Generating NK cell receptor-Fc chimera proteins from 293T cells and considerations of appropriate glycosylation.
The use of recombinant receptors as a scientific tool has become widespread in many research fields. Of particular interest are the natural killer (NK) receptors that play a major role in the immune response against tumors and virus-infected cells. We present here (i) a detailed protocol for the production and purification of soluble recombinant NK cell receptors tagged with human IgG1-Fc (thus termed receptor-Fc chimera or receptor-Ig fusion protein) and (ii) a protocol for cell staining with these recombinant receptor-Fc chimeras. As these recombinant proteins are produced in eukaryotic cells, we further discuss the glycosylation pattern of these receptors that might interfere with their ligand-binding phenotype.
Zilka
Alon
A
Department of Microbiology and Immunology, Ben Gurion University, Beer Sheva, Israel.
Mendelson
Michal
M
Rosental
Benyamin
B
Hershkovitz
Oren
O
Porgador
Angel
A
D000818:Animals; D002460:Cell Line; D005434:Flow Cytometry; D006031:Glycosylation; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D015202:Protein Engineering / Q000379:methods*; D055607:Receptors, Natural Killer Cell / Q000302:isolation & purification / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism*; D013194:Staining and Labeling; D013997:Time Factors; D014162:Transfection
C029152:G-substrate; D007141:Immunoglobulin Fc Fragments; D009419:Nerve Tissue Proteins; D055607:Receptors, Natural Killer Cell; D011993:Recombinant Fusion Proteins