This Mouse ICOS overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of ICOS protein (Cat: 50466-M02H) from the overexpression lysate was verified.
A DNA sequence encoding the mouse ICOS (Q9WVS0) extracellular domain (Met 1-Leu 142) was fused with the Fc region of human IgG1 at the C-terminus.
The recombinant mouse ICOS/Fc is a disulfide-linked homodimer. The reduced monomer consists of 363 amino acids and has a predicted molecular mass of 40.9 kDa. In SDS-PAGE under reducing conditions, the apparent molecular mass of rmICOS/Fc monomer is approximately 40-45 kDa.
Mouse ICOS 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.
Inducible costimulator (ICOS), also called AILIM (activiation-inducible lymphocyte immunomediatory molecule) is a cell-surface receptor, and belongs to the CD28 family of immune costimulatory receptors consisting of CD28, CTLA-4 and PD-1. The interaction of B7-H2/ICOS plays a critical role in Th cell differentiation, T−B cell interactions which is essential for germinal center formation, and humoral immune responses, and as well as the production of cytokine IL-4. In addition, ICOS is more potent in the induction of IL-1 production, a cytokine important for suppressive function of T regulatory cells. The B7-1/B7-2--CD28/CTLA-4 and ICOS-B7RP-1 pathway provides key second signals that can regulate the activation, inhibition and fine-tuning of T-lymphocyte responses. ICOS stimulates both Th1 and Th2 cytokine production but may have a preferential role in Th2 cell development. Moreover, The B7-1/B7-2-CD28/CTLA-4 and ICOS-B7RP-1 pathway has been suggested of being involved in the development of airway inflammation and airway hyperresponsiveness.