Abstract
Until now, custom-made or commercial polyclonal antibody against only one kind of fish IgM limited application of the antibody. During our research on development of vaccine against infection of Clonorchis sinensis (C. sinensis) in several kinds of fish, we were conscious of the urgency of secondary antibody to evaluate immune effect and screen C. sinensis infection with immunological technology instead of labor-intensive and time-consuming squash or artificial digestion of fish flesh. So that, we purified IgM of grass carp, bighead carp, crucian carp, common carp and tilapia which were widely cultured freshwater fishes in most areas of China. On this basis, we generated HRP-conjunct rabbit IgG anti-fish IgMs with high titers. IgM of other freshwater fishes including oshima, yellow catfish, bream, silver carp and so on could be recognized by the IgG sensitively. Additionally, The ELISA detection displayed that the IgG could be more specific and sensitive than custom-made rabbit IgG anti-grass carp IgM. The acquirement of HRP-conjunct rabbit IgG anti-fish IgMs was the cornerstone for studying the immune system of teleost fish, developing immunoassay methods and evaluation of fish vaccine with more convenience.
Highlights
Immunoglobulins (Igs) play important roles in humoral adaptive immunity of vertebrate
Two main bands around 66kDa and 25kDa were observed in each sample, which were approximate to the molecular weight (MW) of H and L chain of fish Immunoglobulin M (IgM) (Figure 1)
The total quantity of IgM from crucian carp or common carp was respectively only 2.856mg or 2.87mg, but it was enough for subsequent immunization
Summary
Immunoglobulins (Igs) play important roles in humoral adaptive immunity of vertebrate. Fish belong to a kind of lower vertebrate. It has been documented that Igs-centric specific immunologic mechanism first appeared in fish (Lobb & Clem 1981). Immunoglobulin M (IgM) from teleost, an analogous to IgG of mammal, is the predominant isotype found in serum and mucus (Solem & Stenvik 2006). Teleost IgM contributes to both innate and adaptive immunity in fish (Mashoof & Criscitiello 2016). It can mediate agglutination for phagocytosis and removing pathogen, and cellular cytotoxicity (Ye et al 2013). Teleost IgM typically is expressed as a tetramer containing two heavy (H) chains with molecular weight (MW) of 70-81 kDa and two light (L) chains with MW of 22-32 kDa (Vesely et al 2006a, Ye 2008)
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