[Preparation of anti-mouse PGRP-L single-epitope polyclonal antibody].
OBJECTIVE
To prepare a single-epitope polyclonal antibody against mouse long peptidoglycan recognition protein (mPGRP-L).


METHODS
B cell dominant epitopes of mPGRP-L predicted by bioinformatics were synthesized, and the immunogen was prepared by conjugation of the synthetic peptide and the carrier protein key-hole limpet hemocyanin (KLH) by MBS method. The single-epitope polyclonal antibody was obtained by immunizing rabbits with the KLH-peptide conjugate, purified by SPG affinity columns or antigenic peptide affinity columns, and identified by ELISA and Western blotting.


CONCLUSIONS
A dominant epitope in N-terminal region of mPGRP-L, with amino acid sequence of NH(2)-(C)DPHSLSPELQALISEVAQHD-COOH, was chosen and synthesized. The titer of anti-serum of the rabbits immunized with the KLH-peptide conjugate was 1:256,000. The polyclonal antibody purified with SPG affinity columns and antigenic peptide affinity columns were named as mPGRP-Ln1 and mPGRP-Ln2, respectively. Western blotting demonstrated that the antibody mPGRP-Ln1 could recognize the recombined N-terminal fragment of mPGRP-Ln with a clear band at relative molecular mass of about 29,000, suggesting the successful preparation of the single-epitope polyclonal antibody against the N-terminal region of mPGRP-L.
He
Zhi
Z
Department of Immunology, Southern Medical University, Guangzhou 510515, China.
Zhang
Li-yun
LY
Chen
Zheng-liang
ZL
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification; D000918:Antibody Specificity / Q000276:immunology; D015153:Blotting, Western; D002352:Carrier Proteins / Q000737:chemistry / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D006433:Hemocyanins / Q000737:chemistry; D007106:Immune Sera / Q000276:immunology; D051379:Mice; D008969:Molecular Sequence Data; D011485:Protein Binding; D011817:Rabbits
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D000939:Epitopes; D007106:Immune Sera; C118897:peptidoglycan recognition protein; D006433:Hemocyanins; C032808:keyhole-limpet hemocyanin