Isolation, characterization and immunocytochemical localization of bovine trophoblast protein-1.
Bovine trophoblast protein-1 (bTP-1) was isolated to 90% purity from culture medium of Day 18-20 conceptuses incubated in vitro (in the presence of L-[3H]leucine) by a combination of Sephacryl S-200 gel filtration chromatography and O-(diethylaminoethyl) (DEAE) anion-exchange high-performance liquid chromatography (DEAE-HPLC). The radiolabeled protein had an Mr of 21,200 +/- 800 by one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and had three isoelectric variants (pI 5.7-6.5) by two-dimensional SDS-PAGE. DEAE-HPLC-enriched bTP-1 cross-reacted with anti-o TP-1 serum on Western blots. A monospecific antiserum against bTP-1 was produced by excising the bTP-1 polypeptide band from preparative SDS-PAGE gels. Antiserum reacted with a single polypeptide with an Mr of 21,200 as determined by Western blotting of SDS-PAGE-separated conceptus medium proteins and by immunoprecipitation from L-[35S]methionine-labeled medium proteins followed by SDS-PAGE and autoradiography. Bovine trophoblast protein-1 was localized by immunocytochemistry in the cytoplasm of both mono- and binuclear trophectoderm cells of Day 20 bovine conceptuses, indicating that it is a product of the trophoblast.
Lifsey
B J
BJ
Department of Animal Science, University of Tennessee, Knoxville 37901.
Baumbach
G A
GA
Godkin
J D
JD
D000818:Animals; D015153:Blotting, Western; D002417:Cattle; D003470:Culture Media / Q000032:analysis; D004586:Electrophoresis / Q000379:methods; D004622:Embryo, Mammalian / Q000032:analysis*; D005260:Female; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D007370:Interferon Type I; D011233:Precipitin Tests; D011247:Pregnancy; D011257:Pregnancy Proteins / Q000302:isolation & purification* / Q000378:metabolism; D014327:Trophoblasts / Q000378:metabolism
D003470:Culture Media; D007370:Interferon Type I; D011257:Pregnancy Proteins; C020947:interferon tau