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United States Department of Agriculture

Agricultural Research Service

Title: A Modified Elisa for Detection of Cutaneous Antibody Against Ichthyophthirius

Authors
item Xu, Dehai
item Klesius, Phillip
item Shelby, Richard

Submitted to: Aquaculture America Conference
Publication Type: Abstract Only
Publication Acceptance Date: September 30, 2002
Publication Date: February 18, 2003
Citation: Xu, D., Klesius, P.H., Shelby, R.A. 2003. A modified elisa for detection of cutaneous antibody against ichthyophthirius. Aquaculture America Conference.

Technical Abstract: Fish surviving Ichthyophthirius multifiliis (Ich) infection develop an immune response against this parasite. Enzyme-linked immunosorbent assays (ELISA) currently used to determine the cutaneous anti-Ich antibody response lack sensitivity. This study modified an ELISA assay for detection of low levels of cutaneous antibody against Ich in channel catfish and in water. The modified ELISA used tetramethylbenzidine (TMB) as substrate for horseradish peroxidase (HRP). The modified ELISA required less theront coating protein, shorter coating time for the plate, and a lower concentration of conjugate. The optical density readings using the modified ELISA were approximately 4-fold higher than the ELISA using o-phenylenediamine (OPD) when measuring anti-Ich antibody in the culture fluid from immune skin. The modified ELISA was able to detect the antibody in water samples collected from tanks with Ich immune fish. The ELISA by using OPD failed to detect the antibody even though the water samples were concentrated 40- fold. The results of this study showed that the modified ELISA using TMB as a HRP substrate was a sensitive, quantitative and time saving assay to measure low concentrations of anti-Ich cutaneous antibody excreted from skin of channel catfish immune to Ich.

Last Modified: 10/30/2014
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