Quantitative determination of intracellular, ferritin-associated radioactive iron by high-performance liquid chromatography and immunoprecipitation.
Antibodies raised against ferritin preparations of diverse origin provide an uncertain reagent for quantitation of the ferritin present in specific cell lysates. Utilizing K562 cells, a human leukemic cell line, techniques are described to resolve and to quantitate the ferritin-bound cytosolic iron. Processing the cell lysates by HPLC employing an anion-exchange or hydrophobic interaction column resulted in recovery of a single, ferritin-containing radioactive peak widely separated from the bulk of the non-ferritin-bound iron. Comparison of the yield obtained by chromatography with that by immunoprecipitation confirmed both the specificity and the quantitation of the antibody technique.
Josic
D
D
Mattia
E
E
Ashwell
G
G
van Renswoude
J
J
D001665:Binding Sites; D002460:Cell Line; D011232:Chemical Precipitation; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D003600:Cytosol / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D005293:Ferritins / Q000032:analysis*; D006801:Humans; D007120:Immunochemistry; D007501:Iron / Q000032:analysis*; D007504:Iron Radioisotopes / Q000032:analysis; D004915:Leukemia, Erythroblastic, Acute / Q000032:analysis; D008715:Methionine / Q000378:metabolism
D007504:Iron Radioisotopes; D005293:Ferritins; D008715:Methionine; D007501:Iron