Development and applications of sensitive enzyme immunoassay for antibodies: A review

Abstract

The sensitivity of the conventional enzyme immunoassay methods is seriously limited by the presence of nonspecific immunoglobulins at high concentrations in test serum. In order to overcome this difficulty, various methods have been developed, and the detection limit of serum antibodies has been lowered to 50-100 ng/l, which is 300,000-fold lower than that by the conventional enzyme immunoassay method using antigen-coated solid phases and anti-immunoglobulin antibody-enzyme conjugates. As a result, anti-insulin antibodies have been demonstrated in most of patients treated with insulin for 0.5-10 months, and anti-thyroglobulin IgG has been demonstrated not only in all patients with Graves' disease and chronic thyroiditis, but also in a large proportion of healthy subjects.

Potential of the immune complex transfer enzyme immunoassay for antigens and antibodies to improve the sensitivity and its limitations

“…The initially developed immune complex transfer enzyme immunoassay was modified in various ways (2)(3)(4)(5)(6). The most sensitive immune complex transfer enzyme immunoassay for antibodies among those developed so far is shown in Fig.…”

“…The immune complex transfer enzyme immunoassay was developed initially for antibody IgG to minimize the nonspecific signal in the conventional ELISA, in which antigencoated solid phase is reacted with antibody IgG to be measured in serum samples and, after washing, with (anti-IgG) antibody-enzyme conjugate (1)(2)(3)(4)(5)(6). In the immune complex transfer enzyme immunoassay initially developed, antibody IgG to be measured in serum samples was reacted with 2,4-dinitrophenyl-biotinyl-antigen, and the immune complex formed was trapped onto colored polystyrene beads coated with (anti-2,4-dinitrophenyl group) IgG.…”

“…However, the sensitivity of noncompetitive immunoassays using labels is limited by the nonspecific signal, that is, the signal in the absence of antigens and antibodies to be measured. A decade ago, a method to markedly reduce the nonspecific signal in a conventional ELISA for antibody IgG was developed (1), and ultrasensitive enzyme immunoassays using this method, namely, the immune complex transfer enzyme immunoassays, have been developed for various antigens and antibodies (2)(3)(4)(5)(6)(7). This article describes the potential of the immune complex transfer enzyme immunoassay to improve the sensitivity and its limitations, using HIV-1 p24 antigen and antibody IgGs to HIV-1 p17 and reverse transcriptase as examples.…”

Assessment of Mucosal Immune Response in Genitourinary Tract Using Urine

“…An immune-complex transfer enzyme immunoassay was developed by us to measure antibodies against infectious microorganisms, autoantigens, and so on with high sensitivity (8), and was demonstrated to be 100 to 300,000-fold more sensitive than ELISA (8). To exactly compare the sensitivity of this assay with that of ELISA for anti-OVA IgA, affinity-purified anti-OVA antibodies were diluted to various extents with pooled urine from naive rats, and were subjected to the two assays.…”

Simultaneous quantitative determination of multiple analytes with fluorescence‐tagged probes by Immunochromatography

“…Since their first introduction in 1950s, immunoassays have become indispensable tools and have achieved great importance in the field of scientific and medical research (Ishikawa and Kohno, 1989;Oellerich, 1984). Immunoassay is rapidly evolving toward more simple approaches for the quick identification and diagnosis of biological specimens (Westermann et al, 2002).…”