Detection of chromogranin A in the adrenal gland extracts of different animal species by an enzyme-linked immunosorbent assay using Thomsen–Friedenreich antigen-specific Amaranthus caudatus lectin

• Published in: Veterinary immunology and immunopathology 2011-12, Vol.144 (3), p.255-258
• Main Authors: Akiyoshi, Hideo, Sugii, Shunji, Nahid, Md. A., Sone, Katsuhito, Tanaka, Toshiyuki, Zheng, Cao, Yijyun, Li, Aoki, Mica, Takenaka, Shigeo, Shimada, Terumasa, Shimizu, Junichiro, Kiyomiya, Ken-ichi, Ohashi, Fumihito
• Format: Article
• Language: English
• Subjects: Adrenal Glands - chemistry; Amaranthus - immunology; Amaranthus caudatus; Animals; Antigens, Tumor-Associated, Carbohydrate - immunology; Blotting, Western; Cattle; Cetacea; Chromogranin A; Chromogranin A - analysis; Chromogranin A - immunology; Dogs; Dolphins; Electrophoresis, Polyacrylamide Gel; Enzyme-Linked Immunosorbent Assay - methods; Horses; Lectin; Plant Lectins - immunology; Rabbits - immunology; Swine; Tissue Extracts - chemistry
• ISSN: 0165-2427; 1873-2534
• DOI: 10.1016/j.vetimm.2011.08.024

The reactivity of different lectins with crude chromogranin A (CgA) obtained from different animals, namely, cow, horse, dog, pig, and dolphin, was examined to identify lectin(s) that would be useful as coating reagent(s) in a sandwich enzyme-linked immunosorbent assay (ELISA). Of the different lectins studied, the Amaranthus caudatus lectin (ACA), which is specific for the Thomsen–Friedenreich (T)-antigen (Galβ1-3GalNAc), was found to react with the CgA from different animals by western blotting. Purified rabbit anti-bovine CgA antibody was also found to cross-react with the crude CgA preparations. On the basis of these findings, a sandwich ELISA was developed with ACA as the coating reagent and anti-bovine CgA antibody as the probing antibody. Using this method, concentration-dependent curves ranging from 0.003 μg/mL to 25 μg/mL and from 0.02 μg/mL to 25 μg/mL were obtained for bovine CgA and canine CgA, respectively. Similarly, concentration-dependent curves were obtained for the equine, swine, and dolphin crude CgA extracts. Thus, ACA is concluded to be a valuable reagent for CgA detection in crude extracts from different animal species, and for CgA isolation/purification.