This HIV HIV gp140 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of HIV gp140 protein (Cat: 40259-V02H) from the overexpression lysate was verified.
A DNA sequence encoding the HIV-1 envelope glycoprotein gp160 (ACY40659.1) extracellular domain (Tyr38-Gln684), termed as gp140, was expressed with the Fc region of human IgG1 at the C-terminus.
The recombinant HIV1 gp140 (group P) consists 885 amino acids and predicts a molecular mass of 99.4 kDa.
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.