Detection of tetanus antitoxin using Eu(3+)-labeled anti-human immunoglobulin G monoclonal antibodies in a time-resolved fluorescence immunoassay

Abstract

Tetanus antitoxin in human sera was detected with solid-phase immunoassays in microtitration modules coated with tetanus toxoid by using Eu3+-labeled anti-human monoclonal antibodies on the basis of an exactly calibrated antibody standard. The use of a time-resolved fluorescence immunoassay (TR-FIA) significantly improved the quantitative detection of tetanus antitoxin over that of the enzyme-linked immunosorbent assay (ELISA) technique because of its high sensitivity and its wide measurement range, detecting antibody levels between 0.001 and 12.5 IU/ml with a single serum dilution of 1:100. For the same purpose, two different serum dilutions (1:100 and 1:1,000) were needed in the ELISA technique. TR-FIA is reproducible and can be performed in 3.5 h. A study of 2,630 serum samples was undertaken to examine the age-dependent distribution of titer levels, indicating the decline of sufficient protection in patients older than 60 years. The wide measurement range of TR-FIA enabled fast examination of large numbers of serum samples without the need for repetition, with further sample dilution, as was often necessary in the ELISA procedure.

Detection of adenovirus in clinical specimens by polymerase chain reaction and liquid-phase hybridization quantitated by time-resolved fluorometry

“…The concept of linking PCR to hybridization with probes that are labeled for detection by TR fluorometry, however, is novel and was generated by the recent advances in europium labeling techniques and in TR-FIA. While the europium labeling of antibodies and other proteins has been established since 1980 (9,13,17,21,24), the chemistry for labeling oligonucleotides with europium chelates has only recently been developed and perfected (4,5,16,18,23). The fact that an amino-modified cytidine phosphoramidite analog can be used to introduce reactive sites onto an oligonucleotide enables the labeling of a single probe with even more than 20 Eu chelates (5,23).…”

Increased maternofoetal transfer of antibodies in a murine model of systemic lupus erythematosus, but no immune activation and neuroimmune sequelae in offspring

“…In contrast to this, levels significantly above those of females across all ages assayed were present in males, already at 13 weeks of age. Finally, our experimental approaches to assay anti-dsDNA were similar: Denenberg and colleagues employed an ELISA-based assay, we employed a TRIFMA; both are solid-phase assays, but TRIFMA is generally considered more sensitive than ELISA (Siitari et al, 1990;Schroder and Kuhlmann, 1991;Bucher et al, 1991).…”

A New Method for Detection African Swine Fever Virus: Time-resolved Fluorescence Immunoassay

“…Compared with ELISA, TRFIA has higher sensitivity and wider measurement range. Schröder and Kuhlmann found that the use of a TRFIA significantly improved the quantitative detection of tetanus antitoxin over that of the ELISA technique, and the wide measurement range of TRFIA enabled fast examination of large numbers of serum samples without the need for repetition [ 23 ]. In this study, a novel TRFIA methodology was developed for the quantitative determination of ASFV antigen in pigs nasal discharge.…”