Partial purification of relaxin from human seminal plasma.
Human seminal plasma contains approximately 50 ng/ml of relaxin (specific activity = 1.3 ng/mg protein). During preliminary purification, semen plasma was delipidated, rehydrated, and loaded onto small octadecyl silica columns that were eluted with a TEAF/acetonitrile gradient system. Results were monitored by radioimmunoassay. The resultant partially purified human relaxin demonstrated biologic activity in the rat uterine segment bioassay. Nine liters of semen plasma was delipidated, rehydrated in TEAF, and subjected to high-performance liquid chromatography by a step gradient followed by a linear gradient. The active eluate was further purified by ion exchange chromatography. Pooled recovery fractions provided a total of 45.8 micrograms of relaxin. An aliquot flash evaporated and desalted by gel filtration chromatography provided 1.85 micrograms of relaxin in 25.2 mg protein, specific activity 73.4. This material is being used as immunogen in the production of antihuman relaxin antibodies by monoclonal technique. Our procedure represents the first and only successful partial purification of human relaxin to yield sufficient quantity and purity for antibody production.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis; D001681:Biological Assay; D002850:Chromatography, Gel / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D005260:Female; D006801:Humans; D008055:Lipids / Q000302:isolation & purification; D008297:Male; D011863:Radioimmunoassay; D051381:Rats; D011919:Rats, Inbred Strains; D012065:Relaxin / Q000276:immunology / Q000302:isolation & purification* / Q000494:pharmacology; D012661:Semen / Q000032:analysis*; D014590:Uterine Contraction / Q000187:drug effects
D000911:Antibodies, Monoclonal; D008055:Lipids; D012065:Relaxin