The physiochemical and immunological characteristics of rat lymph binding protein for 25-hydroxyvitamin D3.
The binding proteins for 25-hydroxyvitamin D3(25-OH-D3) in rat lymph and plasma were purified to apparent homogeneity as determined by sodium dodecyl sulfate disc gel electrophoresis. The purification procedures included the following techniques: gel filtration on Sephadex G-100 and G-200; affinity chromatography on Blue Sepharose CL-6B; ion-exchange chromatography on DEAE-Sepharose CL-6B; chromatofocussing on a Mono P column. Both proteins from lymph and plasma were eluted with the same retention time from an isoelectric column at a pH of approximately 4.53 and showed nearly identical data on the analysis of amino acid composition. When specific anti-lymph 25-OH-D3-binding protein antiserum was prepared in a rabbit, and Ouchterlony immunodiffusion was performed, the same precipitate line was observed on both lymph and plasma binding proteins. These results strongly suggest that a common transport protein exists in both rat lymph and plasma and circulates in the blood-lymph system.