The aim of the study is to explore the possibility of detecting metastatic lymph nodes by FTIR spectra during the surgery of thyroid cancer. The FTIR spectra of 20 metastatic lymph nodes and 69 non-metastatic ones were collected via an ATR (attenuated total reflectance) probe. For each spectrum, 28 variables of 13 bands including peak positions and relative intensities were measured. The variables of metastatic lymph node were compared to those of non-metastatic ones using standard statistic methods. The results indicated that the FTIR spectra of metastatic lymph nodes were significantly different from non-metastatic ones in the bands related to protein,lipid,nucleic acid and carbohydrate. (1) Variations of bands related to protein: The relative intensity ratios of I3 280/I1 460, I1 640/I1 460 and I1 546/I1 460 increased significantly (P＜0.05); (2) Variations of bands related to lipid: The relative intensity ratios of I1 743/I1 460 decreased significantly (P＜0.05). On the contrary, I1 400/I1 460 increased significantly (P＜0.05); (3) The peak positions of 1 165 and 1 120 cm-1 which were all assigned to carbohydrate shifted toward higher wave number (P＜0.05). The relative intensity ratio of I1 165/I1 460 decreased significantly (P＜0.05); (4) The peak positions of 1 085 cm-1 related to nucleic acid shifted to the lower wave number (P＜0.05); (5) Other undetermined bands: The relative intensity ratios of I1 303/I1 460 and I1 303/I1 240 increased significantly (P＜0.05). FTIR spectroscopy could be a reliable and practicable method for metastatic lymph nodes diagnosis in the operation of thyroid cancer. It could be applied in detecting metastatic lymph nodes which can not be determined by palpation in surgery.