An analytical method employing gas chromatography is presented for assessing the concentrations of the sulfur mustard hydrolysis product thiodiglycol (TDG) in cutaneous in vivo microdialysates. The use of a pulsed flame photometric detector allows for selective detection of the analyte following solvent exchange and derivatization with heptafluorobutyric anhydride. Quantitative assessment is performed using thiodipropanol (TDP) as a surrogate internal standard. A linear relationship and a very significant correlation (r2 = 0.9982) between the ratio of TDG and TDP concentrations and the ratio of the square root of peak heights is demonstrated. The suitability of the analytical method is verified by the evaluation of blank in vivo microdialysates spiked with known amounts of TDG. The limit of detection in microdialysates is 0.200 nmol/mL (24.4 ng/mL) and the limit of quantitation was 0.364 nmol/mL (44.4 ng/mL). The presented method provides selective, sensitive, rapid, and high-throughput analysis of microdialysates containing TDG, providing an efficient alternative for high-performance liquid chromatography and capillary electrophoresis techniques.