Clinical evaluation of a new rapid heparin assay using the dye azure A

At the present time, heparin assays are based on biologic measures of activity. They are time-consuming, difficult to reproduce, and require special devices. We have developed a colorimetric assay for chemical heparin in plasma based on the metachromasia of azure A in the presence of heparin. One milliliter of plasma is added to 1 ml of 0.08 percent azure A, vortex mixed, and read at 620 nm in a spectrophotometer. To evaluate the clinical utility of this assay, we compared it to two biologic assays of heparin effect, activated PTT and activated clotting time, and to heparin levels determined by protamine titration in 113 samples from 28 patients undergoing cardiopulmonary bypass. The activated PTT in 94 of 113 samples was greater than 600 seconds, making this test not useful due to the time required and the lack of an end point. The activated clotting time of 80 samples was less than 600 seconds. The azure A measurement of heparin concentration correlated well in this group (correlation coefficient of 0.88, p less than or equal to 0.0001). Protamine titration determinations also had excellent correlation with azure A measurements in the 111 samples tested (correlation coefficient of 0.85, p less than or equal to 0.0001). The azure A assay correlates well with the standard measures of both heparin activity and heparin concentration. It is more rapid, simple, and less expensive than either of these, and it does not depend on a bioassay of coagulation as an end point. Chemical heparin measurements with azure A may be more useful clinically than the biologic assays in determining the reversal dose of protamine.