An Improved Protocol for Coupling Synthetic Peptides to Carrier Proteins for Antibody Production Using DMF to Solubilize Peptides
An Improved Protocol for Coupling Synthetic Peptides to Carrier Proteins for Antibody Production Using DMF to Solubilize Peptides
- Journal of BioMolecular Techniques, 18(3), p.173-176, 2007 .
We present an improved protocol for coupling synthetic peptides to carrier proteins. In this protocol, dimethylformamide is used as the solvent to solubilize peptides instead of phosphate-buffered saline (PBS)or 6 M guanidine-HC l/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 mmaleimidobenzoyl- 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-HC l.
PEPTIDE
ANTIBODY
KLH
BSA
MBS
DMF
MALDI- TOF MS
We present an improved protocol for coupling synthetic peptides to carrier proteins. In this protocol, dimethylformamide is used as the solvent to solubilize peptides instead of phosphate-buffered saline (PBS)or 6 M guanidine-HC l/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 mmaleimidobenzoyl- 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-HC l.
PEPTIDE
ANTIBODY
KLH
BSA
MBS
DMF
MALDI- TOF MS