An improved protocol for coupling synthetic peptides to carrier proteins for antibody production using DMF to solubilize peptides. |
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Authors: | Syed Salman Lateef Shalini Gupta Lasanthi P Jayathilaka Sangeeth Krishnanchettiar Jin-Sheng Huang Bao-Shiang Lee |
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Institution: | Protein Research Laboratory, Research Resources Center, University of illinois at Chicago, 835 S. Wolcott Ave, Chicago, IL, USA. |
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Abstract: | 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. |
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Keywords: | peptide antibody KLH BSA MBS DMF MALDI‐TOF MS |
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