Specific protein content of pools of plasma for fractionation from different sources: impact of frequency of donations.

BACKGROUND AND OBJECTIVES: Plasma pools for the production of human plasma medicinal products are distinguished according to the collection method (recovered or apheresis plasma) and the donor remuneration status. National regulations and the physical status of the donor determine the donation frequency and plasma volume per session. Relevant protein contents of different types of pools have not fully been compared. MATERIALS AND METHODS: We compared the levels of total protein, 15 main relevant plasma protein markers, and anti-B19 and anti-Streptococcus pneumoniae IgG in single-type pools of donations from different countries (Belgium, Finland, France, the Netherlands, Germany, United States). Both recovered plasma from non-remunerated donors and apheresis plasma from remunerated and non-remunerated donors were studied. RESULTS: Pools from paid US high-frequency, high-volume plasmapheresis donors showed significantly lower total protein (-9%), albumin (-15%), total IgG (-24%), IgM (-28%), hemopexin (-11%) and retinol-binding protein (-10%) but higher C1-inhibitor, pre-albumin and C-reactive protein contents than pools from unpaid European Union (EU) or US whole-blood or plasmapheresis donors. In contrast to pools from compensated EU plasmapheresis donors, pools from unpaid whole-blood or plasmapheresis donors showed no significant differences, whatever the collection method or country. Reductions in specific protein contents correlated well with protein half-life. CONCLUSION: These results should be taken into account with regard to donor health management and protein recovery.

Prevalence of human erythrovirus B19 DNA in healthy Belgian blood donors and correlation with specific antibodies against structural and non-structural viral proteins.

BACKGROUND AND OBJECTIVES: Human parvovirus (erythrovirus) B19 is recognized as a major contaminant of blood and blood products. To reduce the risk of contamination, plasma-pool screening and exclusion of highly viraemic donations are recommended. The objectives of this study were to estimate the prevalence of B19 DNA in our blood-donor population, to determine the appropriate pool size to be tested (taking into account parameters such as prevalence, viral load, test sensitivity, and the efficacy of inactivation procedures), and to correlate viral loads with the serological status of donors as regards antibodies against different viral proteins. MATERIALS AND METHODS: Pools of different sizes were tested for B19, using a sensitive nested polymerase chain reaction (PCR) as well as an simple, un-nested, less sensitive PCR. Positive pools were resolved to the level of individual donations, and the viral load and serological markers were determined. RESULTS: Of 16,859 donations, 27 (one of 625) were found to be B19 DNA positive, with viral loads ranging from 10(2) to > 10(7) IU/ml. Twenty-five of the positive donations were tested for VP-specific anti-B19 antibodies, and eight (32%) were negative for both immunoglobulin (Ig)M and IgG. They were probably collected in the preseroconversion window period or from chronic carriers without detectable antibodies. We regarded the seven (28%) IgM-positive donors as being in the early phase of infection. The remaining 10 (40%) IgM-negative, IgG-positive donors were probably carriers of persistent infection (i.e. PCR positive despite the presence of IgG antibodies), as suggested by their low viral loads (< 10(4) IU/ml). Fifteen out of 36 major pools contained one or more contaminated donations. Among these, 12 tested positive by nested PCR and only three by un-nested PCR, this reflecting a viral load of > 10(4) IU/ml. CONCLUSIONS: By testing all donations as pools of 480 by un-nested PCR, and resolving positive pools to identify the responsible donations, it is possible to ensure that the viral load in fractionation pools (5000 donations) remains < 10(3) IU/ml, compatible with the efficacy of inactivation procedures and complying with Food and Drug Administration (FDA) recommendations.

Polychlorinated biphenyls and organochlorine pesticides are eliminated from therapeutic Factor VIII and immunoglobulin concentrates and reduced in albumin by plasma fractionation.

BACKGROUND AND OBJECTIVES: Persistent organochlorine pollutants, such as polychlorinated biphenyls (PCBs) and organochlorine pesticides, are found in the general population and tend to accumulate in blood and tissues. Their distribution was examined in the starting plasma pools for fractionation, Cohn plasma fractions and therapeutic concentrates. MATERIALS AND METHODS: In each process fraction, total protein, cholesterol and triglycerides were measured as well as organochlorine pesticides and PCB congeners. RESULTS: Organochlorine compounds were undetectable in cryoprecipitate, Cohn fraction I, Factor VIII and immunoglobulin concentrates, and reduced in albumin preparations. CONCLUSION: Cohn plasma fractionation is very efficient for removing pollutants present in the starting material. Biological processing techniques should be analysed for their capacity to eliminate/reduce persistent organochlorine pollutants from the therapeutic proteins.

Mapping of natural anti-factor VIII antibodies in plasma pools from healthy donors: use of rationally designed synthetic peptides.

Inhibitor antibodies of blood coagulation factor VIII (FVIII) impair FVIII replacement therapy, constituting a serious complication in haemophilic patients. anti-FVIII antibodies may also develop in a variety of disease-associated autoimmunity. Mapping of human FVIII inhibitors in haemophilia A or autoantibody origin have delineated three major clusters of B-cell inhibitory epitopes (domain A2, A3 and C2). Inhibitory and non-inhibitory FVIII antibodies have also been described in plasma of healthy donors and pools of immunoglobulins. The purpose of this study was to use synthetic FVIII-peptides to more closely define regions of the molecule targeted by natural anti-FVIII antibodies. Predictive algorithms were used for defining the positions of potential continuous epitopes. To investigate the presence of peptide-reactive antibodies in normal plasma pools of healthy donors, a plasma fraction (Cohn fraction II+III) containing all IgG subclasses was purified by affinity chromatography on peptide-Sepharose columns. The results of ELISAs and Western blotting experiments (with the selected peptides and well-defined recombinant FVIII thrombin fragments) confirmed the reaction specificities of the affinity-purified human antibodies. For each IgG preparation, the isotopic subclass was also determined. In the clotting assay, several IgG preparations showed neutralising activity in a dose-dependent manner. Our observations support the recent hypothesis that FVIII inhibitors in haemophilia A and autoimmune disease may originate from the proliferation of natural FVIII-specific B-cell clones.

PCR detects HCV RNA in a plasma pool contaminated by a single preseroconversion donation of genotype 5a.

BACKGROUND AND OBJECTIVES: To determine the prevalence of HCV-RNA-positive plasma pools in Belgium, to validate our PCR method and to increase the safety of the released blood products. MATERIALS AND METHODS: Plasma pools consisting each of about 5,000 donations from Belgian unpaid volunteer blood donors were analysed by PCR for the presence of HCV RNA. Two different extraction methods were compared and validated. RESULTS: Two out of 367 plasma pools were found to be HCV RNA positive and were discarded. For one of these two pools, the look-back procedure identified an anti-HCV-negative contaminated donation. The HCV genotype of both the contaminated pool and the donation was 5a, a genotype rare in Europe. The viral load of the preseroconverted donation was 2.9 x 10(7) gEq/ml according to the bDNA method. CONCLUSION: In the case of plasma derivatives, various important steps are already included to increase safety. Nucleic acid testing of manufacturing plasma pools ensures that viral load in the starting material is as low as possible.