Simplified Determination of Blood Adenosine Triphosphate Using the Firefly System.

DENOSINE TRIPHOSPHATE (ATP) appears to play a critical role in maintaining the integrity of the red cell. In the “Storage lesion,” the disappearance of high energy bonds represented largely by ATP correlates well with the loss of viability of erythrocytes upon reinfusion into the circulation.1’2 In early studies, erythrocyte ATP was estimated by determining “easily hydrolysable” phosphorus.1 ’4 ATP is not the only compound from which phosphorus may be split tinder the conditions employed, and such determinations must be considered, at best, as good approximations. More recently, chromatographic separation of red cell filtrates, either on paper or in columns, has been employed in estimating erythrocyte ATP.5 2 Trichioracetic acid or perchloric acid extracts of erythrocytes have been used and the quantity of ATP has been determined by phosphorus analysis and by measurement of ultraviolet absorption. All of these technics are extremely time-consuming and their accuracy and precision is impaired by the many manipulations required. Several enzymatic technics have also been employed. 2t Although these are less time-consuming than chromatographic technics, they are still quite complex. The relatively recently introduced firefly lantern extract technic for assay of tissue ATP22 is probably the simplest and most sensitive method available. Simon et al.21 have used this method to determine ATP in perchloric acid filtrates made from erythrocytes, but no details were given in their publication. Other investigators may have also adapted this method for the determination of erythrocyte ATP, but we are not aware that any technic has been published. Under proper experimental conditions, we have found that firefly lantern extract may be employed as a basis for a relatively specific, highly reproducible and rapid technic for red cell ATP assay. In the present investigation the variables which influence determination of ATP by the use of firefly lantern extract have been studied and an attempt has been made to elucidate the optimal conditions for carrying out red cell ATP assays in this system.