Of 103,203 donations collected in Scotland and Northern Ireland over a 3-month period and screened for HCV antibody by Ortho or Abbott second-generation ELISAs, 340 were found repeatedly reactive. Supplementary testing with RIBA-2 resulted in 77 being classified as positive, 130 as indeterminate, and 133 as negative. PCR analysis of the positives and indeterminates indicated viraemia in 65 (84%) of the positives and 7 (5.5%) of the indeterminates. To determine if PCR analysis could be eliminated or reduced by further serological testing, all RIBA-2 positives and indeterminates were tested by UBI and Wellcozyme ELISAs and Innolia and RIBA-3 immunoblots. All RIBA-2 positives with bands to more than 1 gene product were detected in all 4 systems, but > 60% of RIBA-2 indeterminates were negative in those tests that contain either recombinant antigens or synthetic peptides derived independently from those used by Ortho/Abbott tests. A comparison of data from the 79 reactive with the core (c22) region revealed only 16 samples reactive in all 4 systems as well as Ortho and Abbott. These 16 included all 6 of the PCR positives in the 79 c22 indeterminate samples. ELISAs and immunoblots using independently derived antigens can offer a useful method of screening out nonspecific reactions in Ortho or Abbott ELISAs, hence reducing the need for PCR testing. Some caution is required as all such tests do not contain identical mixes of antigenic material.