C Vermylen

Evaluation of third-generation screening and confirmatory assays for HCV antibodies.

A third‐generation (gen.) screening and immunoblot assay (Ortho EIA‐3.0; Chiron RIBA‐3 prototype), using antigens derived from the capsid and different nonstructural regions (NS3, NS4 and NS5) of the hepatitis C virus viral genome, were evaluated in comparison with the corresponding second‐gen. assays (Ortho EIA‐2.0; revised Ortho EIA‐2.5; Chiron RIBA‐2). In 203 depository sera of blood donors, positive in EIA‐2.0, specificity of the screening assays was improved as shown by an increase in positive predictive value for viral carrier state from 0.23 (EIA‐2.0) to 0.37 (EIA‐2.5) and 0.52 (EIA‐3.0). Comparing the confirmation patterns on RIBA‐2 and RIBA‐3, this amelioration was mainly due to the specific elimination of false‐positive c22‐3 and c100‐3 reactions. Antibody response to the newly added NS5 antigen was not as prevalent as to the other antigens and had only a minor influence in sample allocation. In contrast, screening of 1,560 volunteer blood donors and 47 hemodialysis patients revealed 3 additional positive sera, only reacting with the NS5 antigen. However none of these isolated NS5 reactions could be confirmed on synthetic peptides [INNO‐LIA: NS5(p)] and none was PCR positive. A documented seroconversion, detected earlier with EIA‐3.0, was related to a better immunological response to the NS3 antigen and not to the additional NS5. From this pilot study third‐gen. assays appeared extremely useful in the reevaluation of HCV‐seropositive depository sera. However the additional value of the NS5 antigen in blood donor screening is still hypothetical and remains to be established in larger screening studies.

HEPATITIS-B IMMUNOGLOBULIN IN PREVENTION OF HBs ANTIGENÆMLA IN HÆMODIALYSIS PATIENTS

In a double-blind study, hepatitis-B immunoglobulin significantly protected patients in a haemodialysis unit against the development of HBs antigenaemia, compared to control patients receiving normal human immunoglobulin (p less than 0-01). Injections were given at the beginning and after 6 months, and observations extended over 16 months. Analysis of antiHBc and anti-HBs antibodies suggested that neutralisation of the virus inoculum, as well as modification of infection, may be implicated in the prevention of HBs antigenaemia.

Interferon production and natural killer (NK) activity in leukocyte cultures from multiple sclerosis patients

Peripheral blood leukocyte (PBL) cultures from only 37% of MS patients produced detectable HuIFN-gamma in response to ConA as opposed to 85% of the cultures derived from normal blood donors. However, the yields in patient-derived cultures that were responsive, were not lower than those in cultures from controls. Production of HuIFN-alpha after stimulation with Sendai virus was not aberrant in cells taken from MS patients. The difference in HuIFN-gamma response rate between MS and normal donor-derived cells was more pronounced when DR2+ carriers were compared amongst each other than when DR2-k carriers were compared. Among the MS patients, the failure of PBLs to produce HuIFN-gamma in response to ConA was not correlated with age, sex, disease duration and type of disease. However, positive correlations were found with current disability indices and past disease progression rates. Unstimulated NK-activities of MS patient-derived PBLs were not different from those of normal donor-derived cells. the degree of augmentation of the activity by stimulation with ConA and interferon-alpha was also normal. Within the MS patients group, but not in the control group, there was a trend for DR2+ carriers to have lower spontaneous and stimulated NK-activities than DR2- individuals.

Confirmation of hepatitis C virus positive blood donors by immunoblotting and polymerase chain reaction.

In a series of 385 sera obtained from volunteer blood donors positive for the first‐generation hepatitis C virus assay (Ortho), the viral genome was detected by polymerase chain reaction (PCR) in 89 sera (23%). Most PCR‐positive sera were found positive with the c100‐3 neutralisation assay (Abbott) and by two second‐generation enzyme immunoassays (Abbott, Ortho). However overall specificity of these assays was rather low. By immunoblotting (Innogenetics and Chiron/Ortho) the specificity could be considerably improved and the best correlation with carrier state was obtained when analysing the results for lane‐specific reaction: all 89 viral carriers and only 9 other donors had antibodies against structural ‘core’ epitopes. From the present data we can conclude that in screening a volunteer blood donor population the confirmation of antibodies against ‘core’ epitopes by immunoblotting is strongly associated with viral carriage.

Immunoglobulin G antibodies to lymphadenopathy-associated virus in differently treated French and Belgian hemophiliacs.

Immunoglobulin G antibodies to lymphadenopathy-associated virus have been detected in two groups of French hemophiliacs and in one group of Belgian hemophiliacs, whose mode of treatment differed. Seropositivity was more frequent (58.9%) in patients heavily transfused with blood products of French and foreign origin than in less frequently transfused persons (10.3%). The Belgian group, treated only with local products, showed the lowest frequency of seropositivity (3.4%). In healthy French controls, 1 of 330 had antibody to the virus. The results indicate transmission of lymphadenopathy-associated virus via blood-derived products.

Activation of natural cytotoxicity of human peripheral blood mononuclear cells by interferon: a kinetic study and comparison of different interferon types

Summary. Overnight incubation of human peripheral blood mononuclear (PBMN) cells with leucocyte interferon (leucocyte IFN) resulted in a 2–5‐fold increase in natural cytotoxicity (NC) against the erythroleukaemic cell line K562. Fibroblast IFN, purified to homogeneity by zinc‐chelate chromatography, stimulated NC to the same extent, while partially purified immune IFN was about twice as active. Upon gel filtration of immune IFN, NC stimulating and antiviral activity co‐eluted. Treatment of PBMN cells with ammonium chloride buffer (AmCl) abrogated NC nearly completely. Incubation of AmCl‐treated cells with leucocyte IFN resulted in a partial regeneration of NC. Kinetic studies revealed that this regeneration required only a short exposure to IFN followed by a longer incubation period. The data are interpreted as indicating that in the process of NC activation IFN mainly acts as a trigger for precursor cells to mature into cells with NC.

A Simple Method for the Assay of Factor VIII, Using 20 Microlitres of Capillary Blood

Summary A simple and sensitive method for Factor‐VIII assay is presented which permits a reproducible quantitative assay on 20μl. capillary blood.

Hepatitis C virus confirmation in blood donor screening.

A combination of different enzyme immunoassays (EIAs) was used for the serological confirmation of sera that were positive in a hepatitis C virus (HCV) second‐generation screening EIA. Different reaction patterns were related with the probability of the HCV‐carrier state as determined by polymerase chain reaction (PCR). Five hundred and eight sera of volunteer blood donors were send for confirmation and at first reexamined with both Abbott and Ortho second‐generation screening EIA. A group of 195 sera, positive in both assays, was further evaluated by the Abbott Supplemental Assay, the Monolisa anti‐HCV and an EIA with only the amino terminal part of the nucleocapsid protein as antigen. In addition PCR on the 5′‐noncoding region of the viral genome was performed. We observed that 75 of the 78 PCR‐positive sera were found in a group of 89 sera that were strongly positive in the four EIAs used. Moreover all but 1 PCR‐positive sera were reactive against the nucleocapsid protein of the virus. Hence we concluded that a genuine antibody response to the nucleocapsid protein is highly suggestive for the HCV‐carrier state.

Ferritin-associated iron induces neutrophil dysfunction in hemosiderosis.

Neutrophils (PMNs) from patients with secondary iron overload have an increased iron and ferritin content as well as a phagocytosis defect. Several serum components might be incriminated in the cellular iron accumulation. We therefore compared the effects on the PMN phagocytosis of total serum as well as the ferritin and transferrin fractions of serum derived from patients with thalassemia major and healthy control subjects. An incubation system of PMNs was developed. PMN phagocytosis was measured before and after incubation. Total serum from patients with thalassemia induced a defect that was prevented by co-incubation with deferoxamine (DFO). Gel-filtration chromatography was performed to separate the serum fraction containing transferrin and albumin from that containing ferritin. The transferrin-albumin fraction had no effect on PMN phagocytosis. On the contrary, the ferritin fraction of normal serum was deleterious to PMN phagocytosis, and the same fraction from thalassemic serum decreased PMN phagocytosis even more. Co-incubation with DFO or catalase improved this defect. Moreover, a cellular increase in the L-type subunit of ferritin was observed after the incubation of PMNs with the ferritin-containing fraction from thalassemic serum. In conclusion, serum from patients with thalassemia is toxic to PMNs, and this toxicity is due to ferritin-associated iron.

Interferon therapy in multiple myeloma: Failure of human fibroblast interferon administration to affect the course of light chain disease

Abstract Fibroblast interferon at a dosage of 28 × 10 6 U/wk failed to influence disease progression in a preterminal case of therapy-resistant light chain myeloma. In a second case, that had not previously been treated, a first course of fibroblast interferon ( 30 × 10 6 U/wk ) associated with corticosteroids remained without effect. In this patient subsequent leukocyte interferon treatment was associated with a decrease in urinary light chain excretion and normalization of calcaemia, all other parameters remaining unaltered. A third patient with light chain disease was resistant to chemotherapy ab origine . None of the disease parameters responded to either fibroblast or leukocyte interferon therapy ( 21 × 10 6 U/wk ). During therapy a downward trend occurred in the relative number of lymphocytes characterizable as B and T cells. Mitogenic reactivity remained unchanged except for a downward trend, during fibroblast interferon therapy, in reactivity to PHA and Con A after 6 days culturing. Spontaneous (background) mitogenesis upon culture showed an upward trend.