Figure 7
Ganglioside dot blot immunoassay employing MAb 10A5. Rows of a PVDF membrane were spotted with: (A) the Folch upper phase extracted from 2 ÎĽg of 25/45 fraction protein, (B) 5 ÎĽg of bovine mixed ganglioside standards, (C) 1 ÎĽg of purified bovine GD1a, (D) 50 ng of the negative control protein RadA-6xHis, and (E) 2.5% BSA in 50% methanol. The antigens were probed with: (1) alkaline phosphatase-conjugated secondary antibody in the absence of primary antibody; (2) MAb 2A2, an isotype-matched negative control antibody specific for the irrelevant bacterial protein RadA-6xHis; and (3) MAb 10A5. Panel 4 was stained with Coomassie Blue R-250 and destained in 40% methanol/10% acetic acid to reveal white ganglioside spots against a blue background.