TM5 and TM5b are the 2 major TM isoforms in human erythrocyte membranes.

(A) A Coomassie blue stain of several purified TM isoforms and their chimeric molecules separated on a 12% gel using SDS-PAGE. (B) The Western blot analysis of the above, with antibody R-41. Samples are 30 μg human ghost membrane (lane 1), chicken leg TM (lane 2), chicken gizzard TM (lane 3), hTM5/2 (lane 4), hTM2 (lane 5), hTM3 (lane 6), hTM5 (lane 7), hTM5/3 (lane 8), and hTM3/5 (lane 9). Approximately 10 μg TM isoforms (except hTM3/5, which has less) are applied. (C) The following lanes depict the Western blot analysis of 30 μg human ghost membrane by mAb Pep3-43^1:100 (lane 3), antibody R-41^1:100 (lane 4), and mAb LC1^1:500(lane 5). The panel also depicts the Coomassie blue stains of human ghost membrane (lane 2) and the molecular weight standards (lane 1). (D) A sequential Western blot analysis demonstrates the distinction between hTM5 and hTM5b. Lane 1 presents the first round of Western blot with mAb Pep3-43,^1:100 while lane 2 shows the same strip after the second round of Western blot with mAb LC1.^1:1000