Preservation of red cell properties after virucidal phototreatment with dimethylmethylene blue

BACKGROUND: All published reports have described methods for virus photoinactivation which significantly alter red cell (RBC) properties during storage. In order to improve virucidal activity and reduce damage to RBCs, a series of phenothiazine derivatives were either synthesized or purified and screened for bacteriophage inactivation and red cell potassium efflux. One compound, 1,9-dimethylmethylene blue (dimethyl-methylene blue), had superior screening results and was chosen for further characterization. STUDY DESIGN AND METHODS:White cell reduced RBC suspensions (30% hematocrit) were deliberately in oculated with extracellular virus or virus-infected VERO cells, incubated with 4 mu M dimethyl-methylene blue and illuminated with cool-white fluorescent light. Control and treated samples were titered for virus inactivation. In par allel studies, RBC suspensions were exposed to dimethylmethylene blue and light under identical conditions and assayed for in vitro RBC storage properties. RESULTS: Phototreatment of RBC suspensions inactivated >4.4 log(10) of extracellular vesicular stomatitis virus (VSV), >3.0 log(10) of intracellular VSV, >5.0 log(10) of extracellular pseudorabies virus (PRV), >4.8 log(10) of intracellular PRV, >4.7 log(10) of extra-cellular bovine virus diarrhea virus, 5.8 log(10) of bacterio-phage phi 6 and >7 log(10) of bactariophage R17. Encephalo-myocarditis virus, a nonenveloped picornavirus, was resistant to photoinactivation. Virucidal conditions resulted in no detectable IgG binding in 11 of 13 samples, unchanged RBC morphology, normal banding patterns of RBC membrane proteins on SDS PAGE, and unaltered characteristics of 12 of 13 RBC antigens during storage as measured by antibody titrations. In addition, minimal changes were observed in RBC osmotic fragility, lysis, potassium efflux, ATP and 2,3-DPG levels: and the strength of one RBC antigen during storage of phototreated samples compared with controls. CONCLUSION: Dimethylmethylene blue photo-treatment can inactivate several intracellular and extracellular model viruses under conditions which minimally alter RBC properties during 42 day storage at 1-6 degrees C.