Simultaneous autoradiography and line immunoelectrophoresis (ARLIE): a novel combination to identify de novo protein synthesis by pregnancy tissues.
A novel combination of two conventional techniques (autoradiography, AR and line immunoelectrophoresis, LIE; ARLIE) for identification of specific proteins synthesized de novo by explants is described. The incorporation rate of [35S]-methionine was linear in proteins derived from cytosol fractions and supernatants of first trimester human trophoblast and gestational endometrium for up to 18 h. SDS-PAGE analysis of these fractions provided further evidence of the protein synthesis and secretion by the tissue explants. The ARLIE system was evaluated by investigating the synthesis and secretion of five test proteins (PP12, PP14, hPL, FA-1 and FA-2) by trophoblast and gestational endometrium. The synthesis P12 and PP14 could be demonstrated by gestational endometrium only. Similarly the synthesis of hPL could be demonstrated by the trophoblast alone. The synthesis of the fetal proteins (FA-1 and FA-2) could not be demonstrated by either tissue. The control procedure, Protein A assisted immunoprecipitation, yielded similar results for PP14 but not hPL. This novel combination (ARLIE) provides a simple technique with which to study the de novo synthesis of several proteins simultaneously which is independent of the subclass and species of origin of antibodies.
Fay
T N
TN
Academic Unit Obstetrics and Gynaecology, London Hospital, UK.
Price
K
K
Teisner
B
B
Jacobs
I
I
Grudzinskas
J G
JG
D000653:Amniotic Fluid / Q000378:metabolism; D001345:Autoradiography; D002352:Carrier Proteins / Q000096:biosynthesis; D002824:Chorionic Villi / Q000378:metabolism*; D004717:Endometrium / Q000378:metabolism*; D005260:Female; D005326:Fetal Proteins / Q000096:biosynthesis*; D000071816:Glycodelin; D006023:Glycoproteins; D006801:Humans; D007122:Immunoelectrophoresis; D018970:Insulin-Like Growth Factor Binding Protein 1; D018969:Insulin-Like Growth Factor Binding Proteins; D010928:Placental Lactogen / Q000096:biosynthesis*; D011247:Pregnancy; D011257:Pregnancy Proteins / Q000096:biosynthesis*; D013997:Time Factors
D002352:Carrier Proteins; D005326:Fetal Proteins; D000071816:Glycodelin; D006023:Glycoproteins; C032231:IGFBP1 protein, human; D018970:Insulin-Like Growth Factor Binding Protein 1; D018969:Insulin-Like Growth Factor Binding Proteins; C045246:PAEP protein, human; D011257:Pregnancy Proteins; D010928:Placental Lactogen