A single solution for non-linear calibration in flow injection spectrophotometry: Kinetic determination of total protein in blood serum

Abstract

Several procedures for the determination of clinical–biochemical parameters are based on commercial kits supplied with only one standard solution because linear calibration graphs are involved. When these procedures are carried out in flow analysis non-linearity of the analytical graph may be observed due to time restrictions. Since at least three standard solutions are then needed, direct implementation of commercial kits in flow injection analysers can be hindered. To overcome these drawbacks, a computer controlled flow injection system exploiting concentration gradients and requiring only one standard solution is proposed for non-linear calibration. Limitations due to non-linearity of the analytical graph were overcome using a dye solution to calibrate the concentration gradient of the injected sample and standard solution. As an application, the spectrophotometric kinetic determination of total protein in blood serum based on commercial kits was selected. The system handled 60 samples per hour and required 17 μl and 1.0 ml of sample and biuret reagent, respectively, per determination. No baseline drift was observed during 8 h operating periods. Within the 40–80 g l −1 range, the overall relative standard deviation of results was estimated as 3.0% ( n = 5). The paired t-test was used and no statistic differences at the 95% probability level between the results for the reference and the proposed procedures was found.