Human Integrin alpha 5 beta 1 HEK293 Overexpression Lysate: 产品信息
This Human Integrin alpha 5 beta 1 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of Integrin alpha 5 beta 1 protein (Cat: CT014-H2508H) from the overexpression lysate was verified.
A DNA sequence encoding the extracellular domain (Met 1-Tyr 995) of human ITGA5 (P08648) was fused with a flag tag at the C-terminus, constructed the plasmid 1; A DNA sequence encoding the extracellular domain (Met 1-Asp 728) of human ITGB1 (P05556-1) was fused with a polyhistidine tag at the C-terminus, constructed the plasmid 2. The two plasmids were co-expressed and the heterodimer was purified
The recombinant heterodimer of human ITGA5/ITGB1 comprises 1745 (996 + 749) amino acids and has a calculated molecular mass of 192 (109 + 83) kDa. As a result of glycosylation, the apparent molecular mass of rh ITGA5/ITGB1 heterodimer is approximately 120 &140 kDa in SDS-PAGE under reducing conditions
Human Integrin alpha 5 beta 1 HEK293 Overexpression Lysate: 使用指南
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.