High-level expression of recombinant Fc chimeric proteins in suspension cultures of stably transfected J558L cells.
Recombinant Fc chimeric proteins are useful tools for studying protein function, including the analysis of molecular interactions by techniques such as expression cloning. Here we describe a method we have used to express the IgLON family proteins, CEPU1 and OBCAM, as recombinant Fc chimeric proteins in stably transfected mouse J558L myeloma cells. The use of this cell line provided the opportunity to maximize protein production, as it secretes antibodies in large quantities and can be grown to high density in small volumes of culture medium. Isolation of recombinant OBCAMFc from the adherent COS7 cell line suggested a minimum level of expression of 0.07 mg OBCAMFc/100 mL culture medium, while the J558L cell line expressed OBCAMFc at approximately 11.4 mg/100 mL culture medium. Purification of IgLON-Fc expressed by J558L cells was simpler than purification from COS7 cells because of the lower volume of culture medium generated. Furthermore, contamination of J558L expressed IgLONFc with bovine IgG from the culture medium was negligible. The method presented, which utilizes a commercially available small-scale bioreactor, provides the nonspecialist protein expression laboratory with the means to produce recombinant proteins quickly and easily in milligram quantities.
Howard
M R
MR
The University of Liverpool, UK.
Lodge
A P
AP
Reed
J E
JE
McNamee
C J
CJ
Moss
D J
DJ
D000818:Animals; D030161:Avian Proteins; D019556:COS Cells; D002352:Carrier Proteins / Q000235:genetics; D015815:Cell Adhesion Molecules / Q000235:genetics; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D002645:Chickens; D058851:GPI-Linked Proteins; D049452:Green Fluorescent Proteins; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000235:genetics*; D007136:Immunoglobulins / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics; D008562:Membrane Glycoproteins / Q000235:genetics; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000235:genetics; D014162:Transfection / Q000379:methods
D030161:Avian Proteins; D002352:Carrier Proteins; D015815:Cell Adhesion Molecules; D058851:GPI-Linked Proteins; D007141:Immunoglobulin Fc Fragments; D007136:Immunoglobulins; D008164:Luminescent Proteins; D008562:Membrane Glycoproteins; C059785:OPCML protein, human; C489214:Opcml protein, mouse; D011993:Recombinant Fusion Proteins; C102493:neural CEPU-1 protein, chicken; D049452:Green Fluorescent Proteins