Bioinformatics Analysis, Prokaryotic Expression, and Antiserum Preparation of hnRNP A1 Protein

Objective This study aimed to optimize the expression conditions of hnRNP A1 protein in BL21 competent cells through methods such as PCR and double enzyme digestion, to obtain high-concentration and high-quality purified protein, and to prepare high-titer polyclonal antibodies against hnRNP A1.Methods Bioinformatics tools were used to analyze the physicochemical properties and structure of hnRNP A1. The pET-28a-hnRNP A1 recombinant plasmid was constructed and transformed into BL21 cells. After optimizing expression conditions, hnRNP A1 protein was purified using nickel column chromatography and identified by Western Blot. The purified protein was used to immunize C57BL mice to produce polyclonal antibodies, and the antibody titer was determined by indirect ELISA.Results The highest expression of hnRNP A1 was achieved under conditions of 0.4 mM IPTG, induction temperature of 42°C, and induction time of 8 hours. The purified protein concentration reached 2.0563 μg/μl, and Western Blot confirmed the target protein. The antibody titer detected by indirect ELISA was 1:409,600.Conclusion The physicochemical properties of hnRNP A1 were successfully analyzed, high-efficiency expression of hnRNP A1 protein was achieved, and high-titer mouse-derived polyclonal antibodies against hnRNP A1 were prepared, providing a valuable tool for further research.