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    Definition of False-Positive Reactions in Screening For Hepatitis C Virus Antibodies

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    Faisal Jamshed
    The study analyzed 1,814 serum samples that tested positive for hepatitis C in an initial screening. 10% of samples were false positives as they were negative on a confirmatory test and PCR, and the patients did not develop hepatitis symptoms. A second screening more accurately identified true positives and negatives, showing the need for confirmation of initial positive hepatitis C screening results.

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    Definition of False-Positive Reactions in Screening For Hepatitis C Virus Antibodies

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    Faisal Jamshed
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    The study analyzed 1,814 serum samples that tested positive for hepatitis C in an initial screening. 10% of samples were false positives as they were negative on a confirmatory test and PCR, and the patients did not develop hepatitis symptoms. A second screening more accurately identified true positives and negatives, showing the need for confirmation of initial positive hepatitis C screening results.

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    07-10-2020

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    The study analyzed 1,814 serum samples that tested positive for hepatitis C in an initial screening. 10% of samples were false positives as they were negative on a confirmatory test and PCR, and the patients did not develop hepatitis symptoms. A second screening more accurately identified true positives and negatives, showing the need for confirmation of initial positive hepatitis C screening results.

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    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    16 views6 pages

    Definition of False-Positive Reactions in Screening For Hepatitis C Virus Antibodies

    Uploaded by

    Faisal Jamshed
    The study analyzed 1,814 serum samples that tested positive for hepatitis C in an initial screening. 10% of samples were false positives as they were negative on a confirmatory test and PCR, and the patients did not develop hepatitis symptoms. A second screening more accurately identified true positives and negatives, showing the need for confirmation of initial positive hepatitis C screening results.

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    of 6

    Definition of False-Positive Reactions in

    Screening for Hepatitis C Virus Antibodies


    ABSTRACT

    The rate of false-positive hepatitis C virus enzyme immunoassay results


    was determined to be at least 10% among 1,814 reactive serum samples
    based on (i) negative results in an independent confirmation assay, (ii)
    negative PCR results, and (iii) no patients developing clinical or
    biochemical signs of hepatitis during a 1-year follow-up.

    In daily laboratory routine, reliable diagnosis of hepatitis C virus (HCV)


    infection is not always possible by sole use of an HCV enzyme
    immunoassay (EIA), since it is well known that for a number of patients
    this assay produces false-positive results (4, 8, 17). Therefore, results
    obtained by EIA need to be confirmed by additional testing. However,
    the commercially available assay RIBA 2.0 (Chiron Corporation) does not
    fulfill the criteria defining a confirmation assay since it consists of
    recombinant proteins identical to those in the EIA (1, 5, 9). HCV PCR
    cannot be used for confirmation of positive EIA results, since a negative
    PCR result does not exclude the possibility of HCV infection with low-
    level viremia (below the limit of detection). Furthermore, PCR is too
    laborious and expensive to be used regularly as a confirmatory assay.
    Therefore, we have established an HCV strip immunoblot assay (SIA)
    (Universitäts-Krankenhaus Eppendorf [UKE] SIA) consisting of four
    recombinant proteins, derived from the core and three nonstructural
    regions (NS3, NS4, and NS5) of HCV, which are different from those used
    in the HCV EIA (5).
    In the present study we compared the results of a second-generation
    HCV EIA with those of the UKE SIA for 2,283 serum samples. The aim was
    to assess the significance of positive results in the HCV EIA to define
    criteria for the performance of further tests to reliably diagnose HCV
    infection in the daily laboratory routine. Sera were drawn from 2,283
    persons living in northern Germany around the city of Hamburg. They
    were sent to our laboratory under suspicion of HCV infection due to
    either elevated liver enzyme values (alanine aminotransferase, >45
    U/liter) or clinical signs of hepatitis (jaundice and upper abdominal pain)
    or risk factors for parenterally transmitted diseases, such as chronic
    hemodialysis, blood transfusion, or intravenous drug use. At the time of
    investigation they tested negative for acute infection with HAV (anti-HAV
    immunoglobulin M antibodies) and HBV (hepatitis B surface antigen).
    Repeated examinations were performed as follow-up every 3 months for
    1 year. For serological screening a second-generation HCV EIA (Abbott
    Laboratories, North Chicago, Ill.) was performed. For confirmation of
    HCV EIA results, sera were tested in parallel by the UKE SIA as previously
    described (5). The immunoblot assay was considered positive when
    antibodies to at least two different recombinant proteins were
    detectable. Reactivity against only a single protein was rated as an
    indeterminate result. For detection of HCV RNA reverse transcription-
    PCR was performed as previously described (6, 7).
    The HCV EIA was negative for 469 samples, of which 456 (97%) were also
    negative by UKE SIA. For 13 samples the UKE SIA was considered
    indeterminate. All 469 of these sera were negative by HCV PCR, and none
    of the patients developed clinical or biochemical signs of hepatitis during
    the follow-up.

    The HCV EIA was reactive for 1,814 samples, of which 1,394 (77%) were
    also positive by the UKE SIA (Table 1). However, in 240 cases (13%) the
    reactivity in the HCV EIA could not be confirmed by UKE SIA. Suitable
    specimens for HCV PCR were available for 193 of these 240 samples, and
    a positive PCR result was obtained with 13 samples. Of these, nine
    became positive by UKE SIA when retested after 3 months, which
    suggests that these patients had acquired HCV infection shortly prior to
    the first examination. In the remaining four patients, who repeatedly
    tested PCR positive despite a negative result by UKE SIA,
    immunosuppressing conditions could be found. One had a B-cell
    lymphoma, one was chronically hemodialyzed, and two practiced
    intravenous drug use. It has been shown earlier that in patients with
    immunosuppressive conditions, serological response is low or even
    absent (10, 14, 15). This could lead to negative or indeterminate results
    in serological assays although the individual suffers from infection with
    HCV (13). Therefore, for patients with known immunosuppressive
    disorders PCR should always be performed. The 180 initially PCR-
    negative subjects remained negative by UKE SIA and HCV PCR in
    repeated examinations during the follow-up. Moreover, these patients
    did not develop clinical or biochemical signs of hepatitis. This indicates
    that in at least these 180 samples (10%), false-positive results occurred.
    We must assume that the EIA was also false positive in the specimens for
    which no suitable material for PCR was available, since the UKE SIA
    remained negative and none of the patients developed clinical or
    biochemical signs of hepatitis during the follow-up. This indicates that as
    long as no better screening assays are commercially available every
    positive HCV EIA result must be confirmed.
    An indeterminate result in the UKE SIA was observed with 180 of the
    1,814 EIA-positive samples (10%). Suitable specimens for HCV PCR were
    obtained for 134 of these 180 samples, and HCV RNA could be detected
    in 58 of them. During the follow-up full seroconversion was observed in
    four patients. All of them initially revealed antibodies directed solely
    against the NS3 protein of the UKE SIA. In follow-up samples, reactivity
    against additional recombinant proteins emerged. These results support
    the previous assumption that antibody reactivity against NS3 plays an
    important role in the early serological detection of HCV infection (5).
    Furthermore, a particularly high correlation has been found between
    HCV viremia and antibody reactivity against the c33c antigen of the
    commercially available RIBA (2). Samples with a positive result by HCV
    EIA and an indeterminate result by immunoblot assay must be subjected
    to PCR, since we detected HCV RNA in 43% of samples (58 of 134). The
    percentage of indeterminate results by the UKE SIA is remarkably low
    compared to that by RIBA 2.0 or 3.0 (2, 3, 11, 16). One reason for this
    might be that local isolates were used to establish the UKE SIA, since
    serological tests containing recombinant proteins of local isolates have
    been shown to have better sensitivity and specificity than commercially
    available assays (5, 12). However, this is unlikely to be the only reason,
    since the UKE SIA was evaluated with serum samples containing a variety
    of HCV genotypes as previously described (5).
    The diagnosis HCV positive has a deep impact on the life of the afflicted
    person. Therefore, it must be reached as reliably as possible. Our data
    indicate that the widely used HCV EIA produces a high percentage (10%)
    of false-positive results. Compared to other screening assays, e.g.,
    human immunodeficiency virus EIAs, this is unacceptably high.
    Therefore, confirmation of every positive HCV EIA result by supplemental
    tests is mandatory. As we have shown with our in-house UKE SIA, one
    possibility for improving the reliability of HCV diagnosis is to introduce
    proteins into the confirmation assay which are different from those used
    in the screening assay.

    FOOTNOTES

     Received 21 January 1998.


     Returned for modification 6 May 1998.
     Accepted 13 October 1998.
     Copyright © 1999 American Society for Microbiology

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