Abstract
The pacu is one of the most important species for Brazilian fish farming and is considered emerging in the global aquaculture. Despite its importance, no effective tool for evaluation of the adaptive immune response of this species has been developed. Therefore, this study aimed the development and standardization of indirect ELISA for the measurement of pacu antigen-specific antibodies using polyclonal rabbit anti-pacu IgM used as detector antibody. For this purpose was isolated and purificated of pacu IgM using mannose-binding protein affinity chromatography and produced specific polyclonal antibodies against heavy and light chains pacu IgM, that showed a molecular weight of 72 kDa and 26 kDa, respectively. Polyclonal antibodies obtained demonstrated specificity with heavy and light Ig chains of pacu serum in western blotting. These polyclonal antibodies allowed the development of an indirect ELISA assay of high sensitivity and specificity for the detection and quantification of pacu IgM antibodies immunized with bovine IgG. In conclusion, this approach has great potential to improve the monitoring of vaccine-induced immune responses and help develop immunodiagnostic and epidemiological studies of infectious diseases in pacu systems.
Highlights
Pacu (Piaractus mesopotamicus) is a native species of the Paraná-Paraguay River Basin in South America and one of the main fish species cultured in Brazil (Valladão et al 2016)
The titration of rabbit anti-pacu IgM antibodies determined by indirect enzyme-linked immunosorbent assay (ELISA) showed high level of specific antibodies against the purified pacu IgM and a positive reaction was detected up to the highest serum dilution tested (1:16000)
Pacu IgM was effectively purified by affinity chromatography with the lectin known as mannose binding protein (MBP)
Summary
Pacu (Piaractus mesopotamicus) is a native species of the Paraná-Paraguay River Basin in South America and one of the main fish species cultured in Brazil (Valladão et al 2016). Pacu is an studies during the process of chemical registrations in Brazil (Castro et al 2014). The intensification of pacu culture has led to outbreaks of several diseases caused mainly by bacterial pathogens resulting in reduced yields and significant economic losses (Farias et al 2016). The lack of information on the pacus immune system is a limiting factor for the development of serological tests and vaccines for immunodiagnosis and monitoring of bacterial diseases treatment, which hinder the implementation of more effective biosecurity measures in intensive production systems. To date no studies have been reported on the isolation of pacu immunoglobulins
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