Development of gel staining techniques for detecting the secretion of procathepsin D (52-kDa protein) in MCF-7 human breast cancer cells Academic Article uri icon

abstract

  • 17 beta-Estradiol stimulates the secretion of the 34- and 52-kDa protein (i.e., cathepsin D and procathepsin D, respectively) from MCF-7 human breast cancer cells and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) inhibits this estrogen-stimulated response. A comparison of the effects of 17 beta-estradiol, TCDD, and their combinations on the secretion of these two proteins was determined using four different assay procedures, namely autoradiographic analysis of the 35S-labeled proteins (from [35S]methionine) separated by polyacrylamide gel electrophoresis (PAGE), densitometric analysis of the silver- and double-stained proteins separated by PAGE, and radioimmunoassay of the proteins using commercially available antibodies to the 52-kDa protein. The results showed that the autoradiographic, staining, and radioimmunoassay procedures gave comparable results with only a few minor differences in the relative amounts of the 52-kDa detected in the various treatment groups. In the medium obtained from 17 beta-estradiol-treated cells that was serially diluted, there was an excellent linear correlation for the relative concentrations of the 52-kDa protein using the double-staining/densitometric procedure and the radioimmunoassay. These results indicate that the double- or silver-staining method may be a useful and rapid method for screening new compounds as antiestrogens in MCF-7 cells.

author list (cited authors)

  • Krishnan, V., Narasimhan, T., & Safe, S.

citation count

  • 21

publication date

  • July 1992