Efficacy comparison of gel-based, membrane and glass array techniques to detect human antibodies isotypes among the...

Hepatitis C virus is a major public health problem leading to cirrhosis and increased risk for development of hepatocellular carcinoma, both leading indications for liver transplantation. Egypt has the highest prevalence of hepatitis C of worldwide. Anti-HCV antibody is usually detected by enzyme immunoassay (EIA). Microarray analysis of 8 Anti-HCV antibody isotypes and 5 HCV peptides considered separately on 5 different matrixes was carried out on 50 hepatitis C Egyptian patients. The optimal substrate kind was chosen based on the greatest amount of antibody bonded with the minimal background. Mercaptosilane activated slides, agarose-slides and polyvinylidene difluoride membranes were the best substrates, while polyacrylamide and nitrocellulose membrane were less sensitive. IgM isotype gave the weakest signals in all patients whatever the substrate type used for antibody immobilization while IgG (total) and its subtypes IgG2, IgG3 and IgG(4) gave the strongest signals with most of the substrates follows by IgA(1), IgG(1) and total IgA, respectively. The results demonstrate that IgG2, IgG3 and IgG(4) are the dominant IgG subtypes, which may indicate that HCV patient immune response shift toward Th-2 immunity. The microarrays permitted the simultaneous serodetection of hepatitis C virus core and envelope peptides by using corresponding rabbit anti-peptides. Hepatitis C virus core and envelope peptides 1, 2, 3, 4 and 5 showed strong signals on all the used substrates except for polyacrylamide slides and nitrocellulose membranes.