[Research for development of immunoassay kits for hapten].

Determination of estrogen concentrations in the urine is widely utilized for a variety of purposes, such as the assessing of fetoplacental unit function and ovarian function, and the monitoring of follicle in inducing ovulation in the treatment of infertility. Since hapten antigens such as estrogen have low molecular weight, unlike high molecular substances, they do not form visible agglutination of antigen-antibody reaction, so there has been no other choice but to depend on radioimmunoassay for the determination of estrogen concentrations. However, radioimmunoassay requires special facilities and apparatus and is complicated in procedure, and these drawbacks have prevented the estrogen assay from routine tests in spite of clinical usefulness. As a result of various researches, a new immunochemical principle called competitive agglutination inhibition reaction has been developed for the determination of hapten concentrations, and succeeded in putting this principle to practical use with a fetoplacental unit function test kit (Estrotec slide test) and highly sensitive semi-quantitative analysis kits for urinary estrogen (Hi-estrotec and Hi-estrotec slide).

Bactericidal activity of M14659 enhanced in low-iron environments.

The bactericidal activity of M14659 against Escherichia coli in low-iron environments was investigated and compared with that of ceftriaxone and ceftazidime. The bactericidal activity of M14659 against E. coli in Mueller-Hinton broth was enhanced 30- to 20,000-fold by addition of transferrin, which is an iron-binding protein, whereas the activity of ceftriaxone or ceftazidime was much less strongly affected. This enhancement by transferrin was completely inhibited by saturating the iron-binding capacity of transferrin with FeCl3. M14659 was taken up markedly into bacterial cells in the presence of transferrin, and its uptake was inhibited by the protonophore dinitrophenol, which inhibits active-transport systems coupled to an energized membrane such as the iron transport systems of E. coli. The bactericidal activity of M14659, which chelates Fe3+, was also enhanced in the presence of other iron-binding compounds such as lactoferrin and alpha,alpha'-dipyridyl or in iron-deficient Mueller-Hinton broth (Fe3+ concentration, less than 2 nM) supplemented with FeCl3 at 0.1 to 1.0 microM, but not in unsupplemented iron-deficient Mueller-Hinton broth. The E. coli used in this study was confirmed to derepress iron transport systems in the presence of transferrin, lactoferrin, and alpha,alpha'-dipyridyl and in the iron-deficient Mueller-Hinton broth supplemented with FeCl3 at 0 to 1.0 microM. M14659 also showed an excellent antibacterial activity in vitro against other gram-negative bacteria in the low-iron environments. These findings indicate that M14659 may be actively taken up with Fe3+ into bacterial cells, probably through the iron transport systems under conditions of low iron and, thus, kills bacteria effectively.