An automatic multidimensional chromatography system for purification of human uterine progesterone receptor and induction of polyclonal antibodies.
This paper reports on the synthesis of Org2058-bonded microparticulate silicas and their use in affinity chromatography as the first step for the purification of human progesterone receptor. The development of microprocessor-controlled instruments allows all the various steps to be performed automatically. The various steps used for the purification of human progesterone receptor were carried out with the FPLC system: affinity chromatography, desalting of eluate on Sephadex G-25, anion-exchange chromatography using a Mono Q column. With this procedure the receptor was purified approx. 10,000-fold within 24 h. The yield of receptor was generally 85-95%. Investigations with induced anti-progesterone receptor antibodies obtained after the fourth immunization show their immunoreactive behaviour towards progesterone receptor in crude cytosol, which was proved by sucrose density gradient centrifugation and by gel filtration on the FPLC system using a Sepharose 12 column. This implies that progesterone receptor was efficiently purified by our purification procedure.
Heubner
A
A
Belovsky
O
O
Grill
H J
HJ
Pollow
K
K
D000906:Antibodies / Q000032:analysis*; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D003900:Desoxycorticosterone / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D008024:Ligands; D011282:Pregnenediones / Q000378:metabolism; D011980:Receptors, Progesterone / Q000276:immunology / Q000302:isolation & purification*; D014599:Uterus / Q000032:analysis*
D000906:Antibodies; D008024:Ligands; D011282:Pregnenediones; D011980:Receptors, Progesterone; C020175:16 alpha-ethyl-21-hydroxy-19-nor-4-pregnene-3,20-dione; D003900:Desoxycorticosterone