Fieldable flow injection analysis system with 1,1′-oxalyldiimidazole chemiluminescence detection capable of quantifying acetylcholine

We developed a highly sensitive biosensor for quantifying acetylcholine (ACh) using flow injection analysis system with 1,1'-oxalyldiimidazole chemiluminescence (ODI-CL) detection designed based on the principle of liquid core waveguide. ACh in Tris-HCl (pH 8.5) was incubated with the mixture of 1.0 U/ml acetylcholinesterase, 0.5 U/ml choline oxidase, 0.04 U/ml horseradish peroxidase, and 1.0 mu M Amp lex Red in PBS (pH 7.4) for 15 min at room temperature. The concentration of resorufin formed from the consecutive enzyme reactions was proportional to the concentration of ACh in analytical sample. The dynamic range of linear calibration curve (y = 12444x + 11617, R-2 = 0.998) for the quantification of ACh using the biosensor with ODI CL detection was 0.7 similar to 11.3 mu M. The limit of detection (LOD = background noise + 3 sigma) of the biosensor was as low as 0.14 mu M. Based on the results of recovery test and linearity study, finally, we confirmed that FIA system with ODI CL detection is accurate, precise, and reproducible.