To quantitatively estimate the influence of cuttlebone on the target strength (TS) of golden cuttlefish, the cuttlebone was carefully extracted from 19 live cuttlefish caught using traps in the inshore waters around Geojedo, Korea, in early May 2010 and the TS was measured using split-beam echosounders (Simrad ES60 and EY500). The TS-length relationships for the cuttlefish (before the extraction of cuttlebone, Fish Aquat Sci. 17:361-7, 2014) and the corresponding cuttlebone were compared. The cuttlebone length (L b ) ranged from 151 to 195 mm (mean L b = 168.3 mm) and the mass (W b ) ranged from 29.3 to 53.2 g (mean W b = 38.8 g). The mean TS values at 70 and 120 kHz were −33.60 dB (std = 1.12 dB) and −32.24 dB (std = 1.87 dB), respectively. The mean TS values of cuttlebone were 0.19 dB and 0.04 dB lower than those of cuttlefish at 70 and 120 kHz, respectively. For 70 and 120 kHz combined, the mean TS value of cuttlebone was −32.87 dB, 0.11 dB lower than that of cuttlefish (−32.76 dB). On the other hand, the mean TS value of cuttlebone predicted by the regression (TS b = 24.86 log 10 L b -4.86 log 10 λ -22.58, r 2 = 0.85, N = 38, P < 0.01) was −33.10 dB, 0.04 dB lower than that of cuttlefish predicted by the regression (TS c = 24.62 log 10 L c -4.62 log 10 λ -22.64, r 2 = 0.85, N = 38, P < 0.01). That is, the contribution of cuttlebone to the cuttlefish TS determined by the measured results was slightly greater than that by the predicted results. These results suggest that cuttlebone is responsible for the TS of cuttlefish, and the contribution is estimated to be at least 99 % of the total echo strength.