ACCURATE QUANTITATIVE-ANALYSIS OF CELLULAR PROTEINS USING A COMPUTERIZED SCANNING-AIDED DOT-BLOT TECHNIQUE

The aim of this study was to use a dot blot technique (DB) aided by a computerised scanning programme (CSDBT) to assess expression of cellul ar proteins in various tumour cell lines. Two groups of proteins, ie. major histocompatibility complex (MHC) Class I and cell adhesion molec ules (e.g. ICAM-1) were investigated. The constitutive expression of C lass I antigens was measured and found to vary for different tumour li nes. Exposure of SKV14 (SV40 transformed epithelial cells) and 5637 li nes (bladder) to interferons (IFN) alpha or gamma resulted in upregula tion of Class I antigens. IFN gamma, but not IFN alpha, induced ICAM-1 on all the lines studied. Thus, the stimulatory indices (SI) for Clas s I antigens in the case of SKV14, J82 (bladder) and 5637 lines after IFN gamma, and IFN alpha stimulation were 2.02, 1.77, 4.68 and 1.65, 1 .36, 2.08 respectively The corresponding values for ICAM-1 were 1.65, 1.87, 3.43 and 1.75, 1.0, 0.96. In all the cases, the P values (using a paired t-test), except ICAM-1 for IFN alpha-stimulated J82 cells, we re below 0.05. Following exposure of cells to combinations of drugs an d IFN alpha, the percentage inhibition for Class I and ICAM-1 in the c ase of SKV14 and 5367 lines in the presence of cycloheximide (10 mu g/ mL) were 82%, 85% and 57%, 45% respectively. The corresponding values for indomethacin (10 mu g/mL) were -23%, -52% and -20%, -24%. In all c ases, the P values (using a paired t-test) were below 0.05. To our kno wledge this is the first report describing a computerised scanning pro gramme for assessing the presence of cellular proteins. The results we re consistent with those obtained by radiobinding and immunocytochemis try. This simple and sensitive approach highlighted two important issu es: (a) that it can be used to detect minor changes, under various con ditions, of cellular proteins whether cytoplasmic or otherwise; and (b ) that the accuracy of the measurements was enhanced by the use of com puter scanning.