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Updated: Jun 15, 2026
Genetic Encoding of a Non-Canonical Amino Acid for the Generation of Antibody-Drug Conjugates Through a Fast Bioorthogonal Reaction
Published on: September 14, 2018
[Prokaryotic expression, purification and preparation of polyclonal antibody for human UNC5CL]
Dan Lü1, Xiao-ping Qian, Xiao-jun Tian
1Department of Immunology, Peking University Health Science Center, Beijing, China.
Aim:
To construct the prokaryotic expression vector of UNC5CL, purify the fusion protein of GST-UNC5CL, prepare the polyclonal antibody of UNC5CL and characterize the reactivity and specificity of the polyclonal antibody.
Methods:
The human UNC5CL nucleotide sequence encoding amino acid from 280 to 518 was amplified and cloned into pGEX-4T-2 vector, then transformed into E.coli BL21, in which the fusion protein GST-UNC5CL (aa280-518) was induced and purified by Glutathione Sepharose-4B. Then the purified protein immunized the rabbits and anti-serum was collected. The anti-serum was further detected by ELISA, Western blot and immunohistochemistry.
Results:
The prokaryotic expression vector of pGEX-4T-2-UNC5CL(aa280-518) was constructed successfully. It can be induced by IPTG and express a fusion protein of GST-UNC5CL (aa280-518) with M(r); 5 2000. The ELISA, Western blot and Immunohistochemistry demonstrated the rabbit antiserum obtained by immunized by the purified fusion protein can detect the target protein.
Conclusion:
The rabbit polyclonal antibody against human UNC5CL is prepared. It can react with UNC5CL specifically and provide basis for further function study of UNC5CL.
