This DENV DENV Envelope overexpression lysate was created in Baculovirus-Insect Cells and intented for use as a Western blot (WB) positive control. Purification of DENV Envelope protein (Cat: 40531-V08B) from the overexpression lysate was verified.
A DNA sequence encoding the Dengue virus (DENV)(type 1, strain US/Hawaii/1944) E (ACF49259.1) (Val580-Lys680) was expressed with a polyhistidine tag at the C-terminus.
The recombinant Dengue virus (DENV)(type 1, strain US/Hawaii/1944) E consists of 112 amino acids and predicts a molecular mass of 12.4 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.