Mid-IR semiconductor lasers of two wavelength bands, 5.4 and 9.6 μm, are applied to measure aqueous glucose concentration ranging from 0 to 500 mg/dL with Intralipid® emulsion (0 to 8%) added as a fat simulator. The absorption coefficient µa is found linear with respect to glucose and Intralipid® concentrations, and their specific absorption coefficients are obtained via linear regression. These coefficients are subsequently used to infer the concentrations and compare with known values. The objective is to evaluate the method accuracy. Glucose concentration is determined within ±21 mg/dL with 90% confidence and ±32 mg/dL with 99% confidence, using <1-mJ laser energy. It is limited by the apparatus mechanical error and not the photometric system noise. The expected uncertainties due to photometric noise are ±6 and ±9 mg/dL with 90 and 99% confidence, respectively. The uncertainty is fully accounted for by the system intrinsic errors, allowing rigorous inference of the confidence level. Intralipid® is found to have no measurable effect on glucose determination. Further analysis suggests that a few mid-IR wavelengths may be sufficient, and that the laser technique offers advantages with regard to accuracy, speed, and sample volume, which can be small, ~0.4×10-7 mL for applications such as microfluidic or microbioarray monitoring.