Large scale purification of intralumenal alkaline phosphatase from calf intestine by immunosorbent affinity chromatography.
Electrophoretically and immunologically homogeneous alkaline phosphatase (AP) can be obtained by a simple and rapid two-step procedure. Step 1: Immunosorbent affinity chromatography using immobilized polyclonal anti-AP-antibodies and elution of the bound enzyme by triethylamine solution, pH 11.4. After affinity chromatography the specific activity of the extracted crude material increases from 3.5 to 793 IU/mg. Step 2: Final purification to a specific activity of 1524 IU/mg by DEAE-cellulose ion exchange chromatography. One investigator is able to purify 100 mg AP within 3 days. The overall recovery is 65%. Methods for characterization and selection of anti-AP-antiserum using immunoinhibition of AP are described.
D000469:Alkaline Phosphatase / Q000302:isolation & purification*; D000818:Animals; D002417:Cattle; D002846:Chromatography, Affinity / Q000379:methods; D002852:Chromatography, Ion Exchange; D007106:Immune Sera / Q000276:immunology; D005779:Immunodiffusion; D007163:Immunosorbent Techniques; D007421:Intestine, Small / Q000201:enzymology*
D007106:Immune Sera; D000469:Alkaline Phosphatase