Riboflavin carrier protein from spotted owlet egg yolk has been isolated and it was assayed using spectrophotometry. The spectral intensity (1.687) of Rf solution showed small changes with a gradually increase in Riboflavin Carrier protein Binding capacity initially 10μl to 40μl and decreased binding capacity later on between 50 to 60 μl concentration. Isolated protein (RCP) binding affinity to Riboflavin was calculated at different concentrations and the results are communicated