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The objective of this study was to discriminate thyroid and parathyroid tissues using Raman spectroscopy combined with an improved support vector machine (SVM) algorithm. In thyroid surgery, there is a risk of inadvertently removing the parathyroid glands. At present, there is a lack of research on using Raman spectroscopy to discriminate parathyroid and thyroid tissues. In this article, samples were obtained from 43 individuals with thyroid and parathyroid tissues for Raman spectroscopy analysis. This study employed partial least squares (PLS) to reduce dimensions of data, and three optimization algorithms are used to improve the classification accuracy of SVM algorithm model in spectral analysis. The results show that PLS‐GA‐SVM algorithm has higher diagnostic accuracy and better reliability. The sensitivity of this algorithm is 94.67% and the accuracy is 94.44%. It can be concluded that Raman spectroscopy combined with the PLS‐GA‐SVM diagnostic algorithm has significant potential for discriminating thyroid and parathyroid tissues. [ABSTRACT FROM AUTHOR]