Characterisation of an antiserum and development of an ELISA for glutathione peroxidase

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Abstract
Sheep red blood cells were fractionated by ion exchange and gel filtration chromatography to yield glutathione peroxidase approximately 99% pure. An antiserum against glutathione peroxidase was raised in the rabbit. The antiserum has been shown to cross-react with both bovine and human glutathione peroxidase by double diffusion. An enzyme linked immunosorbent assay has been developed for glutathione peroxidase which detected 6.15×10−5 IU of the enzyme. The antiserum has also been shown to be effective in the detection of glutathione peroxidase immobolised on strips of nitrocellulose, subsequent to sodium dodecyl sulphate polyacrylamide gel electrophoresis, by second antibody conjugate. Avidin-biotin was also used to detect nitrocellulose immobolised enzyme. These techniques provide an alternative highly sensitive and specific means of assaying glutathione peroxidase which is not dependent on the lability of enzymatic activity nor the chemical specificity of the assay.