Rat S100 beta protein, His tag

Protein S100B is a member of the S100 family. S100 proteins are EF-hand calcium-binding proteins and involved in the regulation of a number of cellular processes such as cell cycle progression and differentiation. Experimental results suggest that the receptor for advanced glycation end products (RAGE) plays important roles in mediating S100 protein-induced cellular signaling. Besides, rat RAGE shares similarities with human RAGE in amino acids sequence with the identity of 80.0% . Thus a binding ELISA assay was conducted to detect the interaction of recombinant rat S100B and recombinant human RAGE. Briefly, S100B were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to RAGE-coated microtiter wells and incubated for 2h at 37ºC. Wells were washed with PBST and incubated for 1h with anti-S100B mAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody, wells were aspirated and washed 3 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37ºC. Finally, add 50 μl stop solution to the wells and read at 450nm immediately. The binding activity of of S100B and RAGE was in a dose dependent manner.