Localization of an EGF receptor in purified chromosomes from A431 and lymphoblast cells

Condensed chromosomes from mitotic cells of A431 and lymphoblast GM131 cells were purified by a flow cytometric sorting and collected directly onto nitrocellulose discs. The blots containing purified chromosomes from A431 cells were then hybridized against iodinated protein ligands. Positive autoradiographic signals were obtained from 125I-epidermal growth factor (EGF), 125I-calmodulin(CAM)-IgG and 125I-number528 monoclonal IgG against EGF receptor, but not from 125I-CAM. Binding of 125I-EGF to the purified chromosomes on the nitrocellulose was specific and reversible upon the addition of excess unlabeled EGF to the hybridization solutions. Blots containing the purified chromosomes of lymphoblast cells gave positive signals for 125I-CAM-IgG, while 125I-CAM signals were not detected. Control experiments showed no detectable contamination from A431 membrane EGF receptor in the purified chromosomal fraction, as demonstrated by the absence of EGF receptor autophosphorylation and the absence of a 125I-EGF hybridization signal in the supernatant solution of the purified chromosomal preparation