Preparation and characterization of the antibodies against human myofibrillogenesis regulator 1 and application in neonatal rat cardiomyocytes
- VernacularTitle:人肌纤生成调节因子-1抗体的制备、鉴定及其在乳鼠心肌细胞中的应用
- Author:
Xiaoreng WANG
;
Xiuhua LIU
;
Quansheng SONG
;
Ting LI
;
Wenling HAN
- Publication Type:Journal Article
- Keywords:
Myofibrillogenesis regulator 1;
Antibodies;
Neonatal rat cardiomyocytes
- From:
Chinese Journal of Pathophysiology
2010;26(1):42-47
- CountryChina
- Language:Chinese
-
Abstract:
AIM: To prepare and purify the polyclonal antibodies against human myofibrillogenesis regulator 1 (hMR-1), then to characterize the purity, titer, specificity and the availability.METHODS: Two polypeptides named peptide 1 and 2 were synthesized based on the bioinformatics analysis of the sequence of hMR-1 by using software TMHMM and DNAStar, then coupled with keyhole limpet hemocyanin (KLH) for immunization. These peptides for immunization were mixed and injected into New Zealand rabbits to prepare antibodies specifically against hMR-1. ELISA assay was used to detect the titers of the antibodies. After purification by immunoaffinity chromatography, antibodies were identified by Western blotting and immunocytofluorescent assays. Applications of the antibodies on neonatal rat cardiomyocytes were also employed.RESULTS: (1)The titers of antibodies were 1:10~5. In WB assay, a specific 17kD band was detected, corresponding to the predicted molecular weight of hMR-1; the positive fluorescent signals were distinct. (2)On the neonatal rat cardiomyocytes model, we observed a peri-nucleus location. The fluorescent signal of hMR-1 overexpression group was much stronger than that in vector control and normal control groups.CONCLUSION: All these results indicate that the antibodies obtained from poly peptides mixture immunization have either human original or rat original antigens. The antibody is available for using in Western blotting or immunofluorescent assays.
