Factors affacting trehalose loading in red blood cells before freeze-drying

Object To study the factors that influence trehalose loading,and to discuss the rlue for loading RBCs with trehalose across to the intracellular trehalose.Methods RBCs were mixed with trehalose and incubated at different temperatures,incubation durations and different trehalose concentrations,and anthrone reaction was used to determine the amount of the intracellular trehalose,meanwhile we evaluated hemolysis of loaded RBCs.Results With extracellular trehalose concentration of 1 000 mmol/L,incubation duration9 h and the temperature37 ℃,trehalose loading in RBCs was correlated with the temperature,duration and extracellular trehalose concentration.When extracellular trehalose concentration were 0,200,400,600,800,and 1 000 mmol/L,trehalose loading in RBCs were 0,10.03,14.5,41.7,55.3 and 71.6 mmol/L,respectively;When the incubation duration were 0,1,3,5,7,9 h,trehalose loading in RBCs were 0,5.73,6.11,55.7and 61.2 mmol/L,respectively.For these factors,the statistical results showed that the temperature had the most effects on trehalose loading.Conclusion Fresh RBCs incubated with 800 mmol/L trhalose at 37℃ for a time span of 7 h results in the highest intracellular trehalose concentration of about 52 mmol/L.This study provides an important first step in the long-term preservation of RBCs.