Chemiluminescent enzyme immunoassay for alpha‐fetoprotein using β‐D‐galactosidase as label and 5‐bromo‐4‐chloro‐3‐indolyl‐β‐D‐galactopyranoside as substrate

Abstract

We developed a sensitive chemiluminescent sandwich-type enzyme immunoassay (CL-EIA) of alpha-fetoprotein (AFP) using beta-D-galactosidase (beta-gal) as a label and 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside as a substrate. The CL-EIA for AFP was performed using two monoclonal antibodies, one antibody is labelled with beta-gal, the other is coated onto the inside surface of a polystyrene tube. The detection limit for AFP was 0.5 ng/mL, equivalent to 10 pg/assay tube. The coefficient of variation for within and between assay imprecision were 2.0%-4.9% (n = 10) and 4.4%-9.8% (n = 5), respectively. AFP values in serum determined by this method correlated well with those obtained by radioimmunoassay (n = 26, r = 0.99). This sensitive AFP assay can be performed within 4 h and can be used as a routine assay in clinical diagnosis.

Development of functionalized terbium fluorescent nanoparticles for antibody labeling and time-resolved fluoroimmunoassay application

“…As a new fluorescence reagent, the advantages of the nanoparticles including smaller size, high hydrophilicity and biocompatibility, strong fluorescence, long fluorescence lifetime, and direct availability for biolabeling, make them particularly useful as a fluorescence probe for time-resolved fluorescence bioassay. The results of TR-FIA for AFP using the nanoparticle-labeled antibody show that the method is simple and has enough sensitivity [31,32] for determination of AFP in human serum samples with good precision and accuracy. It can be expected that the new nanoparticle fluorescence probe would be widely useful for highly sensitive clinical analysis, time-resolved fluorescence bioimaging technology, and other biotechnology applications.…”

Development of Highly Sensitive Analytical Methods for Biologically Relevant Materials and Their Pharmaceutical Applications

“…The sensitivity of the CL reaction was improved by adding an enhancer to the luminescence method, and the resulting improved method is used globally. 4) In 1991, we developed a highly sensitive assay for alkaline phosphatase (ALP) and β-Dgalactosidase (β-gal) using indoxyl derivatives as substrates [5][6][7] ( Fig. 3).…”

Capillary electrophoretic immunoassay for alpha-fetoprotein with chemiluminescence detection

“…A method of high sensitivity and selectivity is thus greatly needed for the determination of AFP in biofluids such as serum. AFP levels are often measured by enzyme-linked immunosorbent assay (ELISA) [26], radioimmunoassay (RIA) [27], chemiluminescent enzyme immunoassay (CLEIA) [28], flow-injection chemiluminescence immunoassay (FI-CLIA) [21] immunosensor [29,30], and CEIA-LIF [31]. However, CEIA-CL detection of AFP has not been reported so far.…”