Characterization of an antigen from the myelogenous leukemia cell line K-562.
A protein was solubilized from the myelogenous leukemia cell line K-562 WITh 3 M KCl that specifically inhibited the antibody-dependent, complement-mediated cytolysis of 51Cr-labeled K-562 cells by a monkey antiserum to K-562. When the crude 3 M KCl extract uas fractionated with ammonium sulfate, an eightfold increase in specific activity (U inhibition/mg protein) resulted. This purified fraction migrated as a single protein band after polyacrylamide gel electrophoresis (PGE) with no detectable carbohydrate or lipid. The molecular weight of the denatured protein determined by sodium dodecyl sulfate-PGE was 77,000, similar to that of the native protein (80,000) determined by Sephadex exclusion chromatography. The protein was stable at pH 6-8, with an apparent isoelectric point between pH 5 and 6. In addition to being irreversibly denatured at pH 5 or less, it was unstable at osmolarities below 0.25 M (NaCl). It was denatured at temperatures of 56 degrees C or above. Normal human peripheral blood leukocytes were extracted similarly with 3 M KCl and fractionated with ammonium sulfate. Neither the crude preparation nor any fraction purified as described for the specific antigen inhibited the cytolytic assay, which indicated at least a quantitative lack of the protein on the surfaces of normal leukocytes.
D000645:Ammonium Sulfate; D000818:Animals; D000951:Antigens, Neoplasm / Q000302:isolation & purification*; D002460:Cell Line; D005591:Chemical Fractionation; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007942:Leukemia, Experimental / Q000276:immunology; D007951:Leukemia, Myeloid / Q000276:immunology*; D009994:Osmolar Concentration
D000951:Antigens, Neoplasm; D000939:Epitopes; D000645:Ammonium Sulfate