LDHB activity was measured from Hela cell lysate, which contains hetero-tetramer isozyme composed of both (M) LDHA and (H) LDHB subunits. However, the liver derived cell line, HepG2, which contains mainly homo-tetramer of subunit M (LDHA), shows no enzymatic activity.
The species cross reactivity of this assay was demonstrated by testing the sera from multiple species. The kit shows a good reaction with human, goat and bovine samples, but shows no reaction with mouse or rat sera which may also contain LDHB. This species cross reaction specificity is due to the species specificity of the capture antibody.
Lane 1 - HeLa cell Lysate Lane 2 - HepG2 Cell Lysate Lane 3 - LDHA (purified from rabbit skeletal muscle) Lane 4 - LDHB (recombinant) Lane 5 - Hela – In-well extracted with capture mAb Lane 6 - HepG2 – In-well extracted with capture mAb Lane 7 - LDHA – In-well extracted with a capture mAb Lane 8 - LDHB – In-well extracted with capture mAb The western blot results indicate that the capture antibody isolates active LDHB, but does not isolate LDHA (which would be detected by the anti-LDH antibody).
