Rat SLIT2 protein, His tag

Slit is a family of secreted extracellular matrix proteins which play an important signalling role in the neural development of most bilaterians. Humans, mice and other vertebrates possess three Slit homologs: Slit1, Slit2 and Slit3. The Slit2 protein has recently been discovered to be associated with the development of new blood vessels from pre-existing vessels, or angiogenesis. Slit2 has been implicated in promoting angiogenesis in mice (both in vitro and in vivo), in the human placenta, and in tumorigenesis. Besides, Gremlin 1 (GREM1) has been identified as an interactor of Slit2, thus a binding ELISA assay was conducted to detect the interaction of recombinant rat Slit2 and recombinant rat GREM1. Briefly, Slit2 were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to GREM1-coated microtiter wells and incubated for 2h at 37ºC. Wells were washed with PBST and incubated for 1h with anti-Slit2 pAb, 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 Slit2 and GREM1 was in a dose dependent manner.