Preparation and molecular characterization of a polyclonal antibody as an efficient cutworm reference protein
2018
HuanYu, Hunan Agricultural University, Changsha, Hunan, China | Lei He, Hunan Agricultural University, Changsha, Hunan, China | Yi-Yi Ou-Yang, Hunan Agricultural University, Changsha, Hunan, China | Guo-Hua Huang, Hunan Agricultural University, Changsha, Hunan, China
Antibodies, which are wildly used in molecular researches, are proteins secreted by plasma cells that can specifically recognize specific antigens. To obtain a stable reference antibodies are important for protein analysis. In this study, a capable glyceraldehyde phosphate dehydrogenase (GAPDH) antiserum against most noctuid larval protein samples was prepared. Firstly, the full-length gapdh was amplified from Spodoptera exigua larvae to construct the prokaryotic expression vector. The purified His-tag fused GAPDH protein was used to immunize rabbits for the antiserum preparation. Protein samples extracted from 11 insect species distributed across seven families and three orders were used to perform the Western bolt analysis. The bright specific bands were detected in S. exigua, S. litura, Helicoverpa armigera, and Mythimna seperata protein samples, indicating that this antiserum may be capable of detecting noctuid larval encoded GAPDH. Further immunofluorescence identification of H. armigera and S. exigua paraffin section slides with GAPDH antiserum exhibited an identical cyto-stained image. The GAPDH antiserum prepared in this study is useful and efficient as reference antibodies in studies involving noctuid larval protein samples in other related fields.
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