Immunoblot analysis of tyrosine-phosphorylated proteins in the lysates of CLL cells.
Cell lysates from 5 × 106 cells were loaded onto separate lanes of the polyacrylamide gel for immunoblot analysis with the anti–P-Tyr antibody 4G10. Each cell sample is indicated by a number above the pair of lanes for lysates made from cells before (−) or after (+) BCR stimulation with anti-μ F(ab′)2. Lysates from ZAP-70+ cases CLL1 (1) and CLL2 (2) are compared with those of CLL13 (13) and CLL14 (14) in the left panel. Lysates from samples obtained from monozygous twins, CLL16 (16) and CLL17 (17), but that were discordant for expression of ZAP-70, are provided in the right panel. Molecular weight markers are indicated on the left and right margins. The brackets define the areas between 70 and 72 kDa in size that were scanned for density with NIH image software. The blots were stripped and then probed with anti–β-actin to monitor for uniformity of protein loading (bottom row labeled “β-actin” on the right-hand margin).
