Biomedical subjects

Karen M Byrne

Publications and source records attributed to Karen M Byrne.

8 recordsLinked to original sources

PubMed2006

Comparison of proteomic profiles of serum, plasma, and modified media supplements used for cell culture and expansion.

BACKGROUND: The culture and expansion of human cells for clinical use requires the presence of human serum or plasma in culture media. Although these supplements have been extensively characterized in their chemical composition, only recently it has been possible to provide by high throughput protein analysis, a comprehensive profile of the soluble factors contributing to cell survival. This study analyzed and compared the presence of 100 proteins including chemokines, cytokines and soluble factors in six different types of media supplements: serum, plasma, recalcified plasma, heat inactivated serum, heat inactivated plasma and heat inactivated recalcified plasma. METHODS: Serum, plasma, recalcified plasma, and heat inactivated supplements were prepared from ten healthy subjects. The levels of 100 soluble factors were measured in each sample using a multiplexed ELISA assay and compared by Eisen hierarchical clustering analysis. RESULTS: A comparison of serum and plasma levels of soluble factors found that 2 were greater in plasma but 18 factors were greater in serum including 11 chemokines. The levels of only four factors differed between recalcified plasma and plasma. Heat inactivation had the greatest effect on soluble factors. Supervised Eisen hierarchical clustering indicated that the differences between heat inactivated supplements and those that were not were greater than the differences within these two groups. The levels of 36 factors differed between heat inactivated plasma and plasma. Thirty one of these factors had a lower concentration in heat inactivated plasma including 12 chemokines, 4 growth factors, 4 matrix metalloproteases, and 3 adhesion molecules. Heat inactivated decalcified plasma is often used in place of heat inactivated serum and the levels of 19 soluble factors differed between these two supplements. CONCLUSION: Our report provides a comprehensive protein profile of serum, plasma recalcified plasma, and heat inactivated supplements. This profile represents a qualitative and quantitative database that can aid in the selection of the appropriate blood derived supplement for human cell cultures with special requirements.

Journal Article↗

PubMed2006

In vitro variables of red blood cell components collected by apheresis and frozen 6 and 14 days after collection.

BACKGROUND: An automated cell processing system (ACP 215, Haemonetics Corp.) can be used for the glycerolization and deglycerolization of RBC components, but the components must be 6 or fewer days old. Depending on the anticoagulant (CP2D)/additive solution (AS) used, deglycerolized RBCs can be stored at 1 to 6 degrees C for up to 14 days. This study evaluated in vitro variables of apheresis RBC stored for 6 and 14 days at 1 to 6 degrees C before glycerolization and 14 days after deglycerolization. STUDY DESIGN AND METHODS: Two units of CP2D/AS-3 leukoreduced RBCs were collected by apheresis from seven donors. One unit was glycerolized and frozen 6 days and the other 14 days after collection. All units were deglycerolized with the ACP 215 and stored at 1 to 6 degrees C for 14 days in AS-3. Several in vitro variables were evaluated during postdeglycerolization storage. RESULTS: All components had postdeglycerolization RBC recoveries greater than 81 percent and osmolalities of less than 400 mOsm per kg. No significant differences were noted in potassium and supernatant hemoglobin after 14 days of postdeglycerolization storage between RBCs frozen at 6 and 14 days after collection. After 14 days of postdeglycerolization storage, however, the pH, lactate, and ATP levels were slightly lower in RBCs frozen after 14 days. CONCLUSION: The ACP 215 can be used to glycerolize and deglycerolize apheresis RBC components that are up to 14 days of age. It is likely that apheresis components glycerolized at 14 days of age or less can be stored up to 14 days in AS-3 after deglycerolization, but this should be confirmed with in vivo survival studies.

Adenosine Triphosphate↗

PubMed2006

KEL6 and KEL7 genotyping with sequence-specific primers.

BACKGROUND: Although antibodies to Js(a) and Js(b) are clinically significant, reagent-quality anti-Js(a) and anti-Js(b) are not readily available. A sequence-specific primer-polymerase chain reaction (SSP-PCR) genotyping assay was tested that makes use of two single-nucleotide polymorphisms (SNPs) at positions 1910 and 2019 of KEL. These SNPs distinguish the gene encoding Js(a), KEL6; and Js(b), KEL7. STUDY DESIGN AND METHODS: Four primer sets that selectively amplified KEL6 and KEL7 from genomic DNA were developed. Two sets detected the SNP at bp 1910 and two sets detected the bp 2019 SNP. KEL6 and KEL7 genotyping and Js(a) and Js(b) phenotyping results were compared among 64 subjects. RESULTS: The SSP-PCRs were specific for KEL6 and KEL7 when testing DNA for three donors of known Js phenotype: Js(a+b-), Js(a-b+), and Js(a+b+). Genotyping results for the 1910 SNP were identical to the phenotyping results in all 64 subjects, but for the 2019 SNP, the genotyping and phenotyping results were identical for only 49 subjects. In 12 subjects with the Js(a-b+) phenotype, the 2019 SNP was heterozygous KEL6, KEL7; in 2 with Js(a-b+) and in 1 with Js(a+b+), the 2019 SNP was homozygous KEL6. CONCLUSION: KEL 2019-bp SNP does not always correlate with the Js phenotype owing to the presence of an atypical KEL gene with a KEL7 polymorphism at 1910 and a KEL6 polymorphism at 2019. The KEL polymorphism at 2019 is silent and this allele yields a Js(a-b+) phenotype. Only analysis of the 1910-bp SNP can be used to genotype KEL6 and KEL7.

Black or African American↗

PubMed2005

Factors affecting the formation of white particulate matter in red blood cell components.

BACKGROUND: Recently white particulate matter (WPM) in red blood cell (RBC) components has received increased attention. The nature and causes of WPM formation were investigated. STUDY DESIGN AND METHODS: Whole-blood units were collected from 18 healthy subjects with three different types of collection sets. Six units were collected into each type. Units were divided into four equal parts and stored for 4 hours: two parts at room temperature and two at 4 degrees C. RBCs were prepared from each quarter-unit: two by heavy centrifugation (5000 x g) and two by light centrifugation (2000 x g). Whole blood was inspected for WPM over 4 hours and RBCs over 1 hour. RESULTS: No WPM was detected in whole blood, but WPM was detected in at least one RBC component from 9 of the 18 donations. The 36 components prepared by heavy centrifugation were more likely to contain WPM than the 36 prepared by light centrifugation (50% vs. 19%; p < 0.02). The incidence of WPM was similar among RBCs stored at room temperature and 4 degrees C. Donors of RBCs with WPM had higher total cholesterol levels than donors of components without WPM (191 +/- 20 mg/dL vs. 163 +/- 32 mg/dL; p < 0.04), but there was no difference in triglyceride levels between the two groups. CONCLUSIONS: WPM is an expected consequence of standard RBC manufacturing methods, but it is more frequent in RBCs prepared by heavy centrifugation and from donors with higher cholesterol levels.

Adult↗

PubMed2004

Increasing hemoglobin oxygen saturation levels in sickle trait donor whole blood prevents hemoglobin S polymerization and allows effective white blood cell reduction by filtration.

UNLABELLED: BACKGROUND Red blood cell (RBC) components from donors with sickle cell trait (Hb AS) often occlude white blood cell (WBC) reduction filters. Techniques were investigated to successfully filter Hb AS donor blood by increasing the Hb oxygen saturation with storage bags and conditions suitable for transfusion products. STUDY DESIGN AND METHODS: Oxygenation kinetics were measured over 3 days in whole-blood units stored in standard-sized 600-mL polyvinylchloride (PVC) bags and whole-blood units divided into three equal parts and stored in standard-sized blood bags made from PVC, tri-2-(ethylhexyl)trimellitate (CLX) plastic, or Teflon. The filterability of Hb AS blood stored for 3 days was tested with whole-blood filters. RESULTS: Oxygen saturation levels did not increase in full whole-blood units from donors without sickle cell trait during 3 days of storage in 600-mL PVC bags. In divided Hb AS whole-blood units stored for 3 days, oxygen saturation levels increased from baseline levels of 45 to 56, 66, and 94 percent after storage in 600-mL PVC, CLX, and Teflon bags, respectively (n = 5, p < 0.02), and all components filtered completely. When full Hb AS whole-blood units from eight donors were stored for 3 days in 1.5-L CLX bags, all units filtered completely, but one had a high residual WBC count. CONCLUSION: Storage of Hb AS whole blood in large-capacity oxygen-permeable bags increases oxygen tension and allows more effective WBC reduction by filtration.

Benzoates↗

PubMed2003

Increasing oxygen tension improves filtration of sickle trait donor blood.

A major cause of filter failure of red cell (RBC) components from donors with sickle cell trait (HbAS) is the polymerization of haemoglobin. The oxygen saturation (sO2) of blood stored in various plastics and different volumes of air was assessed. Blood from 10 HbAS donors was collected and divided into two bags, one with air added, one without. Bags with added air had increased sO2 levels (from 49 +/- 10% to 76 +/- 6%). Filtration was successful for nine of 10 components with air, and one of 10 without air. Successful filtration of RBC components occurs when sO2 is increased.

Adolescent↗

PubMed2002

Detection of antibodies in acid eluates with the gel microcolumn assay.

BACKGROUND: Gel microcolumns can be used to detect unexpected serum antibodies and to determine ABO blood group and Rh phenotype. DATs can also be performed with this system. The purpose of this study was to compare the gel microcolumn to the tube IAT using anti-IgG for the detection of antibodies eluted from RBCs. STUDY DESIGN AND METHODS: Acid eluates were prepared from 30 peripheral blood and 41 umbilical cord blood samples. Twelve of the 71 eluates made were from control samples (known DAT negative). Specificities of eluted antibodies were determined by both tube and gel assays with a three-cell screen plus A1 and B cells, as determined by blood type. RESULTS: Ten of 30 peripheral blood eluates were reactive in both assays. Eighteen were nonreactive in both assays, and two from patients with autoimmune hemolytic anemia were reactive by gel assays and nonreactive by tube assays. Thirty three of the 41 cord blood eluates were reactive in both assays. Eluates from 2 of the 35 DAT-positive samples reacted with A1 and B cells by the tube method but were nonreactive by the gel method. Of the 33 cord blood eluates that were reactive by both assays, antibody specificity differed for two samples. When tested by tube assay, these eluates reacted with both A1 and B cells, whereas the same eluates tested by gel assay showed one reacting with only A1 cells and the other with only B cells. CONCLUSIONS: Results of testing eluates in gel assays were similar to those obtained in tube assays. The gel assays may be better at detecting antibodies eluted from RBCs from patients with autoimmune hemolytic anemia, and tube assays may be better at detecting isohemagglutinins eluted from umbilical cord blood.

Adult↗

PubMed2002

Sickle Hb polymerization in RBC components from donors with sickle cell trait prevents effective WBC reduction by filtration.

BACKGROUND: RBC components collected from donors with sickle cell trait frequently occlude WBC-reduction filters. In vitro, sickle trait RBCs have the potential for sickle Hb (Hb S) polymerization at low oxygen saturations and high Hb concentrations. STUDY DESIGN AND METHOD: To determine if the low pH and high osmolarity of the CP2D used in the collection contributed to filter failures, the filterability of sickle trait donor RBCs collected in CP2D was compared with RBCs from the same donors collected in heparin. RESULTS: Five of six sickle trait components collected in CP2D did not complete filtration, but all six RBC components collected in heparin filtered completely. RBC components collected in CP2D from four other sickle trait donors were divided in two, and one-half was treated with carbon monoxide to convert Hb S to its liganded form to prevent Hb S polymerization. All four carbon monoxide-treated components filtered within 9 minutes, but only one untreated component filtered completely. RBC components collected by apheresis contained less CP2D, and five of seven sickle trait apheresis components filtered completely; four of the five filtered rapidly (<15 min) and one filtered in 100 minutes. Hb oxygen saturation was greater in the four rapidly filtering apheresis RBC components (68 +/- 9%) than in the three that filtered slowly or incompletely (37 +/- 5%, p = 0.03). CONCLUSIONS: Hb S polymerization appears responsible for RBC WBC-reduction filter failures. Citrate anticoagulant and low oxygen saturation are responsible in part for Hb S polymerization in this setting.

Anticoagulants↗