Purification of Vitamin D Binding Protein and Its Role in the Uptake of 25-Hydroxyvitamin D by Cultured Kidney Cells (Mdck Cells, Tissue Culture, Dbp)
This thesis reports the isolation of vitamin D binding protein (DBP) and its use in studies investigating the uptake of 25(OH)D(,3) by kidney cell lines. DBP was isolated from pig plasma in three steps: chromatography on DEAE cellulose and DEAE Sephadex, followed by chromatofocusing. The procedure represents a shorter purification scheme than those previously reported. Antibodies to DBP were produced in rabbits and coupled to cyanogen bromide-activated Sepharose 4B. With this antibody affinity column, DBP was isolated fron pig plasma to a purity similar to that in the initial purification in a one-step procedure. Holo- and apo-DBP did not co-chromatograph upon chromatofocusing, which was used to separate the two forms. Holo-DBP was used as the source of 25(OH)D(,3) to study its uptake by kidney cells.
University of Illinois - Urbana-Champaign
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University of Illinois at Urbana-Champaign