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This study demonstrates that the luciferin of the firefly squid <i>Watasenia scintillans</i>, which generally reacts with <i>Watasenia</i> luciferase, reacted with human albumin to emit light in proportion to the albumin concentration. The luminescence showed a peak wavelength at 540 nm and was eliminated by heat or protease treatment. We used urine samples collected from patients with diabetes to quantify urinary albumin concentration, which is essential for the early diagnosis of diabetic nephropathy. Consequently, we were able to measure urinary albumin concentrations by precipitating urinary proteins with acetone before the reaction with luciferin. A correlation was found with the result of the immunoturbidimetric method; however, the <i>Watasenia</i> luciferin method tended to produce lower albumin concentrations. This may be because the <i>Watasenia</i> luciferin reacts with only intact albumin. Therefore, the quantification method using <i>Watasenia</i> luciferin is a new principle of urinary albumin measurement that differs from already established methods such as immunoturbidimetry and high-performance liquid chromatography.