Karen M. Byrne

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

BackgroundThe 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.MethodsSerum, 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.ResultsA 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.ConclusionOur 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.

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.

Red blood cell preservation by droplet freezing with polyvinylpyrrolidone or sucrose-dextrose and by bulk freezing with glycerol.

BACKGROUND: Red blood cell (RBC) preservation is essential to transfusion medicine. Many blood group reference laboratories need a method to preserve rare blood samples for serologic testing at a later date. This study offers a comparison of three common cryoprotective agents and protocols used today: bulk preservation with glycerol and droplet freezing with sucrose‐dextrose (S+D) or polyvinylpyrrolidone (PVP).

The value of pH as a quality control indicator for apheresis platelets.

BACKGROUND: Standards and regulations require measurement of pH as an apheresis platelet (PLT) component quality monitor. The usefulness of this quality control (QC) measure was investigated.

Investigation of whether the acute hemolysis associated with Rho(D) immune globulin intravenous (human) administration for treatment of immune thrombocytopenic purpura is consistent with the acute hemolytic transfusion reaction model

BACKGROUND: Immune thrombocytopenic purpura and secondary thrombocytopenia patients treated with Rho(D) immune globulin intravenous (human; anti‐D IGIV) have experienced acute hemolysis, which is inconsistent with the typical presentation of extravascular hemolysis—the presumed mechanism of action of anti‐D IGIV. Although the mechanism of anti‐D‐IGIV–associated acute hemolysis has not been established, the onset, signs/symptoms, and complications appear consistent with the intravascular hemolysis of acute hemolytic transfusion reactions (AHTRs). In transfusion medicine, the red blood cell (RBC) antigen‐antibody incompatibility(‐ies) that precipitate AHTRs can be detected in vitro with compatibility testing. Under the premise that anti‐D‐IGIV–associated acute hemolysis results from RBC antigen‐antibody–mediated complement activation, this study evaluated whether the incompatibility(‐ies) could be detected in vitro with a hemolysin assay, which would support the AHTR model as the hemolytic mechanism.

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

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.

Increasing oxygen tension improves filtration of sickle trait donor blood

Summary. 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.

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.

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.

Analysis of the recovery of cryopreserved and thawed CD34+ and CD3+ cells collected for hematopoietic transplantation

Cryopreservation is often used to store cellular therapies, but little is known about how well CD3+ or CD34+ cells tolerate this process.