Heterophilic antibodies in human sera causing falsely increased results in the CA 125 immunofluorometric assay

Abstract

An in-house OC 125 monoclonal antibody-based "sandwich" immunofluorometric assay (IFMA) described previously (Clin Chem 1987; 33:2191-4) gave higher results for CA 125 in 37 of 123 serum samples than did a commercially available immunoradiometric assay (IRMA). Discordant results between the two assays became concordant when measurements of the samples were repeated with normal mouse serum (100 mL/L) included in the IFMA reagent. The murine immunoglobulins are thought to block the ability of the heterophilic antibodies in the human serum samples to cross-link the labeled antibody with the solid-phase antibody. Using an enzyme immunoassay, we demonstrated human anti-mouse antibodies (HAMA) in most of the discordant samples examined. We tested the ability of nonimmune sera from other animal species to lower the apparent CA 125 concentrations of the spurious samples and observed that rat, goat, and sheep serum were less effective than mouse serum. One serum sample was discovered to give a falsely increased CA 125 result with the IRMA, but this increase could be prevented by adding murine serum to the IRMA reagent. We conclude that falsely increased CA 125 results are best prevented by adding murine serum (or murine antibodies) to the assay buffer.

Development of a Human Gamma Interferon Enzyme Immunoassay and Comparison with Tuberculin Skin Testing for Detection ofMycobacterium tuberculosisInfection

“…Specificity for IFN-␥. To assess the possibility that heterophile antibodies (4,5,13,16) caused interference in the IFN-␥ assay, unstimulated human plasma samples from 201 healthy blood donations were tested in sample diluent without NMS. Of these plasma samples, 6% were highly reactive, generating an optical density in the EIA greater than twice that of the negative control (data not shown).…”

“…It is well-established that two-site sandwich EIAs which employ MAbs as both the solid-phase capture antibody and the HRP-conjugated antibody often suffer from false-positive problems due to the presence of heterophile antibodies in many serum-plasma samples (4,5,13,16). To circumvent this problem, we followed the procedure described by Jones et al (13) and coated microtiter plates with F(abЈ) 2 fragments of the solid-phase capture antibody and added 20% NMS to the sample diluent.…”

Unusual results from immunoassays and the role of the clinical endocrinologist

“…Some antibodies that interfere with immunoassays are highly specific and produced in response to immunizations. These are specific human anti‐animal antibodies (HAAAs) that, most commonly, interfere with sandwich immunoassays by linking the capture to the detection antigen, giving a false positive result 78–84 . Kaplan and Levinson 85 suggest a distinction is made between these different forms of endogenous antibody interference as follows:…”

Multiplex Analysis of Heterophil Antibodies in Patients With Indeterminate HIV Immunoassay Results

“…Heterophil antibodies are common in the general population [14][15][16] and frequently cause interference and false-positive results in many types of immunoassays. [2][3][4][5]7,[17][18][19] Heterophil antibodies have not previously been shown definitively to interfere in HIV immunoassays. 6 With a unique multiplexed immunoassay, we have demonstrated that heterophil antibodies can cause false-positive HIV ELISA results in cases in which there are no known risk factors and clinical suspicion is low.…”

“…1 Heterophil antibodies can create a false-positive result in an immunoassay by binding to the capture and signal antibodies in the assay and mimicking the behavior of the analyte of interest. Heterophil interference has been observed in many immunoassays, including creatine kinase MB, 2 human chorionic gonadotropin, 3 CA-125, 4 thyroid stimulating hormone, 5 and others, but has not been well characterized in HIV immunoassays. 6 False-positive HIV enzyme-linked immunosorbent assay (ELISA) results can occur with serious medical, social, and legal consequences, 7 and false-positive results are thought to account for as many as 4.6% of positive Western blots in a low-risk screening setting.…”