This EV-D68 EV-D68 VP4 overexpression lysate was created in Baculovirus-Insect Cells and intented for use as a Western blot (WB) positive control. Purification of EV-D68 VP4 protein (Cat: 40467-V31B) from the overexpression lysate was verified.
A DNA sequence encoding the VP4 of Enterovirus D68 (EV-D68)(strain Fermon) (AAR98503.1) (Met1-Lys69) was expressed with the Fc region of rabbit IgG at the C-terminus.
The recombinant VP4 of Enterovirus D68 (EV-D68)(strain Fermon) consists of 293 amino acids and predicts a molecular mass of 32.7 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.