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Terahertz absorption spectroscopy based on attenuated total reflection (ATR) from a microfluidic sample cell was designed and implemented to detect gene mutations leading to Huntington's disease (HD). The self‐developed compact ATR microfluidic system was employed to detect two groups of base‐repeated DNA molecules combined with a terahertz time‐domain spectrometer in a marker‐free manner. The first group featured different repetition patterns of oligonucleotide fragments, and the second group included the HD gene. For the oligonucleotides of different repetition patterns, there were significant differences among the three oligonucleotides with three repeats of the double bases, which could be unambiguously classified and identified; For the HD gene, it was found that the magnitude of the terahertz absorption coefficients of the four oligonucleotide solutions was, in ascending order, CAG‐4 < CAG‐16 < CAG‐32 < CAG‐40 (the numbers are the repeat times of the CAG base segment, with 40 repeats belonging to the HD gene), when the concentration of oligonucleotide was 1 mg/mL. Principal component analysis result indicated that the spectral differences of the four oligonucleotide solutions with different CAG repeat times were statistically significant and clearly distinguishable. These results demonstrate the potential of terahertz spectroscopy as a noninvasive, unmarked, fast and low‐cost assay for gene diagnosis and clinical disease detection. [ABSTRACT FROM AUTHOR]