Resumo

The simplest and most widely used procedure for the detection of endotoxin is the LAL. As in many enzymatic reactions, LAL test demands optimal pH, temperature and time of incubation. Slight changes in these parameters may result in an increased frequency of false-positive and false-negative results, which will affect the uncertainty of LAL test. The aim of this paper is to evaluate how changes in the pH, temperature and time of incubation will affect the uncertainty of LAL test. Aliquots of 100 μL of solutions having concentrations of 0.5, 0.25, 0.125, 0.06 and 0.03 EU mL^-1 were transferred to apyrgenic tubes containing 100 μL of LAL. To test the effect of pH 1 μL of 0.1 M HCl or 1 μL of 0.1 M NaOH were added to the tubes before incubation. To test the effect of temperature some tubes were incubated for 60 minutes in a water-bath at 36, 37 or 38ºC. To test the effect of time of incubation some tubes were incubated at 37ºC for 58, 60 or 62 minutes. After incubation, each tube was inverted 180º to verify the presence or absence of gel formation. The frequencies of true and false responses obtained in extreme conditions of temperature, time of incubation and pH were compared to those obtained in nominal conditions. Slight differences in pH increase frequencies of false responses in 5-fold. Temperature and time of incubation seem to affect less the LAL test. The increased frequencies of false responses observed in extreme conditions will affect the uncertainty of LAL test. Slight differences in pH increase the estimated uncertainty to 7.6%. The estimated uncertainty obtained in extreme conditions of temperature and time of incubation were similar to that obtained in nominal conditions. Slight changes in pH may affect significantly the estimated uncertainty of LAL test. Slight differences in temperature and time of incubation seem to affect less the LAL test.