Immunocytochemical detection of structural and regulatory proteins in rat adrenal nuclear matrix.
The nuclear matrix is a specific cell structure consisting of a residual nucleoskeleton that extends from the nucleoli to the nuclear envelope. The nuclear matrix of steroidogenic cells was isolated previously from a purified nuclear fraction. We present here an in situ extraction method, modified Lutz's method, for rat glandular adrenal cell nuclear matrix. This residual organelle was characterized and studied using immunocytochemical methods. The adrenal glands were removed, the cells prepared in suspension and deposited by cytospin onto Poly-L-lysine glass slides. The nuclear matrix was extracted with Nonidet P-40, DNase I and high and low ionic strength buffers. Structural proteins, nuclear lamins, coilin and fibrillarin were detected immunocytochemically. The adrenal fasciculata cells were easily identified by this method because of their large nuclei and abundant lipid droplets in the cytoplasm. After immunocytochemical detection by antibodies against lamins A and C, a marked brown layer at the periphery of the nucleus was observed. The intensity of the staining was lower using the antibody against nuclear lamin B. Immunocytochemical detection of the protein coilin revealed punctuated stained areas, 2-6 per nucleus, that probably correspond to the coiled bodies. The protein fibrillarin was detected at the nucleolus and coiled bodies. Our technique is simple, reveals well preserved adrenal nuclear matrices, and may be a useful method for immunocytochemical analysis and in situ hybridization.
Neves
D
D
Institute of Histology and Embryology, Faculty of Medicine of Oporto, Porto, Portugal.
Magalhães
M M
MM
Magalhães
M C
MC
D000311:Adrenal Glands / Q000166:cytology*; D000818:Animals; D002868:Chromosomal Proteins, Non-Histone / Q000032:analysis*; D007150:Immunohistochemistry; D034921:Lamin Type B; D034882:Lamins; D008297:Male; D015530:Nuclear Matrix; D009687:Nuclear Proteins / Q000032:analysis*; D051381:Rats; D017207:Rats, Sprague-Dawley
D002868:Chromosomal Proteins, Non-Histone; D034921:Lamin Type B; D034882:Lamins; D009687:Nuclear Proteins; C047458:fibrillarin