Simple and Sensitive Spectrofluorometric Method for the Determination of Protein Using an Europium-thenoyltrifluoroacetone Probe

The fluorescence intensity of the europium (Eu3+)-thenoyltrifluoroacetone (TTA) complex can be remarkably enhanced by human serum albumin (HSA) in a Britton–Robinson buffer solution. Based on this fact, a simple, rapid, and sensitive method has been developed for the determination of proteins at the nanogram level by fluorescence spectroscopy. Under the optimum conditions, the enhanced fluorescence intensity is proportional to the concentration of HSA. The linear ranges for HSA are 0 – 5.1 and 5.1 – 44.4 µg ml–1 and the limit of detection is 20.7 ng ml–1. Moreover, there is very little interference from common inorganic ions and other coexisting compounds. The binding site number and the binding constant of Eu3+-TTA used as a fluorescence probe to HSA were calculated by using the Rosenthal graphic method. This method has been applied to the determination of total protein in human serum samples. The results are good agreement with data obtained by clinical physicians.