Chimaeric protein A/protein G and protein G/alkaline phosphatase as reporter molecules.
The IgG binding domains of staphylococcal protein A and streptococcal protein G were expressed as a chimaera using the pGEX vector which has been advocated because its fusion proteins tend to be soluble and easily isolated on immobilised glutathione. This chimaera was soluble and abundant (yield = 18 mg/l of bacterial culture) and was tested by double diffusion in agarose and by ELISA. It was found to bind IgG of all species that either parent could bind. It was superior to protein A or protein G in binding mouse Ig. A chimaera of protein G and alkaline phosphatase was also constructed and found to be soluble and abundant (yield = 20 mg/l of bacterial culture). This protein could be used as a secondary reagent in ELISA at 5 micrograms/ml for human, rabbit and mouse and at 25 micrograms/ml for sheep.
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National Animal Quarantine Institute, Qingdao, China.
Lew
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AM
D000469:Alkaline Phosphatase / Q000235:genetics* / Q000276:immunology; D000818:Animals; D001483:Base Sequence; D002678:Chimera / Q000276:immunology*; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay; D005822:Genetic Vectors; D006801:Humans; D005779:Immunodiffusion; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000235:genetics* / Q000276:immunology; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins; D013205:Staphylococcal Protein A / Q000235:genetics* / Q000276:immunology
C029152:G-substrate; D009419:Nerve Tissue Proteins; D011994:Recombinant Proteins; D013205:Staphylococcal Protein A; D000469:Alkaline Phosphatase