E. Martinezgalisteo et al., CHARACTERIZATION OF MAMMALIAN THIOREDOXIN REDUCTASE, THIOREDOXIN AND GLUTAREDOXIN BY IMMUNOCHEMICAL METHODS, Comparative biochemistry and physiology. B. Comparative biochemistry, 111(1), 1995, pp. 17-25
Specific polyclonal antibodies towards the oxidized form of bovine thi
oredoxin reductase (TR) have been obtained in rabbits, and purified. T
he antigenicity was lost upon reduction of TR by NADPH indicating a la
rge conformational change upon reduction of the redox-active disulfide
in the enzyme, The antibodies did not cross-react with other bovine N
ADPH-dependent dehydrogenases. No reactivity was observed with TR from
bacteria, yeast or rat and only a slight reaction was obtained with T
R from horse, Immunoaffinity purified anti-thioredoxin and anti-glutar
edoxin antibodies were used to develop competitive indirect ELISA assa
ys that were validated giving very good linearity, reproducibility, se
nsitivity and parallelism, The glutaredoxin (Grx) immunoassay is the f
irst quantitative method described to measure the protein. When applie
d to a battery of calf tissues the contents of Grx varied from 7 to 12
0 mu g per gram of fresh tissue. Skeletal and heart muscles gave the l
owest values and spleen and salivary glands the highest. However, skel
etal muscle showed the highest gluthathione-hydroxyethyl disulfide oxi
doreductase specific activity.