سال انتشار: ۱۳۸۷

محل انتشار: اولین کنگره بین المللی مدیریت بهداشتی و بیماریهای آبزیان

تعداد صفحات: ۱

نویسنده(ها):

M Alishahi – Faculty of Veterinary Medicine, Shahid Chamran University, Ahvaz, Iran
M Mesbah –
M Ghorbanpoor Najafabadi –

چکیده:

Objective: Recently Barbus sharpeyi has artificially reproduced in some governmental fish hatchery and was accepted by private farms as a member of polyculture system in cyprinid farms. The analyses of fish immunoglubolins is important for the development of approaches to improve fish immunity against infectious disease. In this study, molecular weight (MW) of Igs and MW of Ig chains was determined. IgM was purified from serum using combination of ammonium sulphate precipitation and affinity chromatography.
Method & Materials: Ten Barbus sharpeyi (weighing 515±۵۸g) were immunized with goat IgG (1mg/fish) 3 times with 2 weeks intervals. 14 days after last injection fresh serum was separated from the blood samples of immune fish. 50% saturated ammonium sulphate precipitation was used to remove the albumin fraction. The purification of sera IgM was carried out on an immunoaffinity column (2 ml) of goat IgGlinked Cyanide Bromide (CN-Br) activated agarose- Sigma. First CN-BR- actvated agarose coupled with goat IgG. Purified proteins were analyzed using Sodium Dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) under reducing and non-reducing condition.
Results & Conclusion: Total protein of serum was 40 mg/ml. Analysis of molecular weight (MW) of purified Igs showed that: Under non-reducing, but denaturing, conditions in 5% SDS-PAGE gels, a high molecular weight (850 kDa) and a low molecular weight (168 kDa) fraction were demonstrated. SDS-PAGE Under reduced condition showed this fraction to be composed of disulphide linked heavy and light chains with MWs of 79 and 27 kDa respectively, suggesting that the native molecule was likely to be tetrameric and monomeric in structure. In conclusion, the results showed that high purity Ig was extracted by these methods and extraction with combination of precipitation with ammonium sulphate precipitation and affinity chromatography gave proper yield. Regarding to Ig structure it appears that Barbus sharpeyi is similar to other teleosts in that they possess the predominant high molecular weight Ig tetramer which dissociates into H chains and L chains.