Hepatitis C Virus Antibodies Cut-Off ELISA 96 Wells Dialab

Hepatitis C virus (HCV) is an enveloped, single-stranded positive sense RNA virus belonging to the Flaviviridae family. Six major genotypes and series of subtypes of HCV have been identified. Isolated in 1989, HCV is now recognised as the major cause for transfusion associated non-A, non-B hepatitis. The disease is characterised with acute and chronic forms, although more than 50% of infected individuals develop severe, life threatening chronic hepatitis with liver cirrhosis and hepatocellular carcinomas. Since the introduction in 1990 of anti-HCV screening of blood donations, the incidence of this infection in transfusion recipients has been significantly reduced. This is a third-generation assay including antigens from the Core (c22), NS3 (c200), NS4(c200) and NS5 regions of the genome. The improved sensitivity of third-generation assays enables detection of HCV antibodies 60 days after infection.

This assay is intended for the qualitative determination of antibodies against HCV in human serum or plasma. The test wells are coated with recombinant HCV antigens expressed in E.coli (recombinant Core and NS3/4/5). During the first incubation step patient sample and Biotin-conjugated HCV antigen are added to the well. If HCV antibodies are present in the patient sample they will be captured in the well as a double antigen sandwich. Free and bound are separated by washing and then horseradish peroxidase (HRP) conjugated anti-human IgG which bind the immobilised antigen- antibody complex. Unbound conjugate is removed by washing and then enzyme substrate added which in the presence of the conjugate hydrolyses to form a blue colour. Addition of stop solution forms a yellow end-product which is photometrically measured at 450nm. The amount of colour is proportional to the concentration of antigen in the patient sample. After validation checks a Cut-Off Value is calculated.
Negative: < 1 Cut-off Value
Positive: ≥ 1 Cut-off Value
Borderline 0.9 - 1.1 Cut-off Value (Re-test to confirm)

Antigen coated Microtiter wells : 12x8 well strips
Positive Control: 1ml
Negative Control: 1ml
Enzyme Conjugate : 12ml
Biotin Conjugate :6ml
Substrate Solution A : 1x6ml
Substrate Solution B : 1x6ml
Stop Solution: 1x6ml
Wash buffer (20X); 1x50ml
Sample volume: 50µl human serum or plasma
Incubation time: 2h (60 min, 30min, 30min all at 37°C)
Store at 2-8°C.