In vitro inactivation of human immunodeficiency virus by ascorbic acid

In vitro inactivation of cell-free human immunodeficiency virus (CFHIV) was investigated by mixing replication-competent virions with aliquots of a culture medium (RPMI) containing increasing amounts (62.5-500 micrograms/ml) of ascorbic acid (AA) at pH7. Similarly, mixtures of CFHIV and 500 micrograms/ml AA in whole blood (WB) and leukocyte depleted blood (LDB) were made; control mixtures containing either CFHIV or AA alone in each experiment were included. After holding the mixtures for 3 h at 4 degrees C, the tubes containing WB and LDB mixtures were centrifuged to remove the blood cells. The respective supernatants, including the control aliquots, were layered over 0.5 x 10(6) MT2 cells in quadruplicate wells in microtitre plates. After 1 h of incubation at 37 degrees C in an atmosphere of 5.0% carbon dioxide to permit contact of viable virions, the fluid in each well was replaced with RPMI containing 20% fetal bovine serum (FBS). The incubation was then continued at 37 degrees C for 5 days. On the basis of (1) absence of syncytia formation, (2) 100% viability of MT2 cells as compared with the cell controls, (3) absence of p24 antigen in the culture supernates, and (4) absence of HIV DNA in MT2 cells, we conclude that 500 micrograms/ml AA, in (a) RPMI, (b) WB, or (c) LDB, inactivated CFHIV in vitro. Furthermore, we determined that addition of 500 micrograms/ml AA to platelet concentrates did not adversely affect the platelet function tests during 5 days of storage at room temperature. These data warrant further work to evaluate the mechanism of CFHIV inactivation by treatment of blood products with AA.