Microplate assay of ascorbic acid in aqueous humor with bicinchoninic acid

An inexpensive microplate assay method was developed to determine ascorbic acid in human aqueous humor samples. The method is based on the selective oxidation of ascorbic acid at alkaline pH and determination of ascorbic acid and other reducing substances in samples with bicinchoninic acid/CuSO4 before and after the alkaline treatment. The two-point measurement eliminates the effect of interfering substances, such as glucose, uric acid and glutathione, which are stable at alkaline pH. The advantages of the method are that it requires small sample volumes and allows handling of a large number of samples simultaneously.