessed one trial report for inclusion. We identified one trial report but this was excluded. No clinical trials met the inclusion criteria for this review. There are no large-scale randomised controlled trials to assess outcomes in the management of pyelonephritis in pregnancy with or without blood cultures. Randomised controlled trials are needed to evaluate the effectiveness of managing pyelonephritis in pregnant women with or without

Human cutaneous vascular responses to whole-body tilting, Gz centrifugation, and LBNP

Science.gov (United States)

Watenpaugh, Donald E.; Breit, Gregory A.; Buckley, Theresa M.; Ballard, Richard E.; Murthy, Gita; Hargens, Alan R.

2004-01-01

We hypothesized that gravitational stimuli elicit cardiovascular responses in the following order with gravitational stress equalized at the level of the feet, from lowest to highest response: short-(SAC) and long-arm centrifugation (LAC), tilt, and lower body negative pressure (LBNP). Up to 15 healthy subjects underwent graded application of the four stimuli. Laser-Doppler flowmetry measured regional skin blood flow. At 0.6 G(z) (60 mmHg LBNP), tilt and LBNP similarly reduced leg skin blood flow to approximately 36% of supine baseline levels. Flow increased back toward baseline levels at 80-100 mmHg LBNP yet remained stable during 0.8-1.0 G(z) tilt. Centrifugation usually produced less leg vasoconstriction than tilt or LBNP. Surprisingly, SAC and LAC did not differ significantly. Thigh responses were less definitive than leg responses. No gravitational vasoconstriction occurred in the neck. All conditions except SAC increased heart rate, according to our hypothesized order. LBNP may be a more effective and practical means of simulating cardiovascular effects of gravity than centrifugation.

Centrifugation. A theoretical study of oxygen enrichment by centrifugation

Energy Technology Data Exchange (ETDEWEB)

Kierkegaard, P.; Raetz, E.

1998-12-01

In the present paper we first investigate what happens if we fill a cylinder with air, close it and rotate it. The results show that no matter which peripheral speed is used, it is not possible by means of the radial separation effect alone, to enrich the oxygen concentration from the previous 21% to more then 23.3%, which is of no practical value. In case of a too low enrichment in one centrifuge, the wanted material from this centrifuge can be used as an input for a second centrifuge and so on, in this way forming a cascade of centrifuges. Oxygen will be enriched in each step, until the desired concentration is reached. Cascading was the technology in the very beginning by enrichment plants for uraniumhexaflouride, used for atomic weapons and nuclear power plants. In this study we try to avoid cascading by aiming for higher separation factors. Therefore, we next investigate the possibilities of using a countercurrent centrifuge where in principle the enriched gas is subjected to several centrifugation in the same centrifuge. The calculations show, that in this way it is possible to produce nearly a 100% pure oxygen (polluted with some heavier molecules like argon) in one machine. Our third step was to calculate the amount of oxygen produced per hour. Using a countercurrent centrifuge of the Zippe type, 100 cm high and 20 cm in diameter, it is or will be possible in the near future to produce 17 g enriched air per hour enriched to 50% oxygen. That corresponds to processing 1 m{sup 3} atmospherical air in the period of approximately 24 hours. This is not very impressive. Our fourth step was to estimate the amount of power used for producing this amount of oxygen. A rough, but complicated, estimate shows that the power consumption at the production level will be about the double of the consumption used today. The overall conclusion is, that centrifugation as a production method for oxygen (or nitrogen) will not be competitive with the currently used method in the

Centrifuge enrichment program

International Nuclear Information System (INIS)

Astley, E.R.

1976-01-01

Exxon Nuclear has been active in privately funded research and development of centrifuge enrichment technology since 1972. In October of 1975, Exxon Nuclear submitted a proposal to design, construct, and operate a 3000-MT SWU/yr centrifuge enrichment plant, under the provisions of the proposed Nuclear Fuel Assurance Act of 1975. The U.S. Energy Research and Development Administration (ERDA) accepted the proposal as a basis for negotiation. It was proposed to build a 1000-MT SWU/yr demonstration increment to be operational in 1982; and after successful operation for about one year, expand the facilities into a 3000-MT SWU/yr plant. As part of the overall centrifuge enrichment plant, a dedicated centrifuge manufacturing plant would be constructed; sized to support the full 3000-MT SWU/yr plant. The selection of the centrifuge process by Exxon Nuclear was based on an extremely thorough evaluation of current and projected enrichment technology; results show that the technology is mature and the process will be cost effective. The substantial savings in energy (about 93%) from utilization of the centrifuge option rather than gaseous diffusion is a compelling argument. As part of this program, Exxon Nuclear has a large hardware R and D program, plus a prototype centrifuge manufacturing capability in Malta, New York. To provide a full-scale machine and limited cascade test capability, Exxon Nuclear is constructing a $4,000,000 Centrifuge Test Facility in Richland, Washington. This facility was to initiate operations in the Fall of 1976. Exxon Nuclear is convinced that the centrifuge enrichment process is the rational selection for emergence of a commercial enrichment industry

The homeostasis of Plasmodium falciparum-infected red blood cells.

Directory of Open Access Journals (Sweden)

Jakob M A Mauritz

2009-04-01

Full Text Available The asexual reproduction cycle of Plasmodium falciparum, the parasite responsible for severe malaria, occurs within red blood cells. A merozoite invades a red cell in the circulation, develops and multiplies, and after about 48 hours ruptures the host cell, releasing 15-32 merozoites ready to invade new red blood cells. During this cycle, the parasite increases the host cell permeability so much that when similar permeabilization was simulated on uninfected red cells, lysis occurred before approximately 48 h. So how could infected cells, with a growing parasite inside, prevent lysis before the parasite has completed its developmental cycle? A mathematical model of the homeostasis of infected red cells suggested that it is the wasteful consumption of host cell hemoglobin that prevents early lysis by the progressive reduction in the colloid-osmotic pressure within the host (the colloid-osmotic hypothesis. However, two critical model predictions, that infected cells would swell to near prelytic sphericity and that the hemoglobin concentration would become progressively reduced, remained controversial. In this paper, we are able for the first time to correlate model predictions with recent experimental data in the literature and explore the fine details of the homeostasis of infected red blood cells during five model-defined periods of parasite development. The conclusions suggest that infected red cells do reach proximity to lytic rupture regardless of their actual volume, thus requiring a progressive reduction in their hemoglobin concentration to prevent premature lysis.

Evaluation of the BioFire® FilmArray® Blood Culture Identification Panel on positive blood cultures in a regional hospital laboratory in KwaZulu-Natal

Directory of Open Access Journals (Sweden)

Mokshanand Fhooblall

2016-09-01

Full Text Available Background: There are presently many non-culture-based methods commercially available to identify organisms and antimicrobial susceptibility from blood culture bottles. Each platform has its benefits and limitations. However, there is a need for an improved system with minimal hands-on requirements and short run times. Objectives: In this study, the performance characteristics of the FilmArray® BCID Panel kit were evaluated to assess the efficiency of the kit against an existing system used for identification and antimicrobial susceptibility of organisms from blood cultures. Methods: Positive blood cultures that had initially been received from hospitalised patients of a large quaternary referral hospital in Durban, South Africa were processed as per routine protocol at its Medical Microbiology Laboratory. Positive blood cultures were processed on the FilmArray BCID Panel kit in parallel with the routine sample processing. Inferences were then drawn from results obtained. Results: Organism detection by the FilmArray BCID panel was accurate at 92.6% when organisms that were on the repertoire of the kit were considered, compared to the combination methods (reference method used in the study laboratory. Detection of the antimicrobial resistance markers provided by the panel and reference method demonstrated 100% consistency. Blood cultures with a single organism were accurately identified at 93.8% by FilmArray, while blood cultures with more than one organism were identified at 85.7%. Conclusion: The FilmArray BCID Panel kit is valuable for detection of organisms and markers of antibiotic resistance for an extensive range of organisms.

Comparative evaluation of paired blood culture (aerobic/aerobic) and single blood culture, along with clinical importance in catheter versus peripheral line at a tertiary care hospital.

Science.gov (United States)

Tarai, B; Das, P; Kumar, D; Budhiraja, S

2012-01-01

Paired blood culture (PBC) is uncommon practice in hospitals in India, leading to delayed and inadequate diagnosis. Also contamination remains a critical determinant in hampering the definitive diagnosis. To establish the need of PBC over single blood culture (SBC) along with the degree of contamination, this comparative retrospective study was initiated. We processed 2553 PBC and 4350 SBC in BacT/ALERT 3D (bioMerieux) between October 2010 and June 2011. The positive cultures were identified in VITEK 2 Compact (bioMerieux). True positivity and contaminants were also analyzed in 486 samples received from catheter and peripheral line. Out of 2553 PBC samples, positivity was seen in 350 (13.70%). In 4350 SBC samples, positivity was seen in 200 samples (4.59%). In PBC true pathogens were 267 (10.45%) and contaminants were 83 (3.25%), whereas in SBC 153 (3.51%) were true positives and contaminants were 47 (1.08%). Most of the blood cultures (99.27 %) grew within 72 h and 95.8% were isolated within 48 h. In 486 PBCs received from catheter/periphery (one each), catheter positivity was found in 85 (true positives were 48, false positives 37). In peripheral samples true positives were 50 and false positives were 8. Significantly higher positive rates were seen in PBCs compared with SBCs. Automated blood culture and identification methods significantly reduced the time required for processing of samples and also facilitated yield of diverse/rare organisms. Blood culture from catheter line had higher false positives than peripheral blood culture. Thus every positive result from a catheter must be correlated with clinical findings and requires further confirmation.

Proposing an Empirically Justified Reference Threshold for Blood Culture Sampling Rates in Intensive Care Units

Science.gov (United States)

Castell, Stefanie; Schwab, Frank; Geffers, Christine; Bongartz, Hannah; Brunkhorst, Frank M.; Gastmeier, Petra; Mikolajczyk, Rafael T.

2014-01-01

Early and appropriate blood culture sampling is recommended as a standard of care for patients with suspected bloodstream infections (BSI) but is rarely taken into account when quality indicators for BSI are evaluated. To date, sampling of about 100 to 200 blood culture sets per 1,000 patient-days is recommended as the target range for blood culture rates. However, the empirical basis of this recommendation is not clear. The aim of the current study was to analyze the association between blood culture rates and observed BSI rates and to derive a reference threshold for blood culture rates in intensive care units (ICUs). This study is based on data from 223 ICUs taking part in the German hospital infection surveillance system. We applied locally weighted regression and segmented Poisson regression to assess the association between blood culture rates and BSI rates. Below 80 to 90 blood culture sets per 1,000 patient-days, observed BSI rates increased with increasing blood culture rates, while there was no further increase above this threshold. Segmented Poisson regression located the threshold at 87 (95% confidence interval, 54 to 120) blood culture sets per 1,000 patient-days. Only one-third of the investigated ICUs displayed blood culture rates above this threshold. We provided empirical justification for a blood culture target threshold in ICUs. In the majority of the studied ICUs, blood culture sampling rates were below this threshold. This suggests that a substantial fraction of BSI cases might remain undetected; reporting observed BSI rates as a quality indicator without sufficiently high blood culture rates might be misleading. PMID:25520442

Comparison of the sensitivity of typhi dot test with blood culture in typhoid

Energy Technology Data Exchange (ETDEWEB)

Rizvi, Q [Hamdard College of Medicine, Karachi (Pakistan). Dept. of Pharmacology

2006-10-15

To evaluate the sensitivity of Typhi Dot test in comparison to Blood Culture for the diagnosis of Typhoid Fever in our setup. Fifty patients who fulfilled the clinical criteria of having Typhoid Fever. The data of all the patients was documented, and they were submitted to the Typhi Dot and Blood Culture tests, apart from other routine investigations. Out of the total 50 patients, 47(94%) had their Blood Culture positive for Typhoid bacillus, while in 49 (98%) the Typhi Dot test was positive. Two patients which were found positive on Typhi dot test, gave negative results on Blood Culture. One patient with the signs and symptoms of Typhoid Fever was found neither positive on Typhi Dot test nor upon Blood Culture. There was no significant difference between the results of Blood Culture and Typhi Dot test in the diagnosis of Typhoid Fever. However, Typhi Dot has the advantages of being less expensive and quicker in giving results with excellent sensitivity. (author)

Comparison of the sensitivity of typhi dot test with blood culture in typhoid

International Nuclear Information System (INIS)

Rizvi, Q.

2006-01-01

To evaluate the sensitivity of Typhi Dot test in comparison to Blood Culture for the diagnosis of Typhoid Fever in our setup. Fifty patients who fulfilled the clinical criteria of having Typhoid Fever. The data of all the patients was documented, and they were submitted to the Typhi Dot and Blood Culture tests, apart from other routine investigations. Out of the total 50 patients, 47(94%) had their Blood Culture positive for Typhoid bacillus, while in 49 (98%) the Typhi Dot test was positive. Two patients which were found positive on Typhi dot test, gave negative results on Blood Culture. One patient with the signs and symptoms of Typhoid Fever was found neither positive on Typhi Dot test nor upon Blood Culture. There was no significant difference between the results of Blood Culture and Typhi Dot test in the diagnosis of Typhoid Fever. However, Typhi Dot has the advantages of being less expensive and quicker in giving results with excellent sensitivity. (author)

Blood culture contamination in hospitalized pediatric patients: a single institution experience

Science.gov (United States)

Min, Hyewon; Park, Cheong Soo; Kim, Dong Soo

2014-01-01

Purpose Blood culture is the most important tool for detecting bacteremia in children with fever. However, blood culture contamination rates range from 0.6% to 6.0% in adults; rates for young children have been considered higher than these, although data are limited, especially in Korea. This study determined the contamination rate and risk factors in pediatric patients visiting the emergency room (ER) or being admitted to the ward. Methods We conducted a retrospective chart review of blood cultures obtained from children who visited Yonsei Severance Hospital, Korea between 2006 and 2010. Positive blood cultures were labeled as true bacteremia or contamination according to Centers for Disease Control and Prevention/National Healthcare Safety Network definitions for laboratory-confirmed bloodstream infection, after exclusion of cultures drawn from preexisting central lines only. Results Among 40,542 blood cultures, 610 were positive, of which 479 were contaminations and 131 were true bacteremia (overall contamination rate, 1.18%). The contamination rate in the ER was significantly higher than in the ward (1.32% vs. 0.66%, P6 years, respectively). Conclusion Overall, contamination rates were higher in younger children than in older children, given the difficulty of performing blood sampling in younger children. The contamination rates from the ER were higher than those from the ward, not accounted for only by overcrowding and lack of experience among personnel collecting samples. Further study to investigate other factors affecting contamination should be required. PMID:24868215

Evaluation of Lysis Methods for the Extraction of Bacterial DNA for Analysis of the Vaginal Microbiota.

Science.gov (United States)

Gill, Christina; van de Wijgert, Janneke H H M; Blow, Frances; Darby, Alistair C

2016-01-01

Recent studies on the vaginal microbiota have employed molecular techniques such as 16S rRNA gene sequencing to describe the bacterial community as a whole. These techniques require the lysis of bacterial cells to release DNA before purification and PCR amplification of the 16S rRNA gene. Currently, methods for the lysis of bacterial cells are not standardised and there is potential for introducing bias into the results if some bacterial species are lysed less efficiently than others. This study aimed to compare the results of vaginal microbiota profiling using four different pretreatment methods for the lysis of bacterial samples (30 min of lysis with lysozyme, 16 hours of lysis with lysozyme, 60 min of lysis with a mixture of lysozyme, mutanolysin and lysostaphin and 30 min of lysis with lysozyme followed by bead beating) prior to chemical and enzyme-based DNA extraction with a commercial kit. After extraction, DNA yield did not significantly differ between methods with the exception of lysis with lysozyme combined with bead beating which produced significantly lower yields when compared to lysis with the enzyme cocktail or 30 min lysis with lysozyme only. However, this did not result in a statistically significant difference in the observed alpha diversity of samples. The beta diversity (Bray-Curtis dissimilarity) between different lysis methods was statistically significantly different, but this difference was small compared to differences between samples, and did not affect the grouping of samples with similar vaginal bacterial community structure by hierarchical clustering. An understanding of how laboratory methods affect the results of microbiota studies is vital in order to accurately interpret the results and make valid comparisons between studies. Our results indicate that the choice of lysis method does not prevent the detection of effects relating to the type of vaginal bacterial community one of the main outcome measures of epidemiological studies

Evaluation of Lysis Methods for the Extraction of Bacterial DNA for Analysis of the Vaginal Microbiota.

Directory of Open Access Journals (Sweden)

Christina Gill

Full Text Available Recent studies on the vaginal microbiota have employed molecular techniques such as 16S rRNA gene sequencing to describe the bacterial community as a whole. These techniques require the lysis of bacterial cells to release DNA before purification and PCR amplification of the 16S rRNA gene. Currently, methods for the lysis of bacterial cells are not standardised and there is potential for introducing bias into the results if some bacterial species are lysed less efficiently than others. This study aimed to compare the results of vaginal microbiota profiling using four different pretreatment methods for the lysis of bacterial samples (30 min of lysis with lysozyme, 16 hours of lysis with lysozyme, 60 min of lysis with a mixture of lysozyme, mutanolysin and lysostaphin and 30 min of lysis with lysozyme followed by bead beating prior to chemical and enzyme-based DNA extraction with a commercial kit.After extraction, DNA yield did not significantly differ between methods with the exception of lysis with lysozyme combined with bead beating which produced significantly lower yields when compared to lysis with the enzyme cocktail or 30 min lysis with lysozyme only. However, this did not result in a statistically significant difference in the observed alpha diversity of samples. The beta diversity (Bray-Curtis dissimilarity between different lysis methods was statistically significantly different, but this difference was small compared to differences between samples, and did not affect the grouping of samples with similar vaginal bacterial community structure by hierarchical clustering.An understanding of how laboratory methods affect the results of microbiota studies is vital in order to accurately interpret the results and make valid comparisons between studies. Our results indicate that the choice of lysis method does not prevent the detection of effects relating to the type of vaginal bacterial community one of the main outcome measures of

Positive blood culture with Plasmodium falciparum : Case report

NARCIS (Netherlands)

De Vries, Jutte J. C.; Van Assen, Sander; Mulder, André B.; Kampinga, Greetje A.

2007-01-01

An adult traveler presented with fever and malaise after returning from Sierra Leone. Young trophozoites of Plasmodium falciparum were seen in a blood smear, with parasitemia being 10%. Moreover, blood cultures drawn on admission signaled as "positive" after 1 day of incubation, but no bacteria were

75 FR 70300 - USEC, Inc.; American Centrifuge Lead Cascade Facility; American Centrifuge Plant; Notice of...

Science.gov (United States)

2010-11-17

... Centrifuge Lead Cascade Facility; American Centrifuge Plant; Notice of Receipt of a License Transfer... SNM-2011, for the American Centrifuge Lead Cascade Facility and the American Centrifuge Plant... USEC Inc., (the Licensee), for its American Centrifuge Lead Cascade Facility (LCF) and American...

Optimized methods to measure acetoacetate, 3-hydroxybutyrate, glycerol, alanine, pyruvate, lactate and glucose in human blood using a centrifugal analyser with a fluorimetric attachment

OpenAIRE

Stappenbeck, R.; Hodson, A. W.; Skillen, A. W.; Agius, L.; Alberti, K. G. M. M.

1990-01-01

Optimized methods are described for the analysis of glucose, lactate, pyruvate, alanine, glycerol, D-3-hydroxybutyrate and acetoacetate in perchloric acid extracts of human blood using the Cobas Bio centrifugal analyser. Glucose and lactate are measured using the photometric mode and other metabolites using the fluorimetric mode. The intra-assay coefficients of variation ranged from 0.7 to 4.1%, except with very low levels of pyruvate and acetoacetate where the coefficients of variation were ...

Large-scale preparation of plasmid DNA.

Science.gov (United States)

Heilig, J S; Elbing, K L; Brent, R

2001-05-01

Although the need for large quantities of plasmid DNA has diminished as techniques for manipulating small quantities of DNA have improved, occasionally large amounts of high-quality plasmid DNA are desired. This unit describes the preparation of milligram quantities of highly purified plasmid DNA. The first part of the unit describes three methods for preparing crude lysates enriched in plasmid DNA from bacterial cells grown in liquid culture: alkaline lysis, boiling, and Triton lysis. The second part describes four methods for purifying plasmid DNA in such lysates away from contaminating RNA and protein: CsCl/ethidium bromide density gradient centrifugation, polyethylene glycol (PEG) precipitation, anion-exchange chromatography, and size-exclusion chromatography.

Low-Cost Energy-Efficient 3-D Nano-Spikes-Based Electric Cell Lysis Chips

KAUST Repository

Riaz, Kashif

2017-05-04

Electric cell lysis (ECL) is a promising technique to be integrated with portable lab-on-a-chip without lysing agent due to its simplicity and fast processing. ECL is usually limited by the requirements of high power/voltage and costly fabrication. In this paper, we present low-cost 3-D nano-spikes-based ECL (NSP-ECL) chips for efficient cell lysis at low power consumption. Highly ordered High-Aspect-Ratio (HAR). NSP arrays with controllable dimensions were fabricated on commercial aluminum foils through scalable and electrochemical anodization and etching. The optimized multiple pulse protocols with minimized undesirable electrochemical reactions (gas and bubble generation), common on micro parallel-plate ECL chips. Due to the scalability of fabrication process, 3-D NSPs were fabricated on small chips as well as on 4-in wafers. Phase diagram was constructed by defining critical electric field to induce cell lysis and for cell lysis saturation Esat to define non-ECL and ECL regions for different pulse parameters. NSP-ECL chips have achieved excellent cell lysis efficiencies ηlysis (ca 100%) at low applied voltages (2 V), 2~3 orders of magnitude lower than that of conventional systems. The energy consumption of NSP-ECL chips was 0.5-2 mJ/mL, 3~9 orders of magnitude lower as compared with the other methods (5J/mL-540kJ/mL). [2016-0305

CENTRIFUGE

Science.gov (United States)

Rushing, F.C.

1960-09-01

A vibration damping mechanism for damping vibration forces occurring during the operation of a centrifuge is described. The vibration damping mechanism comprises a plurality of nested spaced cylindrical elements surrounding the rotating shaft of the centrifuge. Some of the elements are held substantially stationary while the others are held with respect to a pair of hearings spaced along the rotating shaft. A fluid is retained about the cylindrical elements.

The utility of anaerobic blood culture in detecting facultative anaerobic bacteremia in children.

Science.gov (United States)

Shoji, Kensuke; Komuro, Hisako; Watanabe, Yasushi; Miyairi, Isao

2013-08-01

Routine anaerobic blood culture is not recommended in children because obligate anaerobic bacteremia is rare in the pediatric population. However, a number of facultative anaerobic bacteria can cause community and hospital acquired infections in children and the utility of anaerobic blood culture for detection of these organisms is still unclear. We conducted a retrospective analysis of all blood culture samples (n = 24,356) at a children's hospital in Japan from October 2009 to June 2012. Among the samples that had paired aerobic and anaerobic blood cultures, 717 samples were considered clinically significant with 418 (58%) organisms detected from both aerobic and anaerobic cultures, 167 (23%) detected only from aerobic culture and 132 (18%) detected only from anaerobic culture. While most facultative anaerobes were detectable by aerobic culture, over 25% of Enterobacteriaceae and 15% of Staphylococcus sp. were detected from anaerobic cultures bottles only, suggesting its potential role in selected settings. Copyright © 2013 Elsevier Inc. All rights reserved.

RESEARCH CENTRIFUGE- ADVANCED TOOL SEPERATION

OpenAIRE

Mahajan Ashwini; Prof. B.V. Jain; Dr Surajj Sarode

2015-01-01

A centrifuge is a critical piece of equipment for the laboratory. Purpose of this study was to study research centrifuge in detail, its applications, uses in different branches and silent features. Their are two types of research centrifuge study here revolutionary research centrifuge and microprocessor research centrifuge. A centrifuge is a device that separates particles from a solution through use of a rotor. In biology, the particles are usually cells, sub cellular organelles, or large mo...

Pipeline system for gas centrifuge

International Nuclear Information System (INIS)

Masumoto, Tsutomu; Umezawa, Sadao.

1977-01-01

Purpose: To enable effective operation for the gas centrifuge cascade system upon failures in the system not by interrupting the operation of all of the centrifuges in the system but by excluding only the failed centrifuges. Constitution: A plurality of gas centrifuges are connected by way of a pipeline and an abnormal detector for the automatic detection of abnormality such as destruction in a vacuum barrel and loss of vacuum is provided to each of the centrifuges. Bypass lines for short-circuitting adjacent centrifuges are provided in the pipelines connecting the centrifuges. Upon generation of abnormality in a centrifuge, a valve disposed in the corresponding bypass is automatically closed or opened by a signal from the abnormal detector to change the gas flow to thereby exclude the centrifuge in abnormality out of the system. This enables to effectively operate the system without interrupting the operation for the entire system. (Moriyama, K.)

Clinical comparison of two commercial blood culture systems

NARCIS (Netherlands)

Spanjaard, L.; Kuijper, E. J.; Dankert, J.

2000-01-01

A prospective, volume-controlled comparison of the BacT/Alert FAN (Organon Teknika, USA) and Vital (bioMérieux, France) blood culture systems was performed in a university hospital during a period of 11 months. Twenty to 40 ml of blood drawn from an adult patient was distributed equally between a

Plasma centrifuges

International Nuclear Information System (INIS)

Karchevskij, A.I.; Potanin, E.P.

2000-01-01

The review of the most important studies on the isotope separation processes in the rotating plasma is presented. The device is described and the characteristics of operation of the pulse plasma centrifuges with weakly and strongly ionized plasma as well as the stationary plasma centrifuges with the medium weak ionization and devices, applying the stationary vacuum arc with the high ionization rate and the stationary beam-plasma discharge with complete ionization, are presented. The possible mechanisms of the isotope separation in plasma centrifuges are considered. The specific energy consumption for isotope separation in these devices is discussed [ru

Changes in clot lysis levels of reteplase and streptokinase following continuous wave ultrasound exposure, at ultrasound intensities following attenuation from the skull bone

Directory of Open Access Journals (Sweden)

Roijer Anders

2008-08-01

Full Text Available Abstract Background Ultrasound (US has been used to enhance thrombolytic therapy in the treatment of stroke. Considerable attenuation of US intensity is however noted if US is applied over the temporal bone. The aim of this study was therefore to explore possible changes in the effect of thrombolytic drugs during low-intensity, high-frequency continuous-wave ultrasound (CW-US exposure. Methods Clots were made from fresh venous blood drawn from healthy volunteers. Each clot was made from 1.4 ml blood and left to coagulate for 1 hour in a plastic test-tube. The thrombolytic drugs used were, 3600 IU streptokinase (SK or 0.25 U reteplase (r-PA, which were mixed in 160 ml 0.9% NaCl solution. Continuous-wave US exposure was applied at a frequency of 1 MHz and intensities ranging from 0.0125 to 1.2 W/cm2. For each thrombolytic drug (n = 2, SK and r-PA and each intensity (n = 9 interventional clots (US-exposed, n = 6 were submerged in thrombolytic solution and exposed to CW-US while control clots (also submerged in thrombolytic solution, n = 6 were left unexposed to US. To evaluate the effect on clot lysis, the haemoglobin (Hb released from each clot was measured every 20 min for 1 hour (20, 40 and 60 min. The Hb content (mg released was estimated by spectrophotometry at 540 nm. The difference in effect on clot lysis was expressed as the difference in the amount of Hb released between pairs of US-exposed clots and control clots. Statistical analysis was performed using Wilcoxon's signed rank test. Results Continuous-wave ultrasound significantly decreased the effects of SK at intensities of 0.9 and 1.2 W/cm2 at all times (P 2 and at 1.2 W/cm2, following 40 min exposure at 0.3, 0.6, 0.9 and at 1.2 W/cm2, and following 60 min of exposure at 0.05 0.3, 0.6, 0.9 and at 1.2 W/cm2 (all P Conclusion Increasing intensities of CW-US exposure resulted in increased clot lysis of r-PA-treated blood clots, but decreased clot lysis of SK-treated clots.

Comparison of the clinical and microbiological characteristics of Campylobacter and Helicobacter bacteremia: the importance of time to blood culture positivity using the BACTEC blood culture systems.

Science.gov (United States)

Yamamoto, Kei; Hayakawa, Kayoko; Nagashima, Maki; Shimada, Kayo; Kutsuna, Satoshi; Takeshita, Nozomi; Kato, Yasuyuki; Kanagawa, Shuzo; Yamada, Koji; Mezaki, Kazuhisa; Kirikae, Teruo; Ohmagari, Norio

2017-11-28

Campylobacter spp. and Helicobacter spp. are rare but important causes of bacteremia in humans. Distinguishing these bacteria is complicated because of their similar phenotypic profiles. We conducted clinical and microbiological investigations of Campylobacter spp. or Helicobacter spp. bacteremia. Patients diagnosed with bacteremia from 2008 to 2014 were included. The clinical and microbiological characteristics of Campylobacter spp. and Helicobacter spp. bacteremia were compared. The BACTEC system was used in blood cultures. A receiver operating characteristic curve was plotted based on the time to blood culture positivity. Sixteen cases of Helicobacter spp. bacteremia (patient age: 61 ± 18 years) and 14 cases of Campylobacter spp. bacteremia (patient age: 49 ± 21 years) were identified. Median time to blood culture positivity was longer for the Helicobacter spp. cases than the Campylobacter spp. cases (91.4 h vs 55.3 h, p culture positivity > 75 h predicted Helicobacter spp. bacteremia with a sensitivity of 0.88 and a specificity of 0.93 (area under the receiver operating characteristic curve of 0.90). In conclusion, a time to blood culture positivity was useful in distinguishing Helicobacter spp. bacteremia from Campylobacter spp. bacteremia.

Simple method for culture of peripheral blood lymphocytes of Testudinidae.

Science.gov (United States)

Silva, T L; Silva, M I A; Venancio, L P R; Zago, C E S; Moscheta, V A G; Lima, A V B; Vizotto, L D; Santos, J R; Bonini-Domingos, C R; Azeredo-Oliveira, M T V

2011-12-06

We developed and optimized a simple, efficient and inexpensive method for in vitro culture of peripheral blood lymphocytes from the Brazilian tortoise Chelonoidis carbonaria (Testudinidae), testing various parameters, including culture medium, mitogen concentration, mitotic index, culture volume, incubation time, and mitotic arrest. Peripheral blood samples were obtained from the costal vein of four couples. The conditions that gave a good mitotic index were lymphocytes cultured at 37°C in minimum essential medium (7.5 mL), with phytohemagglutinin as a mitogen (0.375 mL), plus streptomycin/penicillin (0.1 mL), and an incubation period of 72 h. Mitotic arrest was induced by 2-h exposure to colchicine (0.1 mL), 70 h after establishing the culture. After mitotic arrest, the cells were hypotonized with 0.075 M KCl for 2 h and fixed with methanol/acetic acid (3:1). The non-banded mitotic chromosomes were visualized by Giemsa staining. The diploid chromosome number of C. carbonaria was found to be 52 in females and males, and sex chromosomes were not observed. We were able to culture peripheral blood lymphocytes of a Brazilian tortoise in vitro, for the preparation of mitotic chromosomes.

Storage time of platelet concentrates and risk of a positive blood culture

DEFF Research Database (Denmark)

Kreuger, Aukje L; Rostgaard, Klaus; Middelburg, Rutger A

2018-01-01

BACKGROUND: Concern of transfusion-transmitted bacterial infections has been the major hurdle to extend shelf life of platelet (PLT) concentrates. We aimed to investigate the association between storage time and risk of positive blood cultures at different times after transfusion. STUDY DESIGN...... AND METHODS: We performed a nationwide cohort study among PLT transfusion recipients in Denmark between 2010 and 2012, as recorded in the Scandinavian Donations and Transfusions (SCANDAT2) database. Linking with a nationwide database on blood cultures (MiBa), we compared the incidence of a positive blood......) of a positive blood culture the day after transfusion of at least one old PLT concentrate was 0.77 (95% confidence interval [CI], 0.54-1.09) compared to transfusion of fresh PLT concentrates. The incidence rate of a positive blood culture was lower the day after receiving one old compared to one fresh PLT...

A Comparative Study of Blood Culture Sampling from Umbilical Catheter Line versus Peripheral Site

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Abdolkarim Hamedi

2010-08-01

Full Text Available Neonatal sepsis is an important cause of death and morbidity in newborns and is diagnosed by isolation of organism in blood culture. In several reports,reliablity of blood cultures were done from umbi lical catheters,have been demonstrated. The objective of the present study was to determine,wether an inde welling umbilical catheter, could be an alternative site for blood culture. In a prospective study over 6 months during 2006,141 paired blood cultures from 134 infant,were done simultaneously from peripheral site and umbilical catheter (mostly U. V. C,during the first four days of life. Majority of these infants were preterm and admitted to NICU for special care. these infants had indwelling umbilical line and had indication of sepsis workup. A total of 141 pairs of blood cultures were obtained from 134 infants. In 16 infants blood culture pairs were positive for one organism in both peripheral vein and umbilical site. 71. 6% of total cultures (n=11pairs were negative in boths site. A total of 22 pairs were positive in one site only,with 5 positive from peripheral vein only and the other 17 from umblical site. Two pairs were positve in boths site with two different organism. In over all 16 infant (11%of blood were considered to be contaminated. Contamination rate were 2. 4% and 9. 2% for peripheral and umbilical catheter site. Contamination rate increased after 48 hours of age in umbilical catheter. The result showed that after 2 days contamination rate for blood culture taken from catheter line increased and specifity decreased. We recommended that blood culture via umblical catheter in first 2 days in sick neonates with indwelling catheter can be a alternate site of blood culture sampelling.

Enrichment of reticulocytes from whole blood using aqueous multiphase systems of polymers.

Science.gov (United States)

Kumar, Ashok A; Lim, Caeul; Moreno, Yovany; Mace, Charles R; Syed, Abeer; Van Tyne, Daria; Wirth, Dyann F; Duraisingh, Manoj T; Whitesides, George M

2015-01-01

This paper demonstrates the enrichment of reticulocytes by centrifuging whole blood through aqueous multiphase systems (AMPSs)-immiscible phases of solutions of polymers that form step-gradients in density. The interfaces of an AMPS concentrate cells; this concentration facilitates the extraction of blood enriched for reticulocytes. AMPS enrich reticulocytes from blood from both healthy and hemochromatosis donors. Varying the osmolality and density of the phases of AMPS provides different levels of enrichment and yield of reticulocytes. A maximum enrichment of reticulocytemia of 64 ± 3% was obtained from donors with hemochromatosis. When used on peripheral blood from normal donors, AMPS can provide a higher yield of enriched reticulocytes and a higher proportion of reticulocytes expressing CD71 than differential centrifugation followed by centrifugation over Percoll. Blood enriched for reticulocytes by AMPS could be useful for research on malaria. Several species of malaria parasites show a preference to invade young erythrocytes and reticulocytes; this preference complicates in vitro cultivation of these species in human blood. Plasmodium knowlesi malaria parasites invade normal human blood enriched for reticulocytes by AMPSs at a rate 2.2 times greater (P blood. Parasite invasion in normal blood enriched by AMPS was 1.8 times greater (P blood enriched to a similar reticulocytemia by differential centrifugation followed by centrifugation over Percoll. The enrichment of reticulocytes that are invaded by malaria parasites demonstrates that AMPSs can provide a label-free method to enrich cells for biological research. © 2014 Wiley Periodicals, Inc.

Study on chromosome aberrations test determinated by micro-whole blood culture in vacuum blood collection tube

International Nuclear Information System (INIS)

Zhong Zhihong; Han Fang'an; Ge Qinjuan; Wu Xiao; Chen Juan

2006-01-01

Objective: To develop an easier and efficient method of culturing the chromosome and analyzing the aberrations in peripheral lymphocytes. Methods: Micro whole was cultured for 54 hours in home-made vacuum blood collection tube, and then collection, slice-making, microscopy detection for the chromosome aberrations was done. The difference of the results was analysed by comparing with the common method. Results: For 60 radiologists and 30 contrasts, the chromosome aberrations in peripheral lymphocytes were examed by this system, the lymphocytes and chromosome were clear and alive and easier to analyse. Compared with the common method, there was no significantly difference between the two analyzing results. Conclusion: The chromosome aberrations test by micro whole blood culture in vacuum blood collection tube is easier and efficient, and is worthy of being widely popularized. (authors)

Profiling post-centrifugation delay of serum and plasma with antibody bead arrays.

Science.gov (United States)

Qundos, Ulrika; Hong, Mun-Gwan; Tybring, Gunnel; Divers, Mark; Odeberg, Jacob; Uhlen, Mathias; Nilsson, Peter; Schwenk, Jochen M

2013-12-16

Several biobanking initiatives have emerged to create extensive collections of specimen for biomedical studies and various analytical platforms. An affinity proteomic analysis with antibody suspension bead arrays was conducted to investigate the influence of the pre-analytical time and temperature conditions on blood derived samples. Serum and EDTA plasma prepared from 16 individuals was centrifuged and aliquots were kept either at 4°C or in ambient temperature for 1h and up to 36h prior to first storage. Multiplexed protein profiles of post-centrifugation delay were generated in 384 biotinylated samples using 373 antibodies that targeted 343 unique proteins. Very few profiles were observed as significantly altered by the studied temperature and time intervals. Single binder and sandwich assays revealed decreasing levels of caldesmon 1 (CALD1) related to EDTA standard tubes and prolonged post-centrifugation delay of 36h. Indications from changes in CALD1 levels require further confirmation in independent material, but the current data suggests that samples should preferentially be frozen during the day of collection when to be profiled with antibody arrays selected for this study. Affinity-based profiling of serum and plasma by microarray assays can provide unique opportunities for the discovery of biomarkers. It is though often not known how differences in sample handling after collection influence the downstream analysis. By profiling three types of blood preparations for alterations in protein profiles with respect to time and temperature post centrifugation, we addressed an important component in the analysis and of such specimen. We believe that this analysis adds valuable information to be considered when biobanking blood derived samples. This article is part of a Special Issue entitled: Standardization and Quality Control in Proteomics. Copyright © 2013 The Authors. Published by Elsevier B.V. All rights reserved.

Fresenius AS.TEC204 blood cell separator.

Science.gov (United States)

Sugai, Mikiya

2003-02-01

Fresenius AS.TEC204 is a third-generation blood cell separator that incorporates the continuous centrifugal separation method and automatic control of the cell separation process. Continuous centrifugation separates cell components according to their specific gravity, and different cell components are either harvested or eliminated as needed. The interface between the red blood cell and plasma is optically detected, and the Interface Control (IFC) cooperates with different pumps, monitors and detectors to harvest required components automatically. The system is composed of three major sections; the Front Panel Unit; the Pump Unit, and the Centrifuge Unit. This unit can be used for a wide variety of clinical applications including collection of platelets, peripheral blood stem cells, bone marrow stem cells, granulocytes, mononuclear cells, and exchange of plasma or red cells, and for plasma treatment.

Lysis-deficient phages as novel therapeutic agents for controlling bacterial infection

Directory of Open Access Journals (Sweden)

Kempashanaiah Nanjundappa

2011-08-01

Full Text Available Abstract Background Interest in phage therapy has grown over the past decade due to the rapid emergence of antibiotic resistance in bacterial pathogens. However, the use of bacteriophages for therapeutic purposes has raised concerns over the potential for immune response, rapid toxin release by the lytic action of phages, and difficulty in dose determination in clinical situations. A phage that kills the target cell but is incapable of host cell lysis would alleviate these concerns without compromising efficacy. Results We developed a recombinant lysis-deficient Staphylococcus aureus phage P954, in which the endolysin gene was rendered nonfunctional by insertional inactivation. P954, a temperate phage, was lysogenized in S. aureus strain RN4220. The native endolysin gene on the prophage was replaced with an endolysin gene disrupted by the chloramphenicol acetyl transferase (cat gene through homologous recombination using a plasmid construct. Lysogens carrying the recombinant phage were detected by growth in presence of chloramphenicol. Induction of the recombinant prophage did not result in host cell lysis, and the phage progeny were released by cell lysis with glass beads. The recombinant phage retained the endolysin-deficient genotype and formed plaques only when endolysin was supplemented. The host range of the recombinant phage was the same as that of the parent phage. To test the in vivo efficacy of the recombinant endolysin-deficient phage, immunocompromised mice were challenged with pathogenic S. aureus at a dose that results in 80% mortality (LD80. Treatment with the endolysin-deficient phage rescued mice from the fatal S. aureus infection. Conclusions A recombinant endolysin-deficient staphylococcal phage has been developed that is lethal to methicillin-resistant S. aureus without causing bacterial cell lysis. The phage was able to multiply in lytic mode utilizing a heterologous endolysin expressed from a plasmid in the propagation host

[EXPRESS IDENTIFICATION OF POSITIVE BLOOD CULTURES USING DIRECT MALDI-TOF MASS SPECTROMETRY].

Science.gov (United States)

Popov, D A; Ovseenko, S T; Vostrikova, T Yu

2015-01-01

To evaluate the effectiveness of direct identification of pathogens of bacteremia by direct matrix assisted laser desorption ionization time-flight mass spectrometry (mALDI-TOF) compared to routine method. A prospective study included 211 positive blood cultures obtained from 116 patients (106 adults and 10 children, aged from 2 weeks to 77 years old in the ICU after open heart surgery. Incubation was carried out under aerobic vials with a sorbent for antibiotics Analyzer BacT/ALERT 3D 120 (bioMerieux, France) in parallel with the primary sieving blood cultures on solid nutrient media with subsequent identification of pure cultures using MALDI-TOF mass spectrometry analyzer Vitek MS, bioMerieux, France routine method), after appropriate sample preparation we carried out a direct (without screening) MALDI-TOF mass spectrometric study of monocomponental blood cultures (n = 201). using a routine method in 211 positive blood cultures we identified 23 types of microorganisms (Staphylococcus (n = 87), Enterobacteria- ceae (n = 71), Enterococci (n = 20), non-fermentative Gram-negative bacteria (n = 18), others (n = 5). The average time of incubation of samples to obtain a signal of a blood culture growth was 16.2 ± 7.4 h (from 3.75 to 51 hours.) During the first 12 hours of incubation, growth was obtained in 32.4% of the samples, and on the first day in 92.2%. In the direct mass spectrometric analysis mnonocomponental blood cultures (n = 201) is well defined up to 153 species of the sample (76.1%), while the share of successful identification of Gram-negative bacteria was higher than that of Gram-positive (85.4 and 69, 1%, respectively p = 0.01). The high degree of consistency in the results of standard and direct method of identifying blood cultures using MALDI-TOF mass spectrometry (κ = 0.96, p direct mass spectrometric analysis, including sample preparation, was no longer than 1 hour: The method of direct MALDI-TOF mass spectrometry allows to significantly speed up

76 FR 9613 - USEC Inc. (American Centrifuge Lead Cascade Facility and American Centrifuge Plant); Order...

Science.gov (United States)

2011-02-18

... NUCLEAR REGULATORY COMMISSION [EA-11-013] USEC Inc. (American Centrifuge Lead Cascade Facility and American Centrifuge Plant); Order Approving Direct Transfer of Licenses and Conforming Amendment I USEC... Centrifuge Lead Cascade Facility (Lead Cascade) and American Centrifuge Plant (ACP), respectively, which...

77 FR 9273 - USEC Inc. (American Centrifuge Lead Cascade Facility and American Centrifuge Plant); Direct...

Science.gov (United States)

2012-02-16

... NUCLEAR REGULATORY COMMISSION [NRC-2010-0355] USEC Inc. (American Centrifuge Lead Cascade Facility and American Centrifuge Plant); Direct Transfer of Licenses In the Matter of USEC INC. (American Centrifuge Lead Cascade Facility and American Centrifuge Plant); Order EA-12- [[Page 9274

Gas Centrifuges and Nuclear Proliferation

Energy Technology Data Exchange (ETDEWEB)

Albright, David

2004-09-15

Gas centrifuges have been an ideal enrichment method for a wide variety of countries. Many countries have built gas centrifuges to make enriched uranium for peaceful nuclear purposes. Other countries have secretly sought centrifuges to make highly enriched uranium for nuclear weapons. In more recent times, several countries have secretly sought or built gas centrifuges in regions of tension. The main countries that have been of interest in the last two decades have been Pakistan, Iraq, Iran, and North Korea. Currently, most attention is focused on Iran, Pakistan, and North Korea. These states did not have the indigenous abilities to make gas centrifuges, focusing instead on illicit and questionable foreign procurement. The presentation covered the following main sections: Spread of centrifuges through illicit procurement; Role of export controls in stopping proliferation; Increasing the transparency of gas centrifuge programs in non-nuclear weapon states; and, Verified dismantlement of gas centrifuge programs. Gas centrifuges are important providers of low enriched uranium for civil nuclear power reactors. They also pose special nuclear proliferation risks. We all have special responsibilities to prevent the spread of gas centrifuges into regions of tension and to mitigate the consequences of their spread into the Middle East, South Asia, and North Asia.

Improved Diagnosis of Prosthetic Joint Infection by Culturing Periprosthetic Tissue Specimens in Blood Culture Bottles

Directory of Open Access Journals (Sweden)

Trisha N. Peel

2016-01-01

Full Text Available Despite known low sensitivity, culture of periprosthetic tissue specimens on agars and in broths is routine. Culture of periprosthetic tissue samples in blood culture bottles (BCBs is potentially more convenient, but it has been evaluated in a limited way and has not been widely adopted. The aim of this study was to compare the sensitivity and specificity of inoculation of periprosthetic tissue specimens into blood culture bottles with standard agar and thioglycolate broth culture, applying Bayesian latent class modeling (LCM in addition to applying the Infectious Diseases Society of America (IDSA criteria for prosthetic joint infection. This prospective cohort study was conducted over a 9-month period (August 2013 to April 2014 at the Mayo Clinic, Rochester, MN, and included all consecutive patients undergoing revision arthroplasty. Overall, 369 subjects were studied; 117 (32% met IDSA criteria for prosthetic joint infection, and 82% had late chronic infection. Applying LCM, inoculation of tissues into BCBs was associated with a 47% improvement in sensitivity compared to the sensitivity of conventional agar and broth cultures (92.1 versus 62.6%, respectively; this magnitude of change was similar when IDSA criteria were applied (60.7 versus 44.4%, respectively; P = 0.003. The time to microorganism detection was shorter with BCBs than with standard media (P < 0.0001, with aerobic and anaerobic BCBs yielding positive results within a median of 21 and 23 h, respectively. Results of our study demonstrate that the semiautomated method of periprosthetic tissue culture in blood culture bottles is more sensitive than and as specific as agar and thioglycolate broth cultures and yields results faster.

Trypanosoma cruzi. Surface antigens of blood and culture forms

International Nuclear Information System (INIS)

Nogueira, N.; Chaplan, S.; Tydings, J.D.; Unkeless, J.; Cohn, Z.

1981-01-01

The surface polypeptides of both cultured and blood forms of Trypanosoma cruzi were iodinated by the glucose oxidase-lactoperoxidase technique. Blood-form trypomastigotes (BFT) isolated form infected mice displayed a major 90,000-Mr component. In contrast, both epimastigotes and trypomastigotes obtained form acellular cultures expressed a smaller 75,000-Mr peptide. Both major surface components were presumably glycoproteins in terms of their binding to concanavalin A-Sepharose 4B. Within a 3-h period, both blood and culture forms synthesized their respective surface glycoproteins (90,000 Mr and 75,000 Mr, respectively in vitro. [/sub 35/S]methionine-labeled surface peptides were immunoprecipitated with immune sera of both human and murine origin. A panel of sera form patients with chronic Chagas' disease and hyperimmunized mice recognized similar surface peptides. These immunogens were the same components as the major iodinated species. The major BFT surface peptide was readily removed by trypsin treatment of the parasites, although the procedure did not affect the 75,000-Mr peptide from the culture forms. Two-dimensional polyacrylamide gel electrophoresis revealed that the 90,000-Mr peptide found on BFT was an acidic protein of isoelectric point (pI) 5.0, whereas, the 75,000-Mr peptide form culture-form trypomastigotes has a pI of 7.2. The 90,000-Mr component is thought to be responsible for the anti-phagocytic properties of the BFT

SEAL FOR HIGH SPEED CENTRIFUGE

Science.gov (United States)

Skarstrom, C.W.

1957-12-17

A seal is described for a high speed centrifuge wherein the centrifugal force of rotation acts on the gasket to form a tight seal. The cylindrical rotating bowl of the centrifuge contains a closure member resting on a shoulder in the bowl wall having a lower surface containing bands of gasket material, parallel and adjacent to the cylinder wall. As the centrifuge speed increases, centrifugal force acts on the bands of gasket material forcing them in to a sealing contact against the cylinder wall. This arrangememt forms a simple and effective seal for high speed centrifuges, replacing more costly methods such as welding a closure in place.

Tumor Lysis Syndrome (TLS following intrathecal chemotherapy in a child with acute myelogenous leukemia (AML

Directory of Open Access Journals (Sweden)

Chana L. Glasser, MD

2017-01-01

Full Text Available Tumor Lysis Syndrome (TLS is a well-known complication of induction therapy for hematologic malignancies. It is characterized by rapid breakdown of malignant white blood cells (WBCs leading to metabolic derangements and serious morbidity if left untreated. Most commonly, TLS is triggered by systemic chemotherapy, however, there have been case reports of TLS following intrathecal (IT chemotherapy, all in patients with acute lymphoblastic leukemia (ALL/lymphoma. Here, we report the first case of a patient with acute myelogenous leukemia (AML who developed TLS following a single dose of IT cytosine arabinoside (Ara-C.

Effect of a training programme on blood culture contamination rate in critical care.

Science.gov (United States)

Sánchez-Sánchez, M M; Arias-Rivera, S; Fraile-Gamo, P; Jareño-Collado, R; López-Román, S; Vadillo-Obesso, P; García-González, S; Pulido-Martos, M T; Sánchez-Muñoz, E I; Cacho-Calvo, J; Martín-Pellicer, A; Panadero-Del Olmo, L; Frutos-Vivar, F

2018-03-30

Blood culture contamination can occur from extraction to processing; its rate should not exceed 3%. To evaluate the impact of a training programme on the rate of contaminated blood cultures after the implementation of sample extraction recommendations based on the best evidence. Prospective before-after study in a polyvalent intensive care unit with 18 beds. Two phases were established (January-June 2012, October 2012-October 2015) with a training period between them. Main recommendations: sterile technique, surgical mask, double skin disinfection (70° alcohol and 2% alcoholic chlorhexidine), 70° alcohol disinfection of culture flasks and injection of samples without changing needles. Including all blood cultures of patients with extraction request. demographic, severity, pathology, reason for admission, stay and results of blood cultures (negative, positive and contaminated). Basic descriptive statistics: mean (standard deviation), median (interquartile range) and percentage (95% confidence interval). Calculated contamination rates per 100 blood cultures extracted. Bivariate analysis between periods. Four hundred and eight patients were included. Eight hundred and forty-one blood cultures were taken, 33 of which were contaminated. In the demographic variables, severity, diagnosis and stay of patients with contaminated samples, no differences were observed from those with uncontaminated samples. Pre-training vs post-training contamination rates: 14 vs 5.6 per 100 blood cultures extracted (P=.00003). An evidence-based training programme reduced the contamination of samples. It is necessary to continue working on the planning of activities and care to improve the detection of pollutants and prevent contamination of samples. Copyright © 2018 Sociedad Española de Enfermería Intensiva y Unidades Coronarias (SEEIUC). Publicado por Elsevier España, S.L.U. All rights reserved.

Biological variation in tPA-induced plasma clot lysis time.

Science.gov (United States)

Talens, Simone; Malfliet, Joyce J M C; Rudež, Goran; Spronk, Henri M H; Janssen, Nicole A H; Meijer, Piet; Kluft, Cornelis; de Maat, Moniek P M; Rijken, Dingeman C

2012-10-01

Hypofibrinolysis is a risk factor for venous and arterial thrombosis, and can be assessed by using a turbidimetric tPA-induced clot lysis time (CLT) assay. Biological variation in clot lysis time may affect the interpretation and usefulness of CLT as a risk factor for thrombosis. Sufficient information about assay variation and biological variation in CLT is not yet available. Thus, this study aimed to determine the analytical, within-subject and between-subject variation in CLT. We collected blood samples from 40 healthy individuals throughout a period of one year (average 11.8 visits) and determined the CLT of each plasma sample in duplicate. The mean (± SD) CLT was 83.8 (± 11.1) minutes. The coefficients of variation for total variation, analytical variation, within-subject variation and between-subject variation were 13.4%, 2.6%, 8.2% and 10.2%, respectively. One measurement can estimate the CLT that does not deviate more than 20% from its true value. The contribution of analytical variation to the within-subject variation was 5.0%, the index of individuality was 0.84 and the reference change value was 23.8%. The CLT was longer in the morning compared to the afternoon and was slightly longer in older individuals (> 40 years) compared to younger (≤40 years) individuals. There was no seasonal variation in CLT and no association with air pollution. CLT correlated weakly with fibrinogen, C-reactive protein, prothrombin time and thrombin generation. This study provides insight into the biological variation of CLT, which can be used in future studies testing CLT as a potential risk factor for thrombosis.

Hemocompatibility of Axial Versus Centrifugal Pump Technology in Mechanical Circulatory Support Devices.

Science.gov (United States)

Schibilsky, David; Lenglinger, Matthias; Avci-Adali, Meltem; Haller, Christoph; Walker, Tobias; Wendel, Hans Peter; Schlensak, Christian

2015-08-01

The hemocompatible properties of rotary blood pumps commonly used in mechanical circulatory support (MCS) are widely unknown regarding specific biocompatibility profiles of different pump technologies. Therefore, we analyzed the hemocompatibility indicating markers of an axial flow and a magnetically levitated centrifugal device within an in vitro mock loop. The HeartMate II (HM II; n = 3) device and a CentriMag (CM; n = 3) adult pump were investigated in a human whole blood mock loop for 360 min using the MCS devices as a driving component. Blood samples were analyzed by enzyme-linked immunosorbent assay for markers of coagulation, complement system, and inflammatory response. There was a time-dependent activation of the coagulation (thrombin-antithrombin complexes [TAT]), complement (SC5b-9), and inflammation system (polymorphonuclear [PMN] elastase) in both groups. The mean value of TAT (CM: 4.0 μg/L vs. 29.4 μg/L, P technologies and a magnetically levitated centrifugal pump design might be superior. Copyright © 2015 International Center for Artificial Organs and Transplantation and Wiley Periodicals Inc.

Tumor lysis syndrome in a patient with metastatic colon cancer after treatment with oxaliplatin and 5-Fu

Directory of Open Access Journals (Sweden)

Ruo-Han Tseng

2016-12-01

Full Text Available Tumor lysis syndrome in solid tumors is a rare occurrence, with a poor prognosis. We present the case of a patient of recurrent colon cancer who received chemotherapy with FOLFOX regimen (lencovorin, fluorouracil, and oxaliplatin with subsequent tumor lysis. We present a recurrent rectal cancer patient suffered from tumor lysis syndrome after salvage FOLFOX regimen. After treat with CVVH with improved conscious status. In this case report, we had review the tumor lysis in solid tumor.

Effects of different centrifugation conditions on clinical chemistry and Immunology test results.

Science.gov (United States)

Minder, Elisabeth I; Schibli, Adrian; Mahrer, Dagmar; Nesic, Predrag; Plüer, Kathrin

2011-05-10

The effect of centrifugation time of heparinized blood samples on clinical chemistry and immunology results has rarely been studied. WHO guideline proposed a 15 min centrifugation time without citing any scientific publications. The centrifugation time has a considerable impact on the turn-around-time. We investigated 74 parameters in samples from 44 patients on a Roche Cobas 6000 system, to see whether there was a statistical significant difference in the test results among specimens centrifuged at 2180 g for 15 min, at 2180 g for 10 min or at 1870 g for 7 min, respectively. Two tubes with different plasma separators (both Greiner Bio-One) were used for each centrifugation condition. Statistical comparisons were made by Deming fit. Tubes with different separators showed identical results in all parameters. Likewise, excellent correlations were found among tubes to which different centrifugation conditions were applied. Fifty percent of the slopes lay between 0.99 and 1.01. Only 3.6 percent of the statistical tests results fell outside the significance level of p < 0.05, which was less than the expected 5%. This suggests that the outliers are the result of random variation and the large number of statistical tests performed. Further, we found that our data are sufficient not to miss a biased test (beta error) with a probability of 0.10 to 0.05 in most parameters. A centrifugation time of either 7 or 10 min provided identical test results compared to the time of 15 min as proposed by WHO under the conditions used in our study.

Diagnosis of blood culture-negative endocarditis and clinical comparison between blood culture-negative and blood culture-positive cases.

Science.gov (United States)

Lamas, Cristiane C; Fournier, Pierre-Edouard; Zappa, Monica; Brandão, Tatiana J D; Januário-da-Silva, Carolina A; Correia, Marcelo G; Barbosa, Giovanna Ianini F; Golebiovski, Wilma F; Weksler, Clara; Lepidi, Hubert; Raoult, Didier

2016-08-01

To analyze the clinical characteristics of blood culture-negative endocarditis (BCNE) and how it compares to those of blood culture-positive endocarditis (BCPE) cases and show how molecular tools helped establish the etiology in BCNE. Adult patients with definite infective endocarditis (IE) and having valve surgery were included. Valves were studied by polymerase chain reaction (PCR). Statistical analysis compared BCNE and BCPE. One hundred and thirty-one patients were included; 53 (40 %) had BCNE. The mean age was 45 ± 16 years; 33 (62 %) were male. BCNE was community-acquired in 41 (79 %). Most patients were referred from other hospitals (38, 73 %). Presentation was subacute in 34 (65 %), with fever in 47/53 (90 %) and a new regurgitant murmur in 34/42 (81 %). Native valves were affected in 74 %, mostly left-sided. All echocardiograms showed major criteria for IE. Antibiotics were used prior to BC collection in 31/42 (74 %). Definite histological diagnosis was established for 35/50 (70 %) valves. PCR showed oralis group streptococci in 21 (54 %), S. aureus in 3 (7.7 %), gallolyticus group streptococci in 2 (5.1 %), Coxiella burnetii in 1 (2.5 %) and Rhizobium sp. in 1 (2.5 %). In-hospital mortality was 9/53 (17 %). Fever (p = 0.06, OR 4.7, CI 0.91-24.38) and embolic complications (p = 0.003, OR 3.3, CI 1.55-6.82) were more frequent in BCPE cases, while new acute regurgitation (p = 0.05, OR 0.3, CI 0.098-0.996) and heart failure (p = 0.02, OR 0.3, CI 0.13-0.79) were less so. BCNE resulted mostly from prior antibiotics and was associated with severe hemodynamic compromise. Valve histopathology and PCR were useful in confirming the diagnosis and pointing to the etiology of BCNE.

Centrifugal pump handbook

CERN Document Server

Pumps, Sulzer

2010-01-01

This long-awaited new edition is the complete reference for engineers and designers working on pump design and development or using centrifugal pumps in the field. This authoritative guide has been developed with access to the technical expertise of the leading centrifugal pump developer, Sulzer Pumps. In addition to providing the most comprehensive centrifugal pump theory and design reference with detailed material on cavitation, erosion, selection of materials, rotor vibration behavior and forces acting on pumps, the handbook also covers key pumping applications topics and operational

Survival and function of phagocytes in blood culture media

DEFF Research Database (Denmark)

Fischer, T K; Prag, J; Kharazmi, A

1999-01-01

The survival and function of human phagocytes in sterile aerobic and anaerobic blood culture media were investigated using neutrophil morphology, white blood cell count in a haemoanalyser, flow cytometry, oxidative burst response, and bactericidal effect in Colorbact and Septi-Chek blood culture...... media and Bact/Alert. When comparing agitation to stationary incubation no difference in phagocytic activity was found. The methods showed the same trends demonstrating that the phagocytes' viability and activity were prolonged by oxygen and shortened by anaerobic conditions and sodium polyethanol...... sulfonate (SPS). Best preserved activity and viability were found in the aerobic media containing less than 0.5 g/l SPS, in which significant phagocyte oxidative burst and bactericidal activity were found up to 4 days after inoculation. Considering that the majority of bacteremias are due to aerobic...

Development of an improved rapid BACpro® protocol and a method for direct identification from blood-culture-positive bottles using matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

Science.gov (United States)

Yonezawa, Takatoshi; Watari, Tomohisa; Ashizawa, Kazuho; Hanada, Daisuke; Yanagiya, Takako; Watanabe, Naoki; Terada, Takashi; Tomoda, Yutaka; Fujii, Satoshi

2018-05-01

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been incorporated into pathogenic bacterial identification methods and has improved their rapidity. Various methods have been reported to directly identify bacteria with MALDI-TOF MS by pretreating culture medium in blood culture bottles. Rapid BACpro® (Nittobo Medical Co., Ltd.) is a pretreatment kit for effective collection of bacteria with cationic copolymers. However, the Rapid BACpro® pretreatment kit is adapted only for MALDI Biotyper (Bruker Daltonics K.K.), and there has been a desire to expand its use to VITEK MS (VMS; bioMerieux SA). We improved the protocol and made it possible to analyze with VMS. The culture medium bacteria collection method was changed to a method with centrifugation after hemolysis using saponin; the cationic copolymer concentration was changed to 30% of the original concentration; the sequence with which reagents were added was changed; and a change was made to an ethanol/formic acid extraction method. The improved protocol enhanced the identification performance. When VMS was used, the identification rate was 100% with control samples. With clinical samples, the identification agreement rate with the cell smear method was 96.3%. The improved protocol is effective in blood culture rapid identification, being both simpler and having an improved identification performance compared with the original. Copyright © 2018 Elsevier B.V. All rights reserved.

Dysrhythmias in Laypersons During Centrifuge-Simulated Suborbital Spaceflight.

Science.gov (United States)

Suresh, Rahul; Blue, Rebecca S; Mathers, Charles H; Castleberry, Tarah L; Vanderploeg, James M

2017-11-01

There are limited data on cardiac dysrhythmias in laypersons during hypergravity exposure. We report layperson electrocardiograph (ECG) findings and tolerance of dysrhythmias during centrifuge-simulated suborbital spaceflight. Volunteers participated in varied-length centrifuge training programs of 2-7 centrifuge runs over 0.5-2 d, culminating in two simulated suborbital spaceflights of combined +Gz and +Gx (peak +4.0 Gz, +6.0 Gx, duration 5 s). Monitors recorded pre- and post-run mean arterial blood pressure (MAP), 6-s average heart rate (HR) collected at prespecified points during exposures, documented dysrhythmias observed on continuous 3-lead ECG, self-reported symptoms, and objective signs of intolerance on real-time video monitoring. Participating in the study were 148 subjects (43 women). Documented dysrhythmias included sinus pause (N = 5), couplet premature ventricular contractions (N = 4), bigeminy (N = 3), accelerated idioventricular rhythm (N = 1), and relative bradycardia (RB, defined as a transient HR drop of >20 bpm; N = 63). None were associated with subjective symptoms or objective signs of acceleration intolerance. Episodes of RB occurred only during +Gx exposures. Subjects had a higher post-run vs. pre-run MAP after all exposures, but demonstrated no difference in pre- and post-run HR. RB was more common in men, younger individuals, and subjects experiencing more centrifuge runs. Dysrhythmias in laypersons undergoing simulated suborbital spaceflight were well tolerated, though RB was frequently noted during short-duration +Gx exposure. No subjects demonstrated associated symptoms or objective hemodynamic sequelae from these events. Even so, heightened caution remains warranted when monitoring dysrhythmias in laypersons with significant cardiopulmonary disease or taking medications that modulate cardiac conduction.Suresh R, Blue RS, Mathers CH, Castleberry TL, Vanderploeg JM. Dysrhythmias in laypersons during centrifuge-stimulated suborbital

Prediction and optimization of the recovery rate in centrifugal separation of platelet-rich plasma (PRP)

Science.gov (United States)

Piao, Linfeng; Park, Hyungmin; Jo, Chris

2016-11-01

We present a theoretical model of the recovery rate of platelet and white blood cell in the process of centrifugal separation of platelet-rich plasma (PRP). For the practically used conditions in the field, the separation process is modeled as a one-dimensional particle sedimentation; a quasi-linear partial differential equation is derived based on the kinematic-wave theory. This is solved to determine the interface positions between supernatant-suspension and suspension-sediment, used to estimate the recovery rate of the plasma. While correcting the Brown's hypothesis (1989) claiming that the platelet recovery is linearly proportional to that of plasma, we propose a new correlation model for prediction of the platelet recovery, which is a function of the volume of whole blood, centrifugal acceleration and time. For a range of practical parameters, such as hematocrit, volume of whole blood and centrifugation (time and acceleration), the predicted recovery rate shows a good agreement with available clinical data. We propose that this model is further used to optimize the preparation method of PRP that satisfies the customized case. Supported by a Grant (MPSS-CG-2016-02) through the Disaster and Safety Management Institute funded by Ministry of Public Safety and Security of Korean government.

Enrichment: centrifuge process

International Nuclear Information System (INIS)

Soubbaramayer.

1989-01-01

This short course is divided into three sections devoted respectively to the physics of the process, some practical problems raised by the design of a centrifuge and the present situation of centrifugation in the World. 31 figs., 18 refs

An improved in vitro blood-brain barrier model: rat brain endothelial cells co-cultured with astrocytes.

Science.gov (United States)

Abbott, N Joan; Dolman, Diana E M; Drndarski, Svetlana; Fredriksson, Sarah M

2012-01-01

In vitro blood-brain barrier (BBB) models using primary cultured brain endothelial cells are important for establishing cellular and molecular mechanisms of BBB function. Co-culturing with BBB-associated cells especially astrocytes to mimic more closely the in vivo condition leads to upregulation of the BBB phenotype in the brain endothelial cells. Rat brain endothelial cells (RBECs) are a valuable tool allowing ready comparison with in vivo studies in rodents; however, it has been difficult to obtain pure brain endothelial cells, and few models achieve a transendothelial electrical resistance (TEER, measure of tight junction efficacy) of >200 Ω cm(2), i.e. the models are still relatively leaky. Here, we describe methods for preparing high purity RBECs and neonatal rat astrocytes, and a co-culture method that generates a robust, stable BBB model that can achieve TEER >600 Ω cm(2). The method is based on >20 years experience with RBEC culture, together with recent improvements to kill contaminating cells and encourage BBB differentiation.Astrocytes are isolated by mechanical dissection and cell straining and are frozen for later co-culture. RBECs are isolated from 3-month-old rat cortices. The brains are cleaned of meninges and white matter and enzymatically and mechanically dissociated. Thereafter, the tissue homogenate is centrifuged in bovine serum albumin to separate vessel fragments from other cells that stick to the myelin plug. The vessel fragments undergo a second enzyme digestion to separate pericytes from vessels and break down vessels into shorter segments, after which a Percoll gradient is used to separate capillaries from venules, arterioles, and single cells. To kill remaining contaminating cells such as pericytes, the capillary fragments are plated in puromycin-containing medium and RBECs grown to 50-60% confluence. They are then passaged onto filters for co-culture with astrocytes grown in the bottom of the wells. The whole procedure takes ∼2

Clot formation and lysis in platelet rich plasma of healthy donors and patients with resistant hypertension

Directory of Open Access Journals (Sweden)

I. I. Patalakh

2018-04-01

Full Text Available Hemostatic balance in blood is affected by numerous factors, including coagulation and fibrinolytic proteins, the wide spectrum of their inhibitors, and blood cells. Since platelets can participate in contradictory processes, they significantly complicate the whole picture. Therefore, nowadays the development of global assays of hemostasis, which can reflect the physiological process of hemostasis and can be used for point-of-care diagnosis of thrombosis, is crucial. This paper outlines a new approach we used to analyze the capabilities of clot waveform analysis tools to distinguish the response of platelet-rich plasma from healthy donors and patients with arterial hypertension caused by stimulation of coagulation and lysis (with exogenous thrombin and recombinant tissue-type plasminogen activator, respectively. In donor plasma, when the clot degradation was accompanied by 40 IU/ml of recombinant tissue-type plasminogen activator, platelets potentiated fibrinolysis more than coagulation, which ultimately shifts the overall balance to a profibrinolytic state. At the same time, for patients with hypertension, platelets, embedded in clot obtained from platelet-rich plasma, showed a weaker ability to stimulate fibrinolysis. The obtained data gives the evidence that platelets can act not only as procoagulants but also as profibrinolytics. By simultaneously amplifying coagulation and fibrinolysis, making their rates comparable, platelets would control plasma procoagulant activity, thereby regulating local hemostatic balance, the size and lifetime of the clot. Moreover, clot waveform analysis may be used to distinguish the effects of platelet-rich plasma on clotting or lysis of fibrin clots in healthy donors and patients with essential hypertension.

Detection of circulating microparticles by flow cytometry: influence of centrifugation, filtration of buffer, and freezing.

Science.gov (United States)

Dey-Hazra, Emily; Hertel, Barbara; Kirsch, Torsten; Woywodt, Alexander; Lovric, Svjetlana; Haller, Hermann; Haubitz, Marion; Erdbruegger, Uta

2010-12-06

The clinical importance of microparticles resulting from vesiculation of platelets and other blood cells is increasingly recognized, although no standardized method exists for their measurement. Only a few studies have examined the analytical and preanalytical steps and variables affecting microparticle detection. We focused our analysis on microparticle detection by flow cytometry. The goal of our study was to analyze the effects of different centrifugation protocols looking at different durations of high and low centrifugation speeds. We also analyzed the effect of filtration of buffer and long-term freezing on microparticle quantification, as well as the role of Annexin V in the detection of microparticles. Absolute and platelet-derived microparticles were 10- to 15-fold higher using initial lower centrifugation speeds at 1500 × g compared with protocols using centrifugation speeds at 5000 × g (P centrifugation speeds. Filtration of buffer with a 0.2 μm filter reduced a significant amount of background noise. Storing samples for microparticle detection at -80°C decreased microparticle levels at days 28, 42, and 56 (P centrifugation speeds should be used to minimize contamination by smaller size platelets.

Paperfuge: An ultralow-cost, hand-powered paper-centrifuge inspired by the mechanics of a whirligig toy

Science.gov (United States)

Bhamla, M. Saad; Benson, Brandon; Chai, Chew; Katsikis, Georgios; Johri, Aanchal; Prakash, Manu

From a global-health context, commercial centrifuges are expensive, bulky and electricity-powered, and thus constitute a critical bottleneck in the development of decentralized, battery-free-point-of-care (POC) diagnostic devices. Here, we report an ultralow-cost (20 cents), lightweight (2 g), human-powered paper centrifuge (which we name 'paperfuge') designed on the basis of a theoretical model inspired by the fundamental mechanics of an ancient whirligig (or buzzer toy; 3300 B.C.E). The paperfuge achieves speeds of 125,000 rpm (and equivalent centrifugal forces of 30,000 g), with theoretical limits predicting one million rpm. We demonstrate that the paperfuge can separate pure plasma from whole blood in less than 1.5 minutes, and isolate malaria parasites in 15 minutes. We also show that paperfuge-like centrifugal microfluidic devices can be made of polymethylsiloxane, plastic and 3D-printed polymeric materials. Ultracheap, power-free centrifuges should open up opportunities for POC diagnostics in resource-poor settings.

Induction and identification of rabbit peripheral blood derived dendritic cells

Science.gov (United States)

Zhou, Jing; Yang, FuYuan; Chen, WenLi

2012-03-01

Purpose: To study a method of the induction of dendritic cells (DCs) from rabbit peripheral blood. Methods: Peripheral blood cells were removed from rabbit, filtered through nylon mesh. Peripheral blood mononuclear cells (PBMC) were isolated from the blood cells by Ficoll-Hypaque centrifugation (density of 1.077g/cm3).To obtain DCs, PBMC were cultured in RPMI1640 medium containing 10% fetal calf serum, 50U/mL penicillin and streptomycin, referred to subsequently as complete medium, at 37°C in 5% CO2 atmosphere for 4 hours. Nonadherent cells were aspirated, adherent cells were continued incubated in complete medium, supplemented with granulocyte/macrophage colony-stimulating factor (GM-CSF, 50ng/ml),and interleukin 4 (IL-4, 50ng/ml) for 9 days. Fluorescein labeled antibodies(anti-CD14, anti-HLA-DR, anti-CD86) were used to sign cells cultured for 3,6,9 days respectively, Then flow cytometry was performed. Results: Ratio of anti-HLA-DR and anti-CD86 labeled cells increased with induction time extension, in contrast with anti-CD14. Conclusion: Dendritic cells can be effectively induced by the method of this experiment, cell maturation status increased with induction time extension.

Optimized methods to measure acetoacetate, 3-hydroxybutyrate, glycerol, alanine, pyruvate, lactate and glucose in human blood using a centrifugal analyser with a fluorimetric attachment.

Science.gov (United States)

Stappenbeck, R; Hodson, A W; Skillen, A W; Agius, L; Alberti, K G

1990-01-01

Optimized methods are described for the analysis of glucose, lactate, pyruvate, alanine, glycerol, D-3-hydroxybutyrate and acetoacetate in perchloric acid extracts of human blood using the Cobas Bio centrifugal analyser. Glucose and lactate are measured using the photometric mode and other metabolites using the fluorimetric mode. The intra-assay coefficients of variation ranged from 0.7 to 4.1%, except with very low levels of pyruvate and acetoacetate where the coefficients of variation were 7.1 and 12% respectively. All seven metabolites can be measured in a perchloric acid extract of 20 mul of blood. The methods have been optimized with regard to variation in the perchloric acid content of the samples. These variations arise from the method of sample preparation used to minimize changes occurring in metabolite concentration after venepuncture.

All in the Blood: A Review of Aboriginal Australians' Cultural Beliefs About Blood and Implications for Biospecimen Research.

Science.gov (United States)

Kowal, Emma; Greenwood, Ashley; McWhirter, Rebekah E

2015-10-01

Public participation in medical research and biobanking is considered key to advances in scientific discovery and translation to improved health care. Cultural concerns relating to blood have been found to affect the participation of indigenous peoples and minorities in research, but such concerns are rarely specified in the literature. This article presents a review of the role of blood in Australian Aboriginal cultures. We discuss the range of meanings and uses of blood in traditional culture, including their use in ceremonies, healing, and sorcery. We draw on more recent literature on Aboriginal Australians and biomedicine to consider how traditional beliefs may be changing over time. These findings provide an empirical basis for researchers and bioethicists to develop culturally grounded strategies to boost the participation of Aboriginal Australians in biomedical research. They also serve as a model for integrating anthropological literature with bioethical concerns that could be applied to other indigenous and minority groups. © The Author(s) 2015.

Effectiveness of a Novel Specimen Collection System in Reducing Blood Culture Contamination Rates.

Science.gov (United States)

Bell, Mary; Bogar, Catherine; Plante, Jessica; Rasmussen, Kristen; Winters, Sharon

2018-04-20

False-positive blood-culture results due to skin contamination of samples remain a persistent problem for health care providers. Our health system recognized that our rates of contamination across the 4 emergency department campuses were above the national average. A unique specimen collection system was implemented throughout the 4 emergency departments and became the mandatory way to collect adult blood cultures. The microbiology laboratory reported contamination rates weekly to manage potential problems; 7 months of data are presented here. There was an 82.8% reduction in false positives with the unique specimen collection system compared with the standard method (chi-squared test with Yates correction, 2-tailed, P = 0.0001). Based on the historical 3.52% rate of blood-culture contamination for our health facilities, 2.92 false positives were prevented for every 100 blood cultures drawn, resulting from adoption of the unique specimen collection system as the standard of care. This unique collection system can reduce the risk of blood culture contamination significantly and is designed to augment, rather than replace, the standard phlebotomy protocol already in use in most health care settings. Copyright © 2018 Elsevier Inc. All rights reserved.

Separation parameters of gas centrifuges

International Nuclear Information System (INIS)

May, W.G.

1977-01-01

Early work on development of the gas centrifuge for separation of uranium isotopes has recently been reviewed. Several configurations were investigated. The preferred configuration eventually turned out to be a countercurrent centrifuge. In this form, an internal circulation is set up, and as a consequence, light isotope concentrates at one end of the centrifuge, heavy isotope at the other. In many ways the effect resembles the separation obtained in packed columns in the chemical and petroleum industries. It is the purpose of this paper to develop this analogy between countercurrent gas centrifuges and packed towers and to illustrate its usefulness in understanding the separation process in the centrifuge. 8 figures

Effect of centrifugation and washing on adipose graft viability: a new method to improve graft efficiency.

Science.gov (United States)

Hoareau, Laurence; Bencharif, Karima; Girard, Anne-Claire; Gence, Lydie; Delarue, Pierre; Hulard, Olivier; Festy, Franck; Roche, Regis

2013-05-01

Adipose tissue grafting is a promising method in the field of surgical filling. We studied the effect of centrifugation on fat grafts, and we propose an optimised protocol for the improvement of adipose tissue viability. Adipose tissue was subjected to different centrifugations, and the volumes of interstitial liquid and oil released were measured to choose the optimal condition. Tissue from this condition was then compared to tissue obtained from two traditional techniques: strong centrifugation (commonly 3 min at 3000 rpm/900 g), and decantation, by injecting into immunodeficient mice. The cytokine interleukin-6 (IL-6) and chemokine monocyte chemotactic protein-1 (MCP-1) were assayed 24 h post-injection, and after 1 month of grafting the state of the lipografts was evaluated through macroscopic and histological analysis, with oil gap area measurement. Strong centrifugation (900 g, 1800 g) is deleterious for adipose tissue because it leads to until threefold more adipocyte death compared to low centrifugation (100 g, 400 g). In addition, mice injected with strong centrifuged and non-centrifuged adipose tissue have higher rates of blood IL-6 and MCP-1, compared to those grafted with soft centrifuged fat. Moreover, extensive lipid vacuoles were detectable on histological sections of the non-centrifuged lipografts, whereas lipografts from soft centrifugation contain a higher amount of connective tissue containing collagen fibres. It is necessary to wash and centrifuge adipose tissue before reinjection in order to remove infiltration liquid and associated toxic molecules, which in the long term are deleterious for the graft. However, strong centrifugation is not recommended since it leads very quickly to greater adipocyte death. Thus, soft centrifugation (400 g/1 min), preceded by washings, seems to be the most appropriate protocol for the reinjection of adipose tissue. Copyright © 2013 British Association of Plastic, Reconstructive and Aesthetic Surgeons. Published

Blood culture cross contamination associated with a radiometric analyzer

International Nuclear Information System (INIS)

Griffin, M.R.; Miller, A.D.; Davis, A.C.

1982-01-01

During a 9-day period in August 1980 in a New Jersey hospital, three pairs of consecutively numbered blood cultures from different patients were identified as positive for the same organism, for each pair, both cultures were positive in the same atmosphere, both organisms had the same sensitivities, and the second of each pair grew at least 2 days after the first and was the only positive blood culture obtained from the patient. When the hospital laboratory discontinued use of its radiometric culture analyzer for 15 days, no more consecutive pairs of positive cultures occurred. Subsequent use of the machine for 9 days with a new power unit but the original circuit boards resulted in one more similar consecutive pair (Staphylococcus epidermidis). After replacement of the entire power unit, there were no further such pairs. Examination of the machine by the manufacturer revealed a defective circuit board which resulted in inadequate needle sterilization. Laboratories which utilize radiometric analyzers should be aware of the potential for cross contamination. Recognition of such events requires alert microbiologists and infection control practitioners and a record system in the bacteriology laboratory designed to identify such clusters

Optofluidic Sensor for Inline Hemolysis Detection on Whole Blood

DEFF Research Database (Denmark)

Zhou, Chen; Keshavarz Hedayati, Mehdi; Zhu, Xiaolong

2018-01-01

Hemolysis is the rupture of red blood cells and constitutes the most common reason for unsuitable blood samples in the clinic. To detect hemolysis, one has to separate the hemoglobin in blood plasma from that in red blood cells. However, current methods entail centrifugation for cell......-time inline detection on whole blood without extra sample preparation like centrifugation. Long-term testing with inline integration in a modified, commercial blood gas analyzer shows high reliability and repeatability of the measurements even with the presence of interference from bilirubin. We envision...... that the present work has large potential in improving diagnosis quality by enabling PoC hemolysis detection in blood gas analyzers and can also lend unique sensing capabilities to other applications dealing with complex turbid media....

Effect of sample type, centrifugation and storage conditions on vitamin D concentration.

Science.gov (United States)

Colak, Ayfer; Toprak, Burak; Dogan, Nese; Ustuner, Fusun

2013-01-01

Studies about vitamin D [25(OH)D] stability in plasma are limited and preanalytical variables such as tube type may affect results. We aimed to evaluate effect of storage conditions, sample type and some preanalytical variables on vitamin D concentration. Blood samples from 15 healthy subjects were centrifuged at different temperatures and stored under different conditions. Serum and plasma 25(OH)D difference, effect of centrifugation temperature and common storage conditions were investigated. There was no difference between serum and plasma vitamin D concentration. Centrifugation temperature had no impact on vitamin D concentration. 25(OH)D is stable under common storage conditions: 4 hours at room temperature, 24 hours at 2-8 degrees C, 7 days at -20 degrees C, 3 months at -80 degrees C. Vitamin D does not require any special storage conditions and refrigeration. Both serum and plasma can be used for measurement.

Improved Diagnosis of Prosthetic Joint Infection by Culturing Periprosthetic Tissue Specimens in Blood Culture Bottles.

Science.gov (United States)

Peel, Trisha N; Dylla, Brenda L; Hughes, John G; Lynch, David T; Greenwood-Quaintance, Kerryl E; Cheng, Allen C; Mandrekar, Jayawant N; Patel, Robin

2016-01-05

Despite known low sensitivity, culture of periprosthetic tissue specimens on agars and in broths is routine. Culture of periprosthetic tissue samples in blood culture bottles (BCBs) is potentially more convenient, but it has been evaluated in a limited way and has not been widely adopted. The aim of this study was to compare the sensitivity and specificity of inoculation of periprosthetic tissue specimens into blood culture bottles with standard agar and thioglycolate broth culture, applying Bayesian latent class modeling (LCM) in addition to applying the Infectious Diseases Society of America (IDSA) criteria for prosthetic joint infection. This prospective cohort study was conducted over a 9-month period (August 2013 to April 2014) at the Mayo Clinic, Rochester, MN, and included all consecutive patients undergoing revision arthroplasty. Overall, 369 subjects were studied; 117 (32%) met IDSA criteria for prosthetic joint infection, and 82% had late chronic infection. Applying LCM, inoculation of tissues into BCBs was associated with a 47% improvement in sensitivity compared to the sensitivity of conventional agar and broth cultures (92.1 versus 62.6%, respectively); this magnitude of change was similar when IDSA criteria were applied (60.7 versus 44.4%, respectively; P = 0.003). The time to microorganism detection was shorter with BCBs than with standard media (P Prosthetic joint infections are a devastating complication of arthroplasty surgery. Despite this, current microbiological techniques to detect and diagnose infections are imperfect. This study examined a new approach to diagnosing infections, through the inoculation of tissue samples from around the prosthetic joint into blood culture bottles. This study demonstrated that, compared to current laboratory practices, this new technique increased the detection of infection. These findings are important for patient care to allow timely and accurate diagnosis of infection. Copyright © 2016 Peel et al.

Complement lysis activity in autologous plasma is associated with lower viral loads during the acute phase of HIV-1 infection.

Directory of Open Access Journals (Sweden)

Michael Huber

2006-11-01

Full Text Available BACKGROUND: To explore the possibility that antibody-mediated complement lysis contributes to viremia control in HIV-1 infection, we measured the activity of patient plasma in mediating complement lysis of autologous primary virus. METHODS AND FINDINGS: Sera from two groups of patients-25 with acute HIV-1 infection and 31 with chronic infection-were used in this study. We developed a novel real-time PCR-based assay strategy that allows reliable and sensitive quantification of virus lysis by complement. Plasma derived at the time of virus isolation induced complement lysis of the autologous virus isolate in the majority of patients. Overall lysis activity against the autologous virus and the heterologous primary virus strain JR-FL was higher at chronic disease stages than during the acute phase. Most strikingly, we found that plasma virus load levels during the acute but not the chronic infection phase correlated inversely with the autologous complement lysis activity. Antibody reactivity to the envelope (Env proteins gp120 and gp41 were positively correlated with the lysis activity against JR-FL, indicating that anti-Env responses mediated complement lysis. Neutralization and complement lysis activity against autologous viruses were not associated, suggesting that complement lysis is predominantly caused by non-neutralizing antibodies. CONCLUSIONS: Collectively our data provide evidence that antibody-mediated complement virion lysis develops rapidly and is effective early in the course of infection; thus it should be considered a parameter that, in concert with other immune functions, steers viremia control in vivo.

Quantifying enzymatic lysis: estimating the combined effects of chemistry, physiology and physics

International Nuclear Information System (INIS)

Mitchell, Gabriel J; Weitz, Joshua S; Nelson, Daniel C

2010-01-01

The number of microbial pathogens resistant to antibiotics continues to increase even as the rate of discovery and approval of new antibiotic therapeutics steadily decreases. Many researchers have begun to investigate the therapeutic potential of naturally occurring lytic enzymes as an alternative to traditional antibiotics. However, direct characterization of lytic enzymes using techniques based on synthetic substrates is often difficult because lytic enzymes bind to the complex superstructure of intact cell walls. Here we present a new standard for the analysis of lytic enzymes based on turbidity assays which allow us to probe the dynamics of lysis without preparing a synthetic substrate. The challenge in the analysis of these assays is to infer the microscopic details of lysis from macroscopic turbidity data. We propose a model of enzymatic lysis that integrates the chemistry responsible for bond cleavage with the physical mechanisms leading to cell wall failure. We then present a solution to an inverse problem in which we estimate reaction rate constants and the heterogeneous susceptibility to lysis among target cells. We validate our model given simulated and experimental turbidity assays. The ability to estimate reaction rate constants for lytic enzymes will facilitate their biochemical characterization and development as antimicrobial therapeutics

Quantifying enzymatic lysis: estimating the combined effects of chemistry, physiology and physics.

Science.gov (United States)

Mitchell, Gabriel J; Nelson, Daniel C; Weitz, Joshua S

2010-10-04

The number of microbial pathogens resistant to antibiotics continues to increase even as the rate of discovery and approval of new antibiotic therapeutics steadily decreases. Many researchers have begun to investigate the therapeutic potential of naturally occurring lytic enzymes as an alternative to traditional antibiotics. However, direct characterization of lytic enzymes using techniques based on synthetic substrates is often difficult because lytic enzymes bind to the complex superstructure of intact cell walls. Here we present a new standard for the analysis of lytic enzymes based on turbidity assays which allow us to probe the dynamics of lysis without preparing a synthetic substrate. The challenge in the analysis of these assays is to infer the microscopic details of lysis from macroscopic turbidity data. We propose a model of enzymatic lysis that integrates the chemistry responsible for bond cleavage with the physical mechanisms leading to cell wall failure. We then present a solution to an inverse problem in which we estimate reaction rate constants and the heterogeneous susceptibility to lysis among target cells. We validate our model given simulated and experimental turbidity assays. The ability to estimate reaction rate constants for lytic enzymes will facilitate their biochemical characterization and development as antimicrobial therapeutics.

Electrochemical lysis at the stage of endoresection for large posterior intraocular tumors

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Yu. A. Belyy

2012-01-01

Full Text Available Purpose: to design the new combined technique of endoresection with intraoperative intraocular electrochemical lysis at the tumor destruction stage for large posterior intraocular tumors.Methods: 3 patients (3 eyes with large choroidal melanomas t3N0M0 (tumor thickness — 8-10 mm, base diameter — 13-15 mm, juxtapapillary localization. Mean age was 55.4 years old. Endoresection with intraoperational intraocular electrochemical lysis of the tumor was performed. Electrochemical lysis was performed with use of the technical unit ECU 300 (Soering, Germany and the original method of combined intratumoral positioning of two platinum electrodes: anode and cathode.Results: the tumor was removal completely in all 3 cases. the anatomical retinal reattachment was reached in all patients. Sclera was safe in all 3 cases. Visual acuity was not changed (NLP. At the place of the removal tumor a surgical choroidal coloboma without pigmentation all over scleral bed and periphery was shown in all cases in distant postoperative period (from 1.5 to 3 years. No local recurrences or metastasis were revealed in all patients.Conclusion: Further investigations in clinical group are necessarily to determinate the real possibilities of the combined method and the indications for endoresection with intraoperative intraocular electrochemical lysis for large intraocular tumors. 

Recent and innovative methods for detection of bacteremia and fungemia

International Nuclear Information System (INIS)

Reller, L.B.

1983-01-01

Advances continue to be made in methods for more reliable or more rapid means of detecting bacteremia and fungemia. The importance of blood sample volume and broth dilution has been established in controlled studies. New technology includes the use of resins that remove antimicrobials from blood samples, detection of radioactivity from organisms given radiolabeled substrate, use of dyes that stain microbial DNA and RNA, use of slides coated with growth media, and lysis-centrifugation for trapping microorganisms. Technology now being considered includes counterimmunoelectrophoresis, head-space gas chromatography, electrical impedance, microcalorimetry, and the use of lasers to detect pH changes and turbidity

Impact of sample processing on the measurement of circulating microparticles: storage and centrifugation parameters.

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Vila-Liante, Virtudes; Sánchez-López, Verónica; Martínez-Sales, Vicenta; Ramón-Nuñez, Luis A; Arellano-Orden, Elena; Cano-Ruiz, Alejandra; Rodríguez-Martorell, Francisco J; Gao, Lin; Otero-Candelera, Remedios

2016-11-01

Microparticles (MPs) have been shown to be markers of cellular activation and interactions. Pre-analytical conditions such as the centrifugation protocol and sample storage conditions represent an important source of variability in determining MPs values. The objectives of this study were to evaluate the influence of sample storage conditions and centrifugation speed and temperature on the determination of MPs in plasma. Citrate-anticoagulated blood samples obtained from 21 healthy subjects were centrifuged under four different protocols involving different speeds (2500 g or 1500 g) and temperatures (4 °C or 20 °C) to isolate platelet-poor plasma (PPP). The number of MPs in fresh and frozen-thawed PPP were analyzed by flow cytometry, and MPs-mediated procoagulant activity was determined by a thrombin generation test and phospholipid-dependent procoagulant tests. The number of MPs and their procoagulant activity were affected by freeze-thaw cycling and centrifugation speed but not by centrifugation temperature. Sample freezing increased MPs number (six-fold) and thrombin generation (four-fold), and decreased clotting time (two-fold). Low centrifugation speed caused an increase in MPs number and a parallel increase in MP-mediated procoagulant activity. Sample storage conditions and centrifugation speed are important processing conditions affecting MPs number and activity. Before any study, the protocol for MPs isolation should be optimized to ensure a reliable characterization of MPs, which could provide important information for diagnostic purposes and for understanding the pathogenesis of diseases.

Organic and Inorganic Nitrogen Impact Chlorella variabilis Productivity and Host Quality for Viral Production and Cell Lysis.

Science.gov (United States)

Cheng, Yu-Shen; Labavitch, John; VanderGheynst, Jean S

2015-05-01

Microalgae have been proposed as a potential feedstock for biofuel production; however, cell disruption is usually required for collection and utilization of cytoplasmic polysaccharides and lipids. Virus infection might be one approach to disrupt the cell wall. The concentration of yeast extract and presence of KNO3 in algae cultivation media were investigated to observe their effects on Chlorella variabilis NC64A physiology and composition and the subsequent effect on production of Chlorella virus and disruption of infected cells. Cytoplasmic starch accumulation increased from 5% to approximately 35% of the total dry weight when yeast extract decreased from 1 to 0.25 g L(-1). When cells were cultured with the lowest nitrogen levels, the total polysaccharide accounted for more than 50% of the cell wall, which was 1.7 times higher than the content in cells cultured with the highest nitrogen levels. The C/N ratio of the algal biomass decreased by a factor of approximately 2 when yeast extract increased from 0.25 to 1 g L(-1). After virus infection, cells with a low C/N ratio produced a 7.6 times higher burst size than cells with a high C/N ratio, suggesting that the nitrogen content in C. variabilis has a large influence on viral production and cell lysis. The results have implications on management of nitrogen for both the synthesis of products from algae and product recovery via viral lysis.

The impact of the centrifuge characteristics and centrifugation protocols on the cells, growth factors, and fibrin architecture of a leukocyte- and platelet-rich fibrin (L-PRF) clot and membrane.

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Dohan Ehrenfest, David M; Pinto, Nelson R; Pereda, Andrea; Jiménez, Paula; Corso, Marco Del; Kang, Byung-Soo; Nally, Mauricio; Lanata, Nicole; Wang, Hom-Lay; Quirynen, Marc

2018-03-01

L-PRF (leukocyte- and platelet-rich fibrin) is one of the four families of platelet concentrates for surgical use and is widely used in oral and maxillofacial regenerative therapies. The first objective of this article was to evaluate the mechanical vibrations appearing during centrifugation in four models of commercially available table-top centrifuges used to produce L-PRF and the impact of the centrifuge characteristics on the cell and fibrin architecture of a L-PRF clot and membrane. The second objective of this article was to evaluate how changing some parameters of the L-PRF protocol may influence its biological signature, independently from the characteristics of the centrifuge. In the first part, four different commercially available centrifuges were used to produce L-PRF, following the original L-PRF production method (glass-coated plastic tubes, 400 g force, 12 minutes). The tested systems were the original L-PRF centrifuge (Intra-Spin, Intra-Lock, the only CE and FDA cleared system for the preparation of L-PRF) and three other laboratory centrifuges (not CE/FDA cleared for L-PRF): A-PRF 12 (Advanced PRF, Process), LW-UPD8 (LW Scientific) and Salvin 1310 (Salvin Dental). Each centrifuge was opened for inspection, two accelerometers were installed (one radial, one vertical), and data were collected with a spectrum analyzer in two configurations (full-load or half load). All clots and membranes were collected into a sterile surgical box (Xpression kit, Intra-Lock). The exact macroscopic (weights, sizes) and microscopic (photonic and scanning electron microscopy SEM) characteristics of the L-PRF produced with these four different machines were evaluated. In the second part, venous blood was taken in two groups, respectively, Intra-Spin 9 ml glass-coated plastic tubes (Intra-Lock) and A-PRF 10 ml glass tubes (Process). Tubes were immediately centrifuged at 2700 rpm (around 400 g) during 12 minutes to produce L-PRF or at 1500 rpm during 14 minutes to produce A

Microwave assisted centrifuge and related methods

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Meikrantz, David H [Idaho Falls, ID

2010-08-17

Centrifuge samples may be exposed to microwave energy to heat the samples during centrifugation and to promote separation of the different components or constituents of the samples using a centrifuge device configured for generating microwave energy and directing the microwave energy at a sample located in the centrifuge.

A lab-on-CD prototype for high-speed blood separation

International Nuclear Information System (INIS)

Zhang Jinlong; Liu Mei; Yang Jun; Guo Qiuquan

2008-01-01

Blood separation is the first step for subsequent blood tests in clinical diagnosis. Lab-on-a-chip technology provides an automatic, cost-effective and fast solution for a wide variety of blood analyses. The objective of this work is to design a new lab-on-CD microstructure capable of separating blood cells from the whole blood into different reservoirs directly. A CD platform including a microchannel network consisting of a straight main microchannel, a curved microchannel and a branching microchannel has been proposed. The merits of this design are its simple structure, less operating time and high separation efficiency because it utilizes multiple separation mechanisms, for instance, two centrifugal forces and Coriolis force. One centrifugal force is due to the system rotation; the other centrifugal force is due to the curvature of the specifically designed curved channel. In this work, systematical evaluation on the functionality and performance of such a design has been done. Ninety-nine per cent separation efficiency is achieved for diluted blood of 6% hematocrit

A lab-on-CD prototype for high-speed blood separation

Science.gov (United States)

Zhang, Jinlong; Guo, Qiuquan; Liu, Mei; Yang, Jun

2008-12-01

Blood separation is the first step for subsequent blood tests in clinical diagnosis. Lab-on-a-chip technology provides an automatic, cost-effective and fast solution for a wide variety of blood analyses. The objective of this work is to design a new lab-on-CD microstructure capable of separating blood cells from the whole blood into different reservoirs directly. A CD platform including a microchannel network consisting of a straight main microchannel, a curved microchannel and a branching microchannel has been proposed. The merits of this design are its simple structure, less operating time and high separation efficiency because it utilizes multiple separation mechanisms, for instance, two centrifugal forces and Coriolis force. One centrifugal force is due to the system rotation; the other centrifugal force is due to the curvature of the specifically designed curved channel. In this work, systematical evaluation on the functionality and performance of such a design has been done. Ninety-nine per cent separation efficiency is achieved for diluted blood of 6% hematocrit.

76 FR 50767 - In the Matter of USEC Inc., American Centrifuge Lead Cascade Facility, and American Centrifuge...

Science.gov (United States)

2011-08-16

...; License Nos. SNM-7003, SNM-2011] In the Matter of USEC Inc., American Centrifuge Lead Cascade Facility, and American Centrifuge Plant; Order Extending the Date by Which the Direct Transfer of Licenses Is To... American Centrifuge Lead Cascade Facility (Lead Cascade) and American Centrifuge Plant (ACP), respectively...

Comparison of three methods for recovery of Brucella canis DNA from canine blood samples.

Science.gov (United States)

Batinga, Maria Cryskely A; Dos Santos, Jaíne C; Lima, Julia T R; Bigotto, Maria Fernanda D; Muner, Kerstin; Faita, Thalita; Soares, Rodrigo M; da Silva, David A V; Oliveira, Trícia M F S; Ferreira, Helena L; Diniz, Jaqueline A; Keid, Lara B

2017-12-01

Brucella canis, a gram-negative, facultative intracellular and zoonotic bacterium causes canine brucellosis. Direct methods are the most appropriate for the detection of canine brucellosis and bacterial isolation from blood samples has been employed as gold-standard method. However, due to the delay in obtaining results and the biological risk of the bacterial culturing, the polymerase chain reaction (PCR) has been successfully used as an alternative method for the diagnosis of the infection. Sample preparation is a key step for successful PCR and protocols that provide high DNA yield and purity are recommended to ensure high diagnostic sensitivity. The objective of this study was to evaluate the performance of PCR for the diagnosis of B. canis infection in 36 dogs by testing DNA of whole blood obtained through different extraction and purification protocols. Methods 1 and 2 were based on a commercial kit, using protocols recommended for DNA purification of whole blood and tissue samples, respectively. Method 3 was an in-house method based on enzymatic lysis and purification using organic solvents. The results of the PCR on samples obtained through three different DNA extraction protocols were compared to the blood culture. Of the 36 dogs, 13 (36.1%) were positive by blood culturing, while nine (25.0%), 14 (38.8%), and 15 (41.6%) were positive by PCR after DNA extraction using methods 1, 2 and 3, respectively. PCR performed on DNA purified by Method 2 was as efficient as blood culturing and PCR performed on DNA purified with in-house method, but had the advantage of being less laborious and, therefore, a suitable alternative for the direct B. canis detection in dogs. Copyright © 2017. Published by Elsevier B.V.

Rare incidence of tumor lysis syndrome in metastatic prostate cancer following treatment with docetaxel.

Science.gov (United States)

Bhardwaj, Sharonlin; Varma, Seema

2018-03-01

Tumor lysis syndrome is a serious and sometimes lethal complication of cancer treatment that is comprised of a set of metabolic disturbances along with clinical manifestations. Initiating chemotherapy in bulky, rapidly proliferating tumors causes rapid cell turnover that in turn releases metabolites into circulation that give rise to metabolic derangements that can be dangerous. This syndrome is usually seen in high-grade hematological malignancies. Less commonly, tumor lysis syndrome can present in solid tumors and even rarely in genitourinary tumors. In this report, the authors describe a specific case of tumor lysis syndrome in a patient with metastatic prostate cancer following treatment with docetaxel.

Preanalytical Biases in the Measurement of Human Blood Sphingolipids

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Robert Brunkhorst

2018-05-01

Full Text Available Dysregulation of blood sphingolipids is an emerging topic in clinical science. The objective of this study was to determine preanalytical biases that typically occur in clinical and translational studies and that influence measured blood sphingolipid levels. Therefore, we collected blood samples from four healthy male volunteers to investigate the effect of storage conditions (time, temperature, long-term storage, freeze–thaw cycles, blood drawing (venous or arterial sampling, prolonged venous compression, and sample preparation (centrifugation, freezing on sphingolipid levels measured by LC-MS/MS. Our data show that sphingosine 1-phosphate (S1P and sphinganine 1-phosphate (SA1P were upregulated in whole blood samples in a time- and temperature-dependent manner. Increased centrifugation at higher speeds led to lower amounts of S1P and SA1P. All other preanalytical biases did not significantly alter the amounts of S1P and SA1P. Further, in almost all settings, we did not detect differences in (dihydroceramide levels. In summary, besides time-, temperature-, and centrifugation-dependent changes in S1P and SA1P levels, sphingolipids in blood remained stable under practically relevant preanalytical conditions.

Evaluation of the antimicrobial removal device when used with the BACTEC blood culture system

International Nuclear Information System (INIS)

Strand, C.L.

1982-01-01

A study to determine the value of the Antimicrobial Removal Device (ARD) used in conjunction with radiometric detection of bacteremia using three media was conducted. During a 12-month period, 622 duplicate ARD/BACTEC blood-culture sets were collected. There were 88 positive cultures that yielded 68 pathogenic isolates and 28 probable contaminant isolates. When all patients were considered, 31 pathogenic isolates were detected by both systems, 25 pathogenic isolates were detected faster or only by the BACTEC system, and 12 pathogenic isolates were detected faster or only when the ARD was employed. This difference is statistically significant (P less than 0.05). Thus, the standard BACTEC blood-culture system using three different media was superior to the same BACTEC system using ARD-processed blood specimens. When only patients receiving antimicrobial therapy were considered, there were more pathogenic isolates detected from unprocessed blood than from blood processed in the ARD; however, this difference was not statistically significant. In conclusion, there appears to be no advantage to using the ARD system in conjunction with the three-bottle BACTEC blood-culture system

The impact of centrifugation in primary care on pseudohyperkalaemia: a retrospective evaluation.

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Turner, Helen E; Peake, Roy W A; Allison, James J

2013-07-01

Spurious hyperkalaemia is a relatively common occurrence in samples originating from primary care. Failure to identify spurious hyperkalaemia carries a significant risk of patient mismanagement. We have carried out a retrospective evaluation to review the impact of the use of centrifuges in primary care for biochemistry blood samples on the management of hyperkalaemia. Serum potassium concentrations in samples received from primary care were reviewed for six months prior to and after the implementation of on-site centrifugation. Samples exhibiting significant hyperkalaemia (serum potassium >6.0 mmol/L) were further investigated to ascertain the degree of patient follow-up. There was a significant decrease in the number of samples exhibiting marked hyperkalaemia following the implementation (2244 versus 524; P centrifugation of serum samples in primary care improves the sample quality and the integrity of the potassium results reported. We have also demonstrated evidence of an improvement in patient management and quality of care.

Direct identification from Bact/Alert™ blood culture bottles by MALDI-TOF

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Vesselina Kroumova

2011-12-01

Full Text Available Bacterial identification from blood culture using traditional methods needs about 48 hours, since positivization, to be performed. Rapid bacterial identification can result in clinical and economic benefits. To provide rapid pathogen identification for targeted antibiotic treatment, in this study we tested an our previously described homemade method for bacterial identification using MALDI-TOF directly from positive BACTEC blood culture, on positive BacT/ALERT blood culture. A total of 108 bacteria were identified by MALDI-TOF with a positive identification obtained for 98% of Gram negative and 84,3% of Gram positive bacteria.The average of identification score obtained using the protocol described in this study was 2,047 for Gram positive and 2,204 for Gram negative microorganisms. Data here described show that this method is also useful when BacT/ALERT bottles are used and even if these bottles have activated charcoal as inhibitor of antibiotics.

Using CF11 cellulose columns to inexpensively and effectively remove human DNA from Plasmodium falciparum-infected whole blood samples

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Venkatesan Meera

2012-02-01

Full Text Available Abstract Background Genome and transcriptome studies of Plasmodium nucleic acids obtained from parasitized whole blood are greatly improved by depletion of human DNA or enrichment of parasite DNA prior to next-generation sequencing and microarray hybridization. The most effective method currently used is a two-step procedure to deplete leukocytes: centrifugation using density gradient media followed by filtration through expensive, commercially available columns. This method is not easily implemented in field studies that collect hundreds of samples and simultaneously process samples for multiple laboratory analyses. Inexpensive syringes, hand-packed with CF11 cellulose powder, were recently shown to improve ex vivo cultivation of Plasmodium vivax obtained from parasitized whole blood. This study was undertaken to determine whether CF11 columns could be adapted to isolate Plasmodium falciparum DNA from parasitized whole blood and achieve current quantity and purity requirements for Illumina sequencing. Methods The CF11 procedure was compared with the current two-step standard of leukocyte depletion using parasitized red blood cells cultured in vitro and parasitized blood obtained ex vivo from Cambodian patients with malaria. Procedural variations in centrifugation and column size were tested, along with a range of blood volumes and parasite densities. Results CF11 filtration reliably produces 500 nanograms of DNA with less than 50% human DNA contamination, which is comparable to that obtained by the two-step method and falls within the current quality control requirements for Illumina sequencing. In addition, a centrifuge-free version of the CF11 filtration method to isolate P. falciparum DNA at remote and minimally equipped field sites in malaria-endemic areas was validated. Conclusions CF11 filtration is a cost-effective, scalable, one-step approach to remove human DNA from P. falciparum-infected whole blood samples.

Centrifuge design and development

International Nuclear Information System (INIS)

Edwards, T.T.; Holmes, M.D.

1987-01-01

BNFL has been enriching uranium on an industrial scale using the centrifuge process for over a decade. Together with its Urenco partners, a joint development programme has been and is being vigorously pursued to reduce specific costs, increase output and maintain competitiveness throughout the 1990s. The paper summarises the development of the centrifuge from its earliest concepts through to the centrifuges of today which are jointly designed by the Urenco partners. The potential for further development is also examined. (author)

Coagulase-negative Staphylococci in Danish blood cultures: species distribution and antibiotic susceptibility.

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Jarløv, J O; Højbjerg, T; Busch-Sørensen, C; Scheibel, J; Møller, J K; Kolmos, H J; Wandall, D A

1996-03-01

The distribution and antibiotic susceptibility of coagulase-negative staphylococci (CoNS) isolated from blood cultures was examined in samples from hospitals covering most of Denmark. A total of 499 CoNS isolates were detected in 477 blood cultures from 340 patients and speciated as Staphylococcus epidermidis, 285; Staphylococcus hominis, 61; Staphylococcus haemolyticus, 43; Staphylococcus warneri, 12; Staphylococcus cohnii, 7; Staphylococcus saprophyticus, 4; Staphylococcus capitis, 2 and Staphylococcus lugdunensis, 1. Seventy-eight isolates could not be identified to species level and six were Micrococcus spp. In 108 (22.6%) blood culture sets, more than one CoNS strain were found, as detected by species identification, antibiogram and biotyping. Significantly more blood cultures from patients in university hospitals were drawn from central venous catheters. Comparing university and non-university hospitals, the overall antibiotic susceptibility among CoNS was only slightly different, except for methicillin and amikacin. The prevalence of methicillin-resistant strains was 35.1% in the university hospital strains vs. 25.3% in the non-university hospital strains. The overall prevalence of methicillin resistance was 32%. Great geographic variation in both species distribution and antibiotic resistance was observed. The high prevalence of S. epidermidis makes subtyping of this species important.

Combined short-arm centrifuge and aerobic exercise training improves cardiovascular function and physical working capacity in humans.

Science.gov (United States)

Yang, Chang-Bin; Zhang, Shu; Zhang, Yu; Wang, Bing; Yao, Yong-Jie; Wang, Yong-Chun; Wu, Yan-Hong; Liang, Wen-Bin; Sun, Xi-Qing

2010-12-01

Musculoskeletal and cardiovascular deconditioning occurring in long-term spaceflight gives rise to the needs to develop new strategies to counteract these adverse effects. Short-arm centrifuge combined with ergometer has been proposed as a strategy to counteract adverse effects of microgravity. This study sought to investigate whether the combination of short-arm centrifuge and aerobic exercise training have advantages over short-arm centrifuge or aerobic exercise training alone. One week training was conducted by 24 healthy men. They were randomly divided into 3 groups: (1) short-arm centrifuge training, (2) aerobic exercise training, 40 W, and (3) combined short-arm centrifuge and aerobic exercise training. Before and after training, the cardiac pump function represented by stroke volume, cardiac output, left ventricular ejection time, and total peripheral resistance was evaluated. Variability of heart rate and systolic blood pressure were determined by spectral analysis. Physical working capacity was surveyed by near maximal physical working capacity test. The 1-week combined short-arm centrifuge and aerobic exercise training remarkably ameliorated the cardiac pump function and enhanced vasomotor sympathetic nerve modulation and improved physical working capacity by 10.9% (Pcentrifuge nor the aerobic exercise group showed improvements in these functions. These results demonstrate that combined short-arm centrifuge and aerobic exercise training has advantages over short-arm centrifuge or aerobic exercise training alone in influencing several physiologically important cardiovascular functions in humans. The combination of short-arm centrifuge and aerobic exercise offers a promising countermeasure to microgravity.

Effects of X-irradiation on artificial blood vessel wall degradation by invasive tumor cells

International Nuclear Information System (INIS)

Heisel, M.A.; Laug, W.E.; Stowe, S.M.; Jones, P.A.

1984-01-01

Artificial vessel wall cultures, constructed by growing arterial endothelial cells on preformed layers of rat smooth muscle cells, were used to evaluate the effects of X-irradiation on tumor cell-induced tissue degradation. Bovine endothelial cells had radiation sensitivities similar to those of rat smooth muscle cells. Preirradiation of smooth muscle cells, before the addition of human fibrosarcoma (HT 1080) cells, did not increase the rate of degradation and destruction by the invasive cells. However, the degradation rate was decreased if the cultures were irradiated after the addition of HT 1080 cells. The presence of bovine endothelial cells markedly inhibited the destructive abilities of fibrosarcoma cells, but preirradiation of artificial vessel walls substantially decreased their capabilities to resist HT 1080-induced lysis. These findings suggest that the abilities of blood vessels to limit extravasation may be compromised by ionizing radiation

Vacuum chamber-free centrifuge with magnetic bearings.

Science.gov (United States)

Park, Cheol Hoon; Kim, Soohyun; Kim, Kyung-Soo

2013-09-01

Centrifuges are devices that separate particles of different densities and sizes through the application of a centrifugal force. If a centrifuge could be operated under atmospheric conditions, all vacuum-related components such as the vacuum chamber, vacuum pump, diffusion pump, and sealing could be removed from a conventional centrifuge system. The design and manufacturing procedure for centrifuges could then be greatly simplified to facilitate the production of lightweight centrifuge systems of smaller volume. Furthermore, the maintenance costs incurred owing to wear and tear due to conventional ball bearings would be eliminated. In this study, we describe a novel vacuum chamber-free centrifuge supported by magnetic bearings. We demonstrate the feasibility of the vacuum chamber-free centrifuge by presenting experimental results that verify its high-speed support capability and motoring power capacity.

Long-term cytokine and growth factor release from equine platelet-rich fibrin clots obtained with two different centrifugation protocols.

Science.gov (United States)

Jiménez-Aristizabal, Román F; López, Catalina; Álvarez, María E; Giraldo, Carlos; Prades, Marta; Carmona, Jorge U

2017-09-01

To compare the temporal release (over three weeks) of tumor necrosis factor alpha (TNF-α), interleukin 4 (IL-4), IL-1 receptor antagonist (IL-1ra), platelet-derived growth factor BB (PDGF-BB) and transforming growth factor beta-1 (TGF-β 1 ) from two platelet-rich fibrin (PRF) preparations from equine blood obtained at either 240g/8min or 416g/10min. Whole blood from 10 horses was used to obtain PRF clots by two different centrifugation protocols. After 1h of rest, PRF clots were deposited in wells with culture medium, which was changed at 6h, 24h and then every 48h to 21days. Cytokines and GFs were measured by ELISA at 1h (serum supernatants from PRF clots) and all time points of culture medium change. A negative control (plasma) and a positive control (blood lysate) were also included. There were no relevant differences between the two protocols for the temporal release of proteins. However, a significant (p=0.01) effect of time was noted. All cytokines were detected after 6h of PRF clot culture until day 21. GF were detected at 1h until day 21. The concentrations for these proteins diminished gradually over time. A highly significant (p=0.01) correlation was noticed between all the proteins evaluated. Leukocytes enmeshed in PRF clots were able to produce cytokines, TGF-β 1 and PDGF-BB. These findings demonstrate a paramount role of leukocytes in wound healing induced or modified by PRF clots in mammals. Copyright © 2017 Elsevier Ltd. All rights reserved.

Flow control arrangements for centrifuges

International Nuclear Information System (INIS)

Alderton, G.W.; Davidge, P.C.

1983-01-01

In a centrifuge plant for the separation of uranium isotopes, when a centrifuge machine breaks down, light gas is produced. This gas can cause adjacent machines to break down, so propagating the fault. The present invention provides flow control arrangements in gas pipes to the centrifuge, whereby sudden egress of gas from a failed machine is inhibited. (author)

Evaluation of enrichment by centrifugal separation: the future of the centrifugal-separation method

International Nuclear Information System (INIS)

Kanagawa, A.

A gas centrifuge plant for uranium enrichment is considered from the point of view of economic competition with other methods. Characteristics of the method are presented including: energy efficiency, the cascade, the separation coefficient, the equilibrium separation process, and capability as centrifugal pump. The structure of an individual gas centrifuge separator is described including the rotating cylinder, mechanisms for gas injection and extraction, mechanisms for counter-streaming of gas, the axle holder mechanism, the gas sealing mechanism, and the driving mechanism. (U.S.)

[Treatment of surface burns with proteolytic enzymes: mathematic description of lysis kinetics].

Science.gov (United States)

Domogatskaia, A S; Domogatskiĭ, S P; Ruuge, E K

2003-01-01

The lysis of necrotic tissue by a proteolytic enzyme applied to the surface of a burn wound was studied. A mathematical model was proposed, which describes changes in the thickness of necrotic tissue as a function of the proteolytic activity of the enzyme. The model takes into account the inward-directed diffusion of the enzyme, the counterflow of interstitial fluid (exudates) containing specific inhibitors, and the extracellular matrix proteolysis. It was shown in terms of the quasi-stationary approach that the thickness of the necrotic tissue layer decreases exponentially with time; i.e., the lysis slows down as the thickness of the necrotic tissue layer decreases. The dependence of the characteristic time of this decrease on enzyme concentration was obtained. It was shown that, at high enzyme concentrations (more than 5 mg/ml), the entire time of lysis (after the establishment of quasi-stationary equilibrium) is inversely proportional to the concentration of the enzyme.

Polymerase chain reaction and blood culture in blood donors screened by ELISA test for Chagas' disease

Directory of Open Access Journals (Sweden)

Andréa Tieko Kinoshita-Yanaga

2011-03-01

Full Text Available The objective of this study was to evaluate, through blood culture and PCR, the results of the ELISA for Chagas' disease in the screening of blood donors in the public blood-supply network of the state of Paraná, Brazil, and to map the epidemiological profile of the donors with respect to their risk of infection by Trypanosoma cruzi. The negative and positive results of the ELISA were confirmed by blood culture and PCR for 190/191 individuals (99.5%. For one individual (0.5%, the ELISA was inconclusive, blood culture and IIF were negative, and IHA and PCR positive. Three individuals (1.6% were positive for T. cruzi on all the tests. Donors were predominantly female, and natives of Paraná, of rural origin, had observed or been informed of the presence of the vector in the municipalities where they resided, had never received a blood transfusion, had donated blood 1 to 4 times, and reported no cases of Chagas' disease in their families. We concluded that PCR and blood culturing have excellent potential for confirming the results of the ELISA, and that candidate blood donors with negative or positive tests have a similar risk of infection by T. cruzi, indicating that the ELISA test is sufficiently safe for screening blood prior to use.O objetivo deste estudo foi avaliar, pela hemocultura e PCR, os resultados do teste ELISA utilizado para doença de Chagas na triagem de doadores de sangue na rede pública do Estado do Paraná, Brasil, e traçar o perfil epidemiológico dos doadores quanto ao risco de infecção pelo Trypanosoma cruzi. Os resultados negativos e positivos do ELISA foram confirmados pela hemocultura e PCR em 190/191 indivíduos (99,5%. Para um indivíduo (0,5%, o teste de ELISA foi inconclusivo, hemocultura e IFI foram negativas, HAI e PCR foram positivas. Três indivíduos (1,6% foram positivos para T. cruzi em todos os testes. A maioria dos doadores era do sexo feminino, oriundos do Estado do Paraná, de origem rural, tinham

Applying of centrifugal chromatography on DEAE cellulose and viscosity measurement to estimate damage caused by gamma irradiation in lymphocyte DNA

International Nuclear Information System (INIS)

Olinski, R.

1977-01-01

DNA isolated from limphocytes of pig blood was irradiated by γ radiation in the range of 0.5-50 Krads. Changes caused by irradiation (single and double breaks) were determined by using viscosity measurement and centrifugal chromatography on DEAE cellulose. Study of DNA chromatograms showed possibility to apply centrifugal chromatography on DEAE cellulose to estimate changes caused by irradiation. (author)

Significance of coagulase negative Staphylococcus from blood cultures: persisting problems and partial progress in resource constrained settings.

Science.gov (United States)

Sidhu, Shailpreet K; Malhotra, Sita; Devi, Pushpa; Tuli, Arpandeep K

2016-12-01

Coagulase negative Staphylococcus (CoNS) is frequently isolated from blood cultures but their significance is difficult to interpret. CoNS bacteria which are often previously dismissed as culture contaminants are attracting greater importance as true pathogens in the past decades. Clinical evaluation of these isolates suggests that although there is a relative increase of CoNS associated bloodstream infections in recent years, the microorganisms still remain the most common contaminants in blood cultures. The objective of this study was to determine the significance of CoNS isolated from blood cultures. A retrospective study was conducted to evaluate the rate of contamination in blood cultures in a tertiary care hospital. The paired specimens of blood were cultured using conventional culture methods and the isolates of coagulase negative staphylococci were identified by standard methodology. Clinical data, laboratory indices, microbiological parameters and patient characteristics were analyzed. Of 3503 blood samples, CoNS were isolated from blood culture of 307 patients (8.76%). The isolates were reported as true pathogens of bloodstream infections in only 74 out of 307 cases (24.1%). In the vast majority, 212 of 307 (69.0%), they were mere blood culture contaminants and reported as insignificant/contaminant. Determining whether a growth in the blood culture is a pathogen or a contaminant is a critical issue and multiple parameters have to be considered before arriving at a conclusion. Ideally, the molecular approach is for the most part a consistent method in determining the significant isolates of CoNS. However, in countries with inadequate resources, species identification and antibiogram tests are recommended when determining significance of these isolates.

A Rapid Centrifugation-Assisted Solid-Phase Extraction and Liquid Chromatography Method for Determination of Loureirin A and Loureirin B of Dragon's Blood Capsules in Rat Plasma and Urine After Oral Administration.

Science.gov (United States)

Chen, Xiaoshuang; Li, Gaofeng; Ma, Shangfang; Hu, Xujia

2015-07-01

A simple, sensitive and rapid centrifugation-assisted solid-phase extraction (SPE) with high-performance liquid chromatography (SPE-HPLC) method was developed for simultaneous determination of the metabolites loureirin A and loureirin B from Dragon's blood in rat plasma and urine. The development of the extraction procedure included optimization of some important extraction phases. After evaluation, the metabolites of Dragon's blood were extracted by centrifugation-assisted SPE and separated by using HPLC. This method showed good linearity (r(2) > 0.99), and in the rat plasma and urine, the recoveries were 93.1 and 95.7% for loureirin A and were 90.1 and 94.2% for loureirin B. The relative standard deviation (RSD) values of intraday and interday precision in rat plasma and urine for loureirin A were <3.84 and 2.01%, respectively. The RSD values of the intraday and interday precision in rat plasma and urine for loureirin B were below 4.25 and 5.83%, respectively. Thus, the established method is suitable for metabolism studies of loureirin A and loureirin B in rat plasma and urine. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Centrifugal Compressor Aeroelastic Analysis Code

Science.gov (United States)

Keith, Theo G., Jr.; Srivastava, Rakesh

2002-01-01

Centrifugal compressors are very widely used in the turbomachine industry where low mass flow rates are required. Gas turbine engines for tanks, rotorcraft and small jets rely extensively on centrifugal compressors for rugged and compact design. These compressors experience problems related with unsteadiness of flowfields, such as stall flutter, separation at the trailing edge over diffuser guide vanes, tip vortex unsteadiness, etc., leading to rotating stall and surge. Considerable interest exists in small gas turbine engine manufacturers to understand and eventually eliminate the problems related to centrifugal compressors. The geometric complexity of centrifugal compressor blades and the twisting of the blade passages makes the linear methods inapplicable. Advanced computational fluid dynamics (CFD) methods are needed for accurate unsteady aerodynamic and aeroelastic analysis of centrifugal compressors. Most of the current day industrial turbomachines and small aircraft engines are designed with a centrifugal compressor. With such a large customer base and NASA Glenn Research Center being, the lead center for turbomachines, it is important that adequate emphasis be placed on this area as well. Currently, this activity is not supported under any project at NASA Glenn.

Blood Culture Testing via a Mobile App That Uses a Mobile Phone Camera: A Feasibility Study.

Science.gov (United States)

Lee, Guna; Lee, Yura; Chong, Yong Pil; Jang, Seongsoo; Kim, Mi Na; Kim, Jeong Hoon; Kim, Woo Sung; Lee, Jae-Ho

2016-10-26

To evaluate patients with fever of unknown origin or those with suspected bacteremia, the precision of blood culture tests is critical. An inappropriate step in the test process or error in a parameter could lead to a false-positive result, which could then affect the direction of treatment in critical conditions. Mobile health apps can be used to resolve problems with blood culture tests, and such apps can hence ensure that point-of-care guidelines are followed and processes are monitored for blood culture tests. In this pilot project, we aimed to investigate the feasibility of using a mobile blood culture app to manage blood culture test quality. We implemented the app at a university hospital in South Korea to assess the potential for its utilization in a clinical environment by reviewing the usage data among a small group of users and by assessing their feedback and the data related to blood culture sampling. We used an iOS-based blood culture app that uses an embedded camera to scan the patient identification and sample number bar codes. A total of 4 medical interns working at 2 medical intensive care units (MICUs) participated in this project, which spanned 3 weeks. App usage and blood culture sampling parameters (including sampler, sampling site, sampling time, and sample volume) were analyzed. The compliance of sampling parameter entry was also measured. In addition, the participants' opinions regarding patient safety, timeliness, efficiency, and usability were recorded. In total, 356/644 (55.3%) of all blood culture samples obtained at the MICUs were examined using the app, including 254/356 (71.3%) with blood collection volumes of 5-7 mL and 256/356 (71.9%) with blood collection from the peripheral veins. The sampling volume differed among the participants. Sampling parameters were completely entered in 354/356 cases (99.4%). All the participants agreed that the app ensured good patient safety, disagreed on its timeliness, and did not believe that it was

Comparison of four methods for rapid identification of Staphylococcus aureus directly from BACTEC 9240 blood culture system.

Science.gov (United States)

Ozen, N S; Ogunc, D; Mutlu, D; Ongut, G; Baysan, B O; Gunseren, F

2011-01-01

Differentiation of Staphylococcus aureus (S. aureus) from coagulase-negative staphylococci is very important in blood stream infections. Identification of S. aureus and coagulase-negative staphylococci (CoNS) from blood cultures takes generally 18-24 h after positive signaling on continuously monitored automated blood culture system. In this study, we evaluated the performance of tube coagulase test (TCT), slide agglutination test (Dry Spot Staphytect Plus), conventional polymerase chain reaction (PCR) and LightCycler Staphylococcus MGrade kit directly from blood culture bottles to achieve rapid identification of S. aureus by using the BACTEC 9240 blood culture system. A total of 129 BACTEC 9240 bottles growing gram-positive cocci suggesting Staphylococci were tested directly from blood culture broths (BCBs) with TCT, Dry Spot Staphytect Plus, conventional PCR and LightCycler Staphylococcus MGrade kit for rapid identification of S. aureus. The sensitivities of the tests were 99, 68, 99 and 100%, respectively. Our results suggested that 2 h TCT was found to be simple and inexpensive method for the rapid identification of S. aureus directly from positive blood cultures.

The preanalytical optimization of blood cultures: a review and the clinical importance of benchmarking in 5 Belgian hospitals.

Science.gov (United States)

Willems, Elise; Smismans, Annick; Cartuyvels, Reinoud; Coppens, Guy; Van Vaerenbergh, Kristien; Van den Abeele, Anne-Marie; Frans, Johan

2012-05-01

Bloodstream infections remain a major challenge in medicine. Optimal detection of pathogens is only possible if the quality of preanalytical factors is thoroughly controlled. Since the laboratory is responsible for this preanalytical phase, the quality control of critical factors should be integrated in its quality control program. The numerous recommendations regarding blood culture collection contain controversies. Only an unambiguous guideline permits standardization and interlaboratory quality control. We present an evidence-based concise guideline of critical preanalytical determinants for blood culture collection and summarize key performance indicators with their concomitant target values. In an attempt to benchmark, we compared the true-positive rate, contamination rate, and collected blood volume of blood culture bottles in 5 Belgian hospital laboratories. The true-positive blood culture rate fell within previously defined acceptation criteria by Baron et al. (2005) in all 5 hospitals, whereas the contamination rate exceeded the target value in 4 locations. Most unexpected, in each of the 5 laboratories, more than one third of the blood culture bottles were incorrectly filled, irrespective of the manufacturer of the blood culture vials. As a consequence of this shortcoming, one manufacturer recently developed an automatic blood volume monitoring system. In conclusion, clear recommendations for standardized blood culture collection combined with quality control of critical factors of the preanalytical phase are essential for diagnostic blood culture improvement. Copyright © 2012 Elsevier Inc. All rights reserved.

Nucleation of holin domains and holes optimizes lysis timing of E. coli by phage λ

Science.gov (United States)

Ryan, Gillian; Rutenberg, Andrew

2007-03-01

Holin proteins regulate the precise scheduling of Escherichia coli lysis during infection by bacteriophage λ. Inserted into the host bacterium's inner membrane during infection, holins aggregate to form rafts and then holes within those rafts. We present a two-stage nucleation model of holin action, with the nucleation of condensed holin domains followed by the nucleation of holes within these domains. Late nucleation of holin rafts leads to a weak dependence of lysis timing on host cell size, though both nucleation events contribute equally to timing errors. Our simulations recover the accurate scheduling observed experimentally, and also suggest that phage-λ lysis of E.coli is optimized.

The magnetic centrifugal mass filter

International Nuclear Information System (INIS)

Fetterman, Abraham J.; Fisch, Nathaniel J.

2011-01-01

Mass filters using rotating plasmas have been considered for separating nuclear waste and spent nuclear fuel. We propose a new mass filter that utilizes centrifugal and magnetic confinement of ions in a way similar to the asymmetric centrifugal trap. This magnetic centrifugal mass filter is shown to be more proliferation resistant than present technology. This filter is collisional and produces well confined output streams, among other advantages.

The magnetic centrifugal mass filter

Energy Technology Data Exchange (ETDEWEB)

Fetterman, Abraham J.; Fisch, Nathaniel J. [Department of Astrophysical Sciences, Princeton University, Princeton, New Jersey 08540 (United States)

2011-09-15

Mass filters using rotating plasmas have been considered for separating nuclear waste and spent nuclear fuel. We propose a new mass filter that utilizes centrifugal and magnetic confinement of ions in a way similar to the asymmetric centrifugal trap. This magnetic centrifugal mass filter is shown to be more proliferation resistant than present technology. This filter is collisional and produces well confined output streams, among other advantages.

A technique for separation of canine lymphocytes and their use in the lymphocytotoxic, blastogenic, and rosette assys.

Science.gov (United States)

Whitacre, C C; Lang, R W

1975-01-01

When the techniques established for preparing lymphocytes from human blood were applied to canine blood, the resulting preparations were contaminated with significant numbers of other blood cells. A three-step technique is described here for the separation of canine lymphocytes from blood which eliminates most granulocytes, platelets, and erythrocytes. Defibrinated blood is incubated with iron particles to allow phagocytosis by granulocytes. When mixed with Plasmagel, erythrocytes and iron-containing granulocytes sediment rapidly, leaving a lymphocyte-rich supernate plasma which is recovered and banded by centrifugation on a Ficoll-Hypaque gradient. Any erythrocytes remaining in the lymphocyte suspension aspirated from the gradient are eliminated by hypotonic lysis and centrifugation. The resulting preparation, representing 30 per cent recovery of the circulating lymphocytes, contained 85 to 94 per cent lymphocytes with a cell viability of 99 per cent. The functional capacity of the lymphocyte preparation was tested in three assays: 1) lymphoblastic transformation using phytohemagglutinin and pokeweed mitogen, 2) E rosette formation with human erythrocytes showed a mean 5 per cent rosette forming lymphocytes, 3) antibody-mediated lymphocytotoxicity using a canine anti-lymphocyte serum showed titers reproducible within one tube dilution.

Hyperphosphatemia during spontaneous tumor lysis syndrome culminate in severe hyphosphatemia at the time of blast crisis of Phneg CML to acute myelomoncytic leukemia

Directory of Open Access Journals (Sweden)

Salomon Ophira

2012-08-01

Full Text Available Abstract Extreme swing of phosphor from severe hyperphosphatemia to severe hypophosphatemia in a patient with blast crisis of myeloid origin was the result of imbalance between massive apoptosis of leukemic cells in the context of spontaneous tumor lysis syndrome and massive production of leukemic cells with only 1% of blast in peripheral blood. The mutated p53 protein suggested acting as oncogene in the presented case and possibly affecting phosphor status.

Open-heart surgery using a centrifugal pump: a case of hereditary spherocytosis.

Science.gov (United States)

Matsuzaki, Yuichi; Tomioka, Hideyuki; Saso, Masaki; Azuma, Takashi; Saito, Satoshi; Aomi, Shigeyuki; Yamazaki, Kenji

2016-08-26

Hereditary spherocytosis is a genetic, frequently familial hemolytic blood disease characterized by varying degrees of hemolytic anemia, splenomegaly, and jaundice. There are few reports on adult open-heart surgery for patients with hereditary spherocytosis. We report a rare case of an adult open-heart surgery associated with hereditary spherocytosis. A 63-year-old man was admitted for congestive heart failure due to bicuspid aortic valve, aortic valve regurgitation, and sinus of subaortic aneurysm. The family history, the microscopic findings of the blood smear, and the characteristic osmotic fragility confirmed the diagnosis of hereditary spherocytosis. Furthermore, splenectomy had not been undertaken preoperatively. The patient underwent a successful operation by means of a centrifugal pump. Haptoglobin was used during the cardiopulmonary bypass, and a biological valve was selected to prevent hemolysis. No significant hemolysis occurred intraoperatively or postoperatively. There are no previous reports of patients with hereditary spherocytosis, and bicuspid aortic valve. We have successfully performed an adult open-heart surgery using a centrifugal pump in an adult patient suffering from hereditary spherocytosis and bicuspid aortic valve.

Pre-storage centrifugation conditions have significant impact on measured microRNA levels in biobanked EDTA plasma samples

DEFF Research Database (Denmark)

Binderup, Helle Glud; Houlind, Kim; Madsen, Jonna Skov

2016-01-01

in platelets this may partly be explained by residual platelets in the plasma samples used. When collecting fresh plasma samples, it is possible to produce cell-free/platelet-poor plasma by centrifugation. In this study, we systematically investigated whether biobanked EDTA plasma samples could be processed...... to be suitable for miRNA analysis. Materials and methods: Blood samples were collected from ten healthy volunteers and centrifuged to produce platelet-poor-plasma (PPP) and standard biobank plasma. After one week at -80 °C the biobanked EDTA plasma was re-centrifuged by different steps to remove residual...... platelets. Using RT-qPCR the levels of 14 miRNAs in the different plasma preparations were compared to that of PPP. Results: We were able to remove residual platelets from biobanked EDTA plasma by re-centrifugation of the thawed samples. Nevertheless, for most of the investigated miRNAs, the miRNA level...

Sustained Accelerated Idioventricular Rhythm in a Centrifuge-Simulated Suborbital Spaceflight.

Science.gov (United States)

Suresh, Rahul; Blue, Rebecca S; Mathers, Charles; Castleberry, Tarah L; Vanderploeg, James M

2017-08-01

Hypergravitational exposures during human centrifugation are known to provoke dysrhythmias, including sinus dysrhythmias/tachycardias, premature atrial/ventricular contractions, and even atrial fibrillations or flutter patterns. However, events are generally short-lived and resolve rapidly after cessation of acceleration. This case report describes a prolonged ectopic ventricular rhythm in response to high G exposure. A previously healthy 30-yr-old man voluntarily participated in centrifuge trials as a part of a larger study, experiencing a total of 7 centrifuge runs over 48 h. Day 1 consisted of two +Gz runs (peak +3.5 Gz, run 2) and two +Gx runs (peak +6.0 Gx, run 4). Day 2 consisted of three runs approximating suborbital spaceflight profiles (combined +Gx and +Gz). Hemodynamic data collected included blood pressure, heart rate, and continuous three-lead electrocardiogram. Following the final acceleration exposure of the last Day 2 run (peak +4.5 Gx and +4.0 Gz combined, resultant +6.0 G), during a period of idle resting centrifuge activity (resultant vector +1.4 G), the subject demonstrated a marked change in his three-lead electrocardiogram from normal sinus rhythm to a wide-complex ectopic ventricular rhythm at a rate of 91-95 bpm, consistent with an accelerated idioventricular rhythm (AIVR). This rhythm was sustained for 2 m, 24 s before reversion to normal sinus. The subject reported no adverse symptoms during this time. While prolonged, the dysrhythmia was asymptomatic and self-limited. AIVR is likely a physiological response to acceleration and can be managed conservatively. Vigilance is needed to ensure that AIVR is correctly distinguished from other, malignant rhythms to avoid inappropriate treatment and negative operational impacts.Suresh R, Blue RS, Mathers C, Castleberry TL, Vanderploeg JM. Sustained accelerated idioventricular rhythm in a centrifuge-simulated suborbital spaceflight. Aerosp Med Hum Perform. 2017; 88(8):789-793.

Displacement of the mitotic apparatuses by centrifugation reveals cortical actin organization during cytokinesis in cultured tobacco BY-2 cells.

Science.gov (United States)

Arima, Kengo; Tamaoki, Daisuke; Mineyuki, Yoshinobu; Yasuhara, Hiroki; Nakai, Tomonori; Shimmen, Teruo; Yoshihisa, Tohru; Sonobe, Seiji

2018-06-19

In plant cytokinesis, actin is thought to be crucial in cell plate guidance to the cortical division zone (CDZ), but its organization and function are not fully understood. To elucidate actin organization during cytokinesis, we employed an experimental system, in which the mitotic apparatus is displaced and separated from the CDZ by centrifugation and observed using a global-local live imaging microscope that enabled us to record behavior of actin filaments in the CDZ and the whole cell division process in parallel. In this system, returning movement of the cytokinetic apparatus in cultured-tobacco BY-2 cells occurs, and there is an advantage to observe actin organization clearly during the cytokinetic phase because more space was available between the CDZ and the distantly formed phragmoplast. Actin cables were clearly observed between the CDZ and the phragmoplast in BY-2 cells expressing GFP-fimbrin after centrifugation. Both the CDZ and the edge of the expanding phragmoplast had actin bulges. Using live-cell imaging including the global-local live imaging microscopy, we found actin filaments started to accumulate at the actin-depleted zone when cell plate expansion started even in the cell whose cell plate failed to reach the CDZ. These results suggest that specific accumulation of actin filaments at the CDZ and the appearance of actin cables between the CDZ and the phragmoplast during cell plate formation play important roles in the guidance of cell plate edges to the CDZ.

Reduction of relative centrifugation force within injectable platelet-rich-fibrin (PRF) concentrates advances patients' own inflammatory cells, platelets and growth factors: the first introduction to the low speed centrifugation concept.

Science.gov (United States)

Choukroun, J; Ghanaati, S

2018-02-01

The aim of this study was to analyze systematically the influence of the relative centrifugation force (RCF) on leukocytes, platelets and growth factor release within fluid platelet-rich fibrin matrices (PRF). Systematically using peripheral blood from six healthy volunteers, the RCF was reduced four times for each of the three experimental protocols (I-III) within the spectrum (710-44 g), while maintaining a constant centrifugation time. Flow cytometry was applied to determine the platelets and leukocyte number. The growth factor concentration was quantified 1 and 24 h after clotting using ELISA. Reducing RCF in accordance with protocol-II (177 g) led to a significantly higher platelets and leukocytes numbers compared to protocol-I (710 g). Protocol-III (44 g) showed a highly significant increase of leukocytes and platelets number in comparison to -I and -II. The growth factors' concentration of VEGF and TGF-β1 was significantly higher in protocol-II compared to -I, whereas protocol-III exhibited significantly higher growth factor concentration compared to protocols-I and -II. These findings were observed among 1 and 24 h after clotting, as well as the accumulated growth factor concentration over 24 h. Based on the results, it has been demonstrated that it is possible to enrich PRF-based fluid matrices with leukocytes, platelets and growth factors by means of a single alteration of the centrifugation settings within the clinical routine. We postulate that the so-called low speed centrifugation concept (LSCC) selectively enriches leukocytes, platelets and growth factors within fluid PRF-based matrices. Further studies are needed to evaluate the effect of cell and growth factor enrichment on wound healing and tissue regeneration while comparing blood concentrates gained by high and low RCF.

Theoretical investigations on plasma centrifuges

International Nuclear Information System (INIS)

Hong, S.H.

1978-01-01

The theoretical analysis of the steady-state dynamics of plasma centrifuges is dealt with to understand the physics of rotating plasmas and their feasibility for isotope separation. The centrifuge systems under consideration employ cylindrical gas discharge chambers with externally-applied axial magnetic fields. The cathode and anode are symmetric about the cylinder axis and arranged in such a way for each system, i.e., (1) two ring electrodes of different radii in the chamber end plates or (2) two ring electrodes embedded in the mantle of the cylinder. They produce converging and/or diverging current density field lines, which intersect the external magnetic field under a nonvanishing angle. The associated Lorentz forces set the plasma, which is produced through an electrical discharge, into rotation around the cylinder axis. Three boundary-value problems for the coupled partial differential equations of the centrifuge fields are formulated, respectively, on the basis of the magnetogasdynamic equations. The electric field, electrostatic potential, current density, induced magnetic field, and velocity distributions are discussed in terms of the Hartmann number, the Hall coefficient, and the magnetic Reynolds number. The plasma centrifuge analyses presented show that the speeds of plasma rotation up to the order of 10 4 m/sec are achievable at typical conditions. The associated centrifugal forces produce a significant spatial isotope separation, which is somewhat reduced in the viscous boundary layers at the centrifuge walls. The speeds of plasma rotation increase with increasing Hartmann number and Hall coefficient. For small Hall coefficient, the induced azimuthal magnetic field does not affect the plasma rotation. For large volumes of rotating isotope mixtures, a multidischarge centrifuge can be constructed by setting up a large number of centrifuge systems in series

Characterization of glucocerebrosidase in peripheral blood cells and cultured blastoid cells

NARCIS (Netherlands)

Aerts, J. M.; Heikoop, J.; van Weely, S.; Donker-Koopman, W. E.; Barranger, J. A.; Tager, J. M.; Schram, A. W.

1988-01-01

We have characterized glucocerebrosidase in various cell types of peripheral blood of control subjects and in cultured human blastoid cells. The intracellular level of glucocerebrosidase in cultured blastoid cells (10-30 nmol substrate hydrolyzed/h.mg protein) resembles closely values observed for

21 CFR 864.5350 - Microsedimentation centrifuge.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Microsedimentation centrifuge. 864.5350 Section 864.5350 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES... § 864.5350 Microsedimentation centrifuge. (a) Identification. A microsedimentation centrifuge is a...

Impact of antibiotic administration on blood culture positivity at the beginning of sepsis: a prospective clinical cohort study.

Science.gov (United States)

Scheer, Christian S; Fuchs, Christian; Gründling, Matthias; Vollmer, Marcus; Bast, Juliane; Bohnert, Jürgen A; Zimmermann, Kathrin; Hahnenkamp, Klaus; Rehberg, Sebastian; Kuhn, Sven-Olaf

2018-06-04

Sepsis guidelines recommend obtaining blood cultures before starting anti-infective therapy in patients with sepsis. However, little is known how antibiotic treatment prior to sampling affects bacterial growth. The aim of this study was to compare the results of blood cultures drawn prior to and under antibiotic therapy. Prospective clinical cohort study of septic patients. Adult ICU patients with 2 or 3 blood culture (BC) sets at the beginning of sepsis between 2010 and 2017 were included. Patients with blood culture samplings obtained prior to antibiotic therapy were compared to patients with samplings under antibiotic therapy. Blood culture positivity, defined as microbiological pathogen finding, was compared between the groups. Logistic regression was performed to adjust the impact of different factors with respect to blood culture positivity. In total, 559 patients with 1364 blood culture sets at the beginning of sepsis were analyzed. BC positivity was 50.6% (78/154) among septic patients who did not receive antibiotics and only 27.7% (112/405) in those who were already under antibiotics (Pcultures under antibiotic therapy is associated with a significant loss of pathogen detection. This strongly emphasizes the current recommendation to obtain blood cultures prior to antibiotic administration in patients with sepsis. Copyright © 2018. Published by Elsevier Ltd.

Uranium enrichment by gas centrifuge

International Nuclear Information System (INIS)

Heriot, I.D.

1988-01-01

After recalling the physical principles and the techniques of centrifuge enrichment the report describes the centrifuge enrichment programmes of the various countries concerned and compares this technology with other enrichment technologies like gaseous diffusion, laser, aerodynamic devices and chemical processes. The centrifuge enrichment process is said to be able to replace with advantage the existing enrichment facilities in the short and medium term. Future prospects of the process are also described, like recycled uranium enrichment and economic improvements; research and development needs to achieve the economic prospects are also indicated. Finally the report takes note of the positive aspect of centrifuge enrichment as far as safeguards and nuclear safety are concerned. 27 figs, 113 refs

Decreased mortality associated with prompt Gram staining of blood cultures.

Science.gov (United States)

Barenfanger, Joan; Graham, Donald R; Kolluri, Lavanya; Sangwan, Gaurav; Lawhorn, Jerry; Drake, Cheryl A; Verhulst, Steven J; Peterson, Ryan; Moja, Lauren B; Ertmoed, Matthew M; Moja, Ashley B; Shevlin, Douglas W; Vautrain, Robert; Callahan, Charles D

2008-12-01

Gram stains of positive blood cultures are the most important factor influencing appropriate therapy. The sooner appropriate therapy is initiated, the better. Therefore, it is reasonable to expect that the sooner Gram stains are performed, the better. To determine the value of timely Gram stains and whether improvement in Gram stain turnaround time (TAT) is feasible, we compared data for matched pairs of patients with cultures processed promptly ( or =1 hour TAT) and then monitored TAT by control charting.In 99 matched pairs, average difference in time to detection of positive blood cultures within a pair of patients was less than 0.1 hour. For the less than 1 hour TAT group, the average TAT and crude mortality were 0.1 hour and 10.1%, respectively; for the 1 hour or longer TAT group, they were 3.3 hours and 19.2%, respectively (P Gram stains.

transfusion transmissible viral infections among potential blood

African Journals Online (AJOL)

boaz

Blood sample was spun on a bench centrifuge at 3,000rpm for 10 minutes to obtain serum. Serum or plasma was separated immediately. Data Collection and Laboratory Methods. A survey of the blood sample of the prospective donors at the Blood Bank, University College. Hospital (UCH) Ibadan was conducted between.

Comparison of four methods for rapid identification of Staphylococcus aureus directly from BACTEC 9240 blood culture system

Directory of Open Access Journals (Sweden)

N S Ozen

2011-01-01

Full Text Available Purpose: Differentiation of Staphylococcus aureus (S. aureus from coagulase-negative staphylococci is very important in blood stream infections. Identification of S. aureus and coagulase-negative staphylococci (CoNS from blood cultures takes generally 18-24 h after positive signaling on continuously monitored automated blood culture system. In this study, we evaluated the performance of tube coagulase test (TCT, slide agglutination test (Dry Spot Staphytect Plus, conventional polymerase chain reaction (PCR and LightCycler Staphylococcus MGrade kit directly from blood culture bottles to achieve rapid identification of S. aureus by using the BACTEC 9240 blood culture system. Materials and Methods: A total of 129 BACTEC 9240 bottles growing gram-positive cocci suggesting Staphylococci were tested directly from blood culture broths (BCBs with TCT, Dry Spot Staphytect Plus, conventional PCR and LightCycler Staphylococcus MGrade kit for rapid identification of S. aureus. Results: The sensitivities of the tests were 99, 68, 99 and 100%, respectively. Conclusion: Our results suggested that 2 h TCT was found to be simple and inexpensive method for the rapid identification of S. aureus directly from positive blood cultures.

Tolerance of centrifuge-simulated suborbital spaceflight by medical condition.

Science.gov (United States)

Blue, Rebecca S; Pattarini, James M; Reyes, David P; Mulcahy, Robert A; Garbino, Alejandro; Mathers, Charles H; Vardiman, Johnené L; Castleberry, Tarah L; Vanderploeg, James M

2014-07-01

We examined responses of volunteers with known medical disease to G forces in a centrifuge to evaluate how potential commercial spaceflight participants (SFPs) might tolerate the forces of spaceflight despite significant medical history. Volunteers were recruited based upon suitability for each of five disease categories (hypertension, cardiovascular disease, diabetes, lung disease, back or neck problems) or a control group. Subjects underwent seven centrifuge runs over 2 d. Day 1 consisted of two +G(z) runs (peak = +3.5 G(z), Run 2) and two +G(x), runs (peak = +6.0 G(x), Run 4). Day 2 consisted of three runs approximating suborbital spaceflight profiles (combined +G(x) and +G(z), peak = +6.0 G(x)/+4.0 G(z)). Data collected included blood pressure, electrocardiogram, pulse oximetry, neurovestibular exams, and post-run questionnaires regarding motion sickness, disorientation, grayout, and other symptoms. A total of 335 subjects registered for participation, of which 86 (63 men, 23 women, age 20-78 yr) participated in centrifuge trials. The most common causes for disqualification were weight and severe and uncontrolled medical or psychiatric disease. Five subjects voluntarily withdrew from the second day of testing: three for anxiety reasons, one for back strain, and one for time constraints. Maximum hemodynamic values recorded included HR of 192 bpm, systolic BP of 217 mmHg, and diastolic BP of 144 mmHg. Common subjective complaints included grayout (69%), nausea (20%), and chest discomfort (6%). Despite their medical history, no subject experienced significant adverse physiological responses to centrifuge profiles. These results suggest that most individuals with well-controlled medical conditions can withstand acceleration forces of launch and re-entry profiles of current commercial spaceflight vehicles.

Effect of ultrasonic frequency on the mechanism of formic acid sono-lysis

International Nuclear Information System (INIS)

Chave, T.; Nikitenko, S.I.; Navarro, N.M.; Pochon, P.; Bisel, I.

2011-01-01

The kinetics and mechanism of formic acid sono-chemical degradation were studied at ultrasonic frequencies of 20, 200, and 607 kHz under argon atmosphere. Total yield of HCOOH sono-chemical degradation increases approximately 6-8-fold when the frequency increased from 20 to 200 or to 607 kHz. At low ultrasonic frequencies, HCOOH degradation has been attributed to oxidation with OH . radicals from water sono-lysis and to the HCOOH decarboxylation occurring at the cavitation bubble-liquid interface. With high-frequency ultrasound, the sono-chemical reaction is also influenced by HCOOH dehydration. Whatever the ultrasonic frequency, the sono-lysis of HCOOH yielded H 2 and CO 2 in the gas phase as well as trace, amounts of oxalic acid and formaldehyde in the liquid phase. However, CO and CH 4 formations were only detected under high frequency ultrasound. The most striking difference between low frequency and high frequency ultrasound is that the sono-lysis of HCOOH at high ultrasonic frequencies initiates Fischer-Tropsch hydrogenation of carbon monoxide. (authors)

Genotoxic damage in cultured human peripheral blood lymphocytes ...

African Journals Online (AJOL)

Falaq Naz

2012-06-29

Jun 29, 2012 ... Genotoxic damage in cultured human peripheral blood lymphocytes of oral ... catechol estrogens and quinines, via redox reactions causes oxidative damage to .... volume was prepared for each donor. About, 0.8 ml of cell sus .... duce the adverse effects of OCs, such as the reduction in the estrogen content.

TNF induction of EL4 hyposensitivity to lysis by recombinant (soluble) and membrane-associated TNFs: TNF binding, internalization, and degradation.

Science.gov (United States)

Fishman, M; Costlow, M

1994-04-01

EL4 mouse thymoma cells sensitive to TNF-mediated lysis only in the presence of cycloheximide (S-EL4) or in the presence or absence of cycloheximide (N-EL4) were used in these experiments. Murine tumor cell line (S-EL4) sensitivity to TNF cytotoxicity is augmented when cycloheximide is added together with TNF or when cycloheximide is added 1 hr before or after TNF. No enhanced sensitivity is observed when target cells are incubated with cycloheximide 2-4 hr before or after the addition of TNF. In the absence of cycloheximide, S-EL4 cells preexposed to murine TNF are less susceptible to lysis by TNF and TNF receptor-conjugated TNF but are lysed by integral membrane TNF. TNF-induced hyposensitivity is partially reversed by actinomycin D or by culturing the preexposed cells for 4 hr prior to TNF lytic assay. TNF preincubation of N- and S-EL4 cells results in an immediate decrease in 125I-TNF binding due to TNF receptor occupancy. Recovery of TNF-R occupancy and TNF internalization were subsequently noted.

The analytic impact of a reduced centrifugation step on chemistry and immunochemistry assays: an evaluation of the Modular Pre-Analytics.

Science.gov (United States)

Koenders, Mieke M J F; van Hurne, Marco E J F; Glasmacher-Van Zijl, Monique; van der Linde, Geesje; Westerhuis, Bert W J J M

2012-09-01

The COBAS 6000 system can be completed by a Modular Pre-Analytics (MPA), an integrated laboratory automation system that streamlines preanalysis. For an optimal throughput, the MPA centrifuges blood collection tubes for 5 min at 1885 × g - a centrifugation time that is not in concordance with the World Health Organization guidelines which suggest centrifugation for 10/15 min at 2000-3000 × g. In this study, the analytical outcome of 50 serum and 50 plasma samples centrifuged for 5 or 10 min at 1885 × g was investigated. The study included routine chemistry and immunochemistry assays on the COBAS 6000 and the Minicap capillary electrophoresis. Deming-fit and Bland-Altman plots of the 5-min and 10-min centrifugation steps indicated a significant correlation in serum samples. The lipaemia index in plasma samples centrifuged for 5 min displayed a statistically significant variation when compared with the 10-min centrifugation. Preanalytical centrifugation can be successfully down-scaled to a duration of 5 min for most routine chemistry and immunochemistry assays in serum and plasma samples. To prevent inaccurate results in plasma samples with an increased lipaemia index from being reported, the laboratory information system was programmed to withhold results above certain lipaemia indices. The presented data support the use of a 5-min centrifugation step to improve turnaround times, thereby meeting one of the desires of the requesting clinicians.

Advancing haemostasis automation--successful implementation of robotic centrifugation and sample processing in a tertiary service hospital.

Science.gov (United States)

Sédille-Mostafaie, Nazanin; Engler, Hanna; Lutz, Susanne; Korte, Wolfgang

2013-06-01

Laboratories today face increasing pressure to automate operations due to increasing workloads and the need to reduce expenditure. Few studies to date have focussed on the laboratory automation of preanalytical coagulation specimen processing. In the present study, we examined whether a clinical chemistry automation protocol meets the preanalytical requirements for the analyses of coagulation. During the implementation of laboratory automation, we began to operate a pre- and postanalytical automation system. The preanalytical unit processes blood specimens for chemistry, immunology and coagulation by automated specimen processing. As the production of platelet-poor plasma is highly dependent on optimal centrifugation, we examined specimen handling under different centrifugation conditions in order to produce optimal platelet deficient plasma specimens. To this end, manually processed models centrifuged at 1500 g for 5 and 20 min were compared to an automated centrifugation model at 3000 g for 7 min. For analytical assays that are performed frequently enough to be targets for full automation, Passing-Bablok regression analysis showed close agreement between different centrifugation methods, with a correlation coefficient between 0.98 and 0.99 and a bias between -5% and +6%. For seldom performed assays that do not mandate full automation, the Passing-Bablok regression analysis showed acceptable to poor agreement between different centrifugation methods. A full automation solution is suitable and can be recommended for frequent haemostasis testing.

Development of a real-time and quantitative thrombus sensor for an extracorporeal centrifugal blood pump by near-infrared light.

Science.gov (United States)

Sakota, Daisuke; Fujiwara, Tatsuki; Ohuchi, Katsuhiro; Kuwana, Katsuyuki; Yamazaki, Hiroyuki; Kosaka, Ryo; Nishida, Masahiro; Mizuno, Tomohiro; Arai, Hirokuni; Maruyama, Osamu

2018-01-01

We developed an optical thrombus sensor for a monopivot extracorporeal centrifugal blood pump. In this study, we investigated its quantitative performance for thrombus detection in acute animal experiments of left ventricular assist using the pump on pathogen-free pigs. Optical fibers were set in the driver unit of the pump. The incident light at the near-infrared wavelength of 810 nm was aimed at the pivot bearing, and the resulting scattered light was guided to the optical fibers. The detected signal was analyzed to obtain the thrombus formation level. As a result, real-time and quantitative monitoring of the thrombus surface area on the pivot bearing was achieved with an accuracy of 3.6 ± 2.3 mm 2 . In addition, the sensing method using the near-infrared light was not influenced by changes in the oxygen saturation and the hematocrit. It is expected that the developed sensor will be useful for optimal anticoagulation management for long-term extracorporeal circulation therapies.

Study of the phage production efficiency in the bacteria lysis processes

International Nuclear Information System (INIS)

Vidania Munoz, R. de; Garces, F.; Davila, C. A.

1979-01-01

In this work we present a search for the best production conditions of λvir andλ clear phages In E coli K12 and E coli C 6 00 infected cells respectively. By keeping fixed some parameters of the process as the bacterial and phage generation times and (he bacterial burst side, we have finder that the lysis yield is strongly dependent on the multiplicity and in a lesser degree on the infection time. It appears from the experimental results that other variables are important, as infection efficiency and approach time from phages to bacteria. We will try to describe the lysis phenomenon by a numerical model on the bases of the se experimental results. (Author) 11 refs

Progress in ultra-centrifuge enrichment technology

International Nuclear Information System (INIS)

Paul Dawson

2006-01-01

Urenco have undertaken a continuous development programme in centrifuge technology for over 35 years. This has seen development from sub-critical machines in the mid 1970's through to the company's world leading TC12 supercritical centrifuge, which has been deployed on a large-scale basis over the last decade. The latest centrifuge to emerge from this programme is Urenco's sixth generation centrifuge, the TC21, which will be commercially deployed from mid-2007 onwards. In recent times Urenco has vested its centrifuge technology in Enrichment Technology Company (ETC) as a vehicle to enable the use of this advanced technology by other operators for commercial purposes. This paper reviews why Urenco and ETC believe this technology represents the best choice for creating new global commercial enrichment capacity and its future development prospects. (author)

Noninvasive prenatal diagnosis. Use of density gradient centrifugation, magnetically activated cell sorting and in situ hybridization

DEFF Research Database (Denmark)

Campagnoli, C; Multhaupt, H A; Ludomirski, A

1997-01-01

OBJECTIVE: To develop a noninvasive method suitable for clinical prenatal diagnosis. STUDY DESIGN: Fetal nucleated erythrocytes were separated from peripheral blood of 17 healthy pregnant women using small magnetically activated cell sorting columns (MiniMACS) following density gradient centrifug...

Radiometric detection of yeasts in blood cultures of cancer patients

International Nuclear Information System (INIS)

Hopfer, R.L.; Orengo, A.; Chesnut, S.; Wenglar, M.

1980-01-01

During a 12-month period, 19,457 blood cultures were collected. Yeasts were isolated from 193 cultures derived from 76 cancer patients. Candida albicans or Candida tropicalis accounted for 79% of isolates. Of the three methods compared, the radiometric method required 2.9 days to become positive, blind subculture required 2.6 days, and Gram stains required 1 day. However, the radiometric method was clearly superior in detecting positive cultures, since 73% of all cultures were first detected radiometrically, 22% were detected by subculture, and only 5% were detected by Gram stain. Although 93% of the isolates were detected by aerobic culture, five (7%) isolates were obtained only from anaerobic cultures. Seven days of incubation appear to be sufficient for the radiometric detection of yeasts

HB&L System: rapid determination of antibiotic sensitivity of bacteria isolated from blood cultures.

Directory of Open Access Journals (Sweden)

Simone Barocci

2010-03-01

Full Text Available Introduction. Blood culture is an important method to detect microbial pathogens on blood, very useful for diagnosing bacterial infections. Unfortunately, classical diagnostic protocols cannot directly identify bacteria responsible for sepsis and accordingly their antimicrobial profiles. This problem causes a delay of almost two days in the availability of a specific antimicrobial profile. Objective. Among the main causes of death, sepsis have a relevant importance. For this reason it is important both to identify pathogens and to perform an antimicrobial susceptibility test in the shortest time as possible. For this purpose, the main aim of this study is the evaluation of the performances of an antimicrobial susceptibility determination directly performed on positive blood cultures. Materials and methods. This study has been performed on 70 positive blood cultures, during the period from January to July 2009. A number of 35 blood cultures were positive for Gram negative bacteria, and 35 were positive for Gram positive bacteria. From these positive blood cultures, after a short sample preparation, it has been possible to directly determine antimicrobial susceptibility profiles by using the HB&L (formerly URO-QUICK instrument. Results. The HB&L system results showed a very good correlation with both the classical disk diffusion method and VITEK 2 automatic system.The performances between the methods carried out in this study were equivalent. Conclusions. From data reported, thanks to the rapidity and simplicity of the method used, we can assert that the direct susceptibility test available with the HB&L system, is useful for a rapid and early choice of the antibiotic treatment.

Tolerance of centrifuge-simulated suborbital spaceflight in subjects with implanted insulin pumps.

Science.gov (United States)

Levin, Dana R; Blue, Rebecca S; Castleberry, Tarah L; Vanderploeg, James M

2015-04-01

With commercial spaceflight comes the possibility of spaceflight participants (SFPs) with significant medical conditions. Those with previously untested medical conditions, such as diabetes mellitus (DM) and the use of indwelling medical devices, represent a unique challenge. It is unclear how SFPs with such devices will react to the stresses of spaceflight. This case report describes two subjects with Type I DM using insulin pumps who underwent simulated dynamic phases of spaceflight via centrifuge G force exposure. Two Type I diabetic subjects with indwelling Humalog insulin pumps, a 23-yr-old man averaging 50 u of Humalog daily and a 27-yr-old man averaging 60 u of Humalog daily, underwent seven centrifuge runs over 48 h. Day 1 consisted of two +Gz runs (peak = +3.5 Gz, run 2) and two +Gx runs (peak = +6.0 Gx, run 4). Day 2 consisted of three runs approximating suborbital spaceflight profiles (combined +Gx and +Gz). Data collected included blood pressure, electrocardiogram, pulse oximetry, neurovestibular evaluation, and questionnaires regarding motion sickness, disorientation, greyout, and other symptoms. Neither subject experienced adverse clinical responses to the centrifuge exposure. Both maintained blood glucose levels between 110-206 mg · dl(-1). Potential risks to SFPs with insulin pump dependent DM include hypo/hyperglycemia, pump damage, neurovestibular dysfunction, skin breakdown, and abnormal stress responses. A search of prior literature did not reveal any previous studies of individuals with DM on insulin pumps exposed to prolonged accelerations. These cases suggest that individuals with conditions dependent on continuous medication delivery might tolerate the accelerations anticipated for commercial spaceflight.

Influence of polymer architecture on antigens camouflage, CD47 protection and complement mediated lysis of surface grafted red blood cells.

Science.gov (United States)

Chapanian, Rafi; Constantinescu, Iren; Rossi, Nicholas A A; Medvedev, Nadia; Brooks, Donald E; Scott, Mark D; Kizhakkedathu, Jayachandran N

2012-11-01

Hyperbranched polyglycerol (HPG) and polyethylene glycol (PEG) polymers with similar hydrodynamic sizes in solution were grafted to red blood cells (RBCs) to investigate the impact of polymer architecture on the cell structure and function. The hydrodynamic sizes of polymers were calculated from the diffusion coefficients measured by pulsed field gradient NMR. The hydration of the HPG and PEG was determined by differential scanning calorimetry analyses. RBCs grafted with linear PEG had different properties compared to the compact HPG grafted RBCs. HPG grafted RBCs showed much higher electrophoretic mobility values than PEG grafted RBCs at similar grafting concentrations and hydrodynamic sizes indicating differences in the structure of the polymer exclusion layer on the cell surface. PEG grafting impacted the deformation properties of the membrane to a greater degree than HPG. The complement mediated lysis of the grafted RBCs was dependent on the type of polymer, grafting concentration and molecular size of grafted chains. At higher molecular weights and graft concentrations both HPG and PEG triggered complement activation. The magnitude of activation was higher with HPG possibly due to the presence of many hydroxyl groups per molecule. HPG grafted RBCs showed significantly higher levels of CD47 self-protein accessibility than PEG grafted RBCs at all grafting concentrations and molecular sizes. PEG grafted polymers provided, in general, a better shielding and protection to ABO and minor antigens from antibody recognition than HPG polymers, however, the compact HPGs provided greater protection of certain antigens on the RBC surface. Our data showed that HPG 20 kDa and HPG 60 kDa grafted RBCs exhibited properties that are more comparable to the native RBC than PEG 5 kDa and PEG 10 kDa grafted RBCs of comparable hydrodynamic sizes. The study shows that small compact polymers such as HPG 20 kDa have a greater potential in the generation of functional RBC for therapeutic

Reliability of direct sensitivity determination of blood cultures

International Nuclear Information System (INIS)

Noman, F.; Ahmed, A.

2008-01-01

The aim of this study was to evaluate the error in interpreting antimicrobial sensitivity by direct method when compared to standard method and find out if specific antibiotic-organism combination had more discrepancies. All blood culture samples received at Microbiology Laboratory from 1st July 2006 to 31st August 2006 were ncluded in the study. All samples were inoculated in automated blood culture system BACTEC 9240 which contained enriched Soybean-Casein Digest broth with CO/sub 2/. Once positive, bottles were removed from system; gram staining of the positive broths was done. Susceptibility test was performed from positive broth, on MHA (Mueller-Hinton Agar), with antibiotics panel according to gram stain result. All positive broths were also sub-cultured on blood agar, chocolate agar and McConkey agar for only gram-negative rods. Next day, the zone sizes of all antibiotics were recorded using measuring scale and at the same time susceptibility test was repeated from isolated colonies from subcultures, with inoculums prepared of McFarland 0.5 standard 0.2 Staphylococcus aureus (ATCC 29213); E.coli (ATCC 25922) and Pseudomonas aeruginosa (ATCC 27853) were included as quality control strain. Zone sizes were interpreted as sensitive (S), resistant (R) and intermediate (I) according to CLSI recommendation. Two results were compared and recorded. Out of a total 1083 combinations, zone diameters by standard method were either equal or greater than direct zone diameter (never smaller). Most of the discrepancies were in b-lactam/b-lactamase combinations and aminoglycosides. While reporting these groups of antibiotics with direct sensitivity test, one should be cautious. These are the major antibiotic used for life-threatening infections. In case of being heavy/lighter standard inoculums or marginal zones, repeating with standard method should be preferred to minimize the chances of error. (author)

Empirical Design Considerations for Industrial Centrifugal Compressors

Directory of Open Access Journals (Sweden)

Cheng Xu

2012-01-01

Full Text Available Computational Fluid Dynamics (CFD has been extensively used in centrifugal compressor design. CFD provides further optimisation opportunities for the compressor design rather than designing the centrifugal compressor. The experience-based design process still plays an important role for new compressor developments. The wide variety of design subjects represents a very complex design world for centrifugal compressor designers. Therefore, some basic information for centrifugal design is still very important. The impeller is the key part of the centrifugal stage. Designing a highly efficiency impeller with a wide operation range can ensure overall stage design success. This paper provides some empirical information for designing industrial centrifugal compressors with a focus on the impeller. A ported shroud compressor basic design guideline is also discussed for improving the compressor range.

Lack of requirement for blind subcultures of BACTEC blood culture media

International Nuclear Information System (INIS)

McLaughlin, J.C.; Evers, J.L.; Officer, J.L.

1981-01-01

To determine the need for blind subculturing of BACTEC (Johnston Laboratories, Cockeysville, Md.) blood culture media, we compared results of radiometric readings, visual inspection, and blind subculturing for nearly 7,500 blood specimens. Visual inspection and radiometric testing were performed on day 1 through 7, and blind subcultures were made on day 3. In the first phase of the study, 402 of 3,896 aerobic bottles were positive by radiometric testing (growth index, greater than 25), visual inspection, or subculturing. Only six bottles were radiometrically negative but subculture positive on day 3. The second phase of the study was designed to determine if aerobic bottles eventually became radiometrically positive in those cases in which they were radiometrically negative but subculture positive on day 3. Two bottles were subculture positive but never gave a growth index of greater than or equal to 25 by day 7. One yielded Staphylococcus epidermidis, and one yielded viridans, Streptococcus sp. A total of 35 anaerobic organisms were isolated from 3,896 blood specimens. All of these anaerobes were detected by both radiometric testing and subculturing. We examined a total of 14,972 blood culture bottles. Twenty-nine bottles considered negative by visual inspection or radiometric readings were found to be positive by subculturing. Fifteen of these were shown, by chart review, to contain contaminants. Organisms in the other negative bottles would not have gone undetected because companion bottles from the same patients were radiometrically or visually positive. We concluded that it is necessary to perform blind subcultures of BACTEC 7B and 8B blood culture bottles

Reproducibility of Serum Potassium Values in Serum From Blood Samples Stored for Increasing Times Prior to Centrifugation and Analysis.

Science.gov (United States)

Harper, Aaron; Lu, Chuanyong; Sun, Yi; Garcia, Rafael; Rets, Anton; Alexis, Herol; Saad, Heba; Eid, Ikram; Harris, Loretta; Marshall, Barbara; Tafani, Edlira; Pincus, Matthew R

2016-05-01

The goal of this work was to determine if immediate versus postponed centrifugation of samples affects the levels of serum potassium. Twenty participants donated normal venous blood that was collected in four serum separator tubes per donor, each of which was analyzed at 0, 1, 2, or 4 hr on the Siemens Advia 1800 autoanalyzer. Coefficients of variation (CVs) for potassium levels ranged from 0% to 7.6% with a mean of 3 ± 2%. ANOVA testing of the means for all 20 samples showed a P-value of 0.72 (>0.05) indicating that there was no statistically significant difference between the means of the samples at the four time points. Sixteen samples were found to have CVs that were ≤5%. Two samples showed increases of potassium from the reference range to levels higher than the upper reference limit, one of which had a 4-hr value that was within the reference or normal range (3.5-5 mEq/l). Overall, most samples were found to have reproducible levels of serum potassium. Serum potassium levels from stored whole blood collected in serum separator tubes are, for the most part, stable at room temperature for at least 4 hr prior to analysis. However, some samples can exhibit significant fluctuations of values. © 2015 Wiley Periodicals, Inc.

Manual centrifuge system: Bearing-based hand spinner made with 3-D printer.

Science.gov (United States)

Sun-Young Yoo; Seung Jae Lee; Jong-Mo Seo

2017-07-01

Compact disk (CD) Microfluidic platforms are being studied for medical applications such as blood tests. However, its size is bulky and electricity is needed to realize centrifuge force. In this paper, bearing-based hand spinner is designed using three-dimensional printer. This spinner does not need electricity and keeps rotating direction unlike paperfuge while it is spinning. The properties of spinner vary depending on bearing type which is positioned at the center. The type of weighting area also affects change in RPM over time. When a separation experiment is implemented, separating mixture into red ink and oil and whole blood into red blood cell and plasma are achieved properly with ceramic ball bearing.

Centrifuge Health Monitoring of the 50gTon beam centrifuge at the University of Sheffield