Relevance of RIBA-3 supplementary test to HCV PCR positivity and genotypes for HCV confirmation of blood donors

Abstract

HCV antibody screening of 624,910 blood donations resulted in 3,832 samples being referred for confirmation. All were tested by RIBA-3 with 2,710 negative, 945 indeterminate and 177 positive results. HCV RNA was detected by PCR in an average of 69.5% of RIBA-3 positives (4 bands 84.1%; 3 bands 74.1%; 2 bands 34.1%) and only 0.53% of RIBA-3 indeterminates. Eighty-four percent of samples with a total RIBA-3 band intensity score (maximum 16) of > or = 8 were PCR positive compared with only 22% of those with a score of < 8. Total mean band intensities for HCV genotype 1 samples (n = 65) were 13.2, genotype 2 (n = 17) 11.4 and genotype 3 (n = 65) 11.2 with type 1 samples showing greater reactivity with c100 and c33 antibodies. No PCR positive type 1 samples were found with RIBA-3 total band scores less than 8, no PCR positive type 2 samples less than 6, whilst PCR positive type 3 samples were found with scores as low as 2. NS5 indeterminates were the most common (40.2%) single band pattern but yielded no PCR positive samples, followed by c33 (23.3%) with one PCR positive and c100 (20.2%) with one PCR positive whilst c22 indeterminates were least common (16.3%) but included three PCR positive donors. All five RIBA-3 indeterminate PCR positive donors were type 3.

Antigenic Heterogeneity of the Hepatitis C Virus NS5A Protein

“…Different studies performed with these assays have shown that the NS5 antigen is not as immunoreactive as the other antigens (4,15) and is potentially less specific than any of the other HCV antigens included in the third generation of commercially available diagnostic assays (11).…”

“…Regions of nonspecific antigenic reactivity can be found within different HCV proteins, including the NS5 protein (11). One study showed that more than 30% of the 543 synthetic peptides derived from different regions of the HCV polyproteins of different genotypes were recognized by antibodies from healthy blood donors (24).…”

“…A recently released third generation of EIA (EIA-3) includes the NS5 protein of 942 aa. EIA-3 is slightly more sensitive than EIA-2, but most of the increased sensitivity appears to be attributable to improvements in detecting anti-C33 (4,11,27). The detection rate of anti-NS5 (59.3%) is much lower than the detection rates of anti-C22 (92.1%) and C33 (88.1%) (11).…”

“…EIA-3 is slightly more sensitive than EIA-2, but most of the increased sensitivity appears to be attributable to improvements in detecting anti-C33 (4,11,27). The detection rate of anti-NS5 (59.3%) is much lower than the detection rates of anti-C22 (92.1%) and C33 (88.1%) (11). Commercial HCV tests based upon proteins or peptides derived solely from HCV genotype 1 may be less effective for the detection of antibody against HCV of other genotypes (8,17,24).…”

High Diagnostic Accuracy of Antigen Microarray for Sensitive Detection of Hepatitis C Virus Infection

“…Because comparing an HCV RNA-based PCR assay with an anti-HCV-based assay is not practical, we cannot directly compare PCR-based results with those obtained using our protein microarray assays. However, the overall accuracy of our protein microarray method is nearly equivalent or better than that of the LIBA-blot based confirmatory test (19 ). The inclusion of additional antigens such as NS4 would improve the specificity of this protein microarray-based assay.…”