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Abstract
For the efficacy test of animal biologics, most vaccines and antibody biologics are evaluated by challenging target animals, which is a more direct and accurate manifestation of immune response in animals. The study aims to find the correlation between challenging protections and sera IgY titers in ducks in order to replace the current challenging ducks (in vivo) method with sera IgY titers detection (in vitro) method for efficacy evaluation of bivalent egg yolk antibody against Riemerella anatipestifer infection. In addition, for those anti-RA IgY products failing to evaluate the efficacy with challenge animals, the detection of sera IgY titers may serve as an alternative for efficacy assessment. The 3Rs principles goal could be achieve gradually by reduction in the number of laboratory animals used.
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Abstract
The Polymerase chain reaction (PCR) has been a routine diagnostic technique used in laboratories concerned with the detection of the etiological agents of infectious diseases. When employing this method, one of the considerations is the potential of false positive PCR results. To minimize the risk of producing false-positive results, nucleic acid positive controls are a high risk factor for contamination that need to be taken into account for, in addition to reagents, equipment, and working areas. The aim of this study was to construct a standard DNA construct as a differentiable positive control when using PCR to detect Neospora. The positive control is a plasmid inserted with a fragment composed of an inner sequence of avian influenza virus and the primer annealing regions of Neospora sequences at both ends. When the construct was employed in the Neospora-AI PCR assay, the reaction generates a product 211 bp longer than that of the normal Neospora positive PCR product. In addition, the product can be identified by nested PCR which is used for the routine identification of avian influenza viruses. The same strategy was used to construct a Parainfluenza 3-goatpox differentiable positive control DNA, followed by transcribing it into RNA with the MEGAscript™ SP6 Kit.
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