An improved protocol for coupling synthetic peptides to carrier proteins for antibody production using DMF to solubilize peptides.
We present an improved protocol for coupling synthetic peptides to carrier proteins. In this protocol, dimethyl-formamide is used as the solvent to solubilize peptides instead of phosphate-buffered saline (PBS) or 6 M guanidine-HCl/0.01 M phosphate buffer (pH 7). Additionally, the last desalting or dialyzing step to remove uncoupled peptides as in the traditional method is eliminated. Finally, 3 ml of 0.1 M ammonium bicarbonate is added to the carrier protein conjugated peptide solution to help the lyophilization process. Coupling of Cys-containing synthetic peptides to keyhole limpet hemocyanin or bovine serum albumin using m-maleimidobenzoyl-N-hydroxysuccinimide ester are used as the test cases. This method produces high-quality antipeptide antibodies. Also, compared to the traditional method, this procedure is simpler and useful for peptides with solubility problems in PBS or 6 M guanidine-HCl.
Lateef
Syed Salman
SS
Protein Research Laboratory, Research Resources Center, University of illinois at Chicago, 835 S. Wolcott Ave, Chicago, IL, USA.
Gupta
Shalini
S
Jayathilaka
Lasanthi P
LP
Krishnanchettiar
Sangeeth
S
Huang
Jin-Sheng
JS
Lee
Bao-Shiang
BS
D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000737:chemistry* / Q000276:immunology* / Q000378:metabolism; D004126:Dimethylformamide; D007136:Immunoglobulins / Q000096:biosynthesis*; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis* / Q000276:immunology / Q000378:metabolism; D012995:Solubility
D002352:Carrier Proteins; D007136:Immunoglobulins; D010455:Peptides; D004126:Dimethylformamide