Objective：The UPLC fingerprint of Mistletoe was established and the similarity evaluation was carried out. Combined with cluster analysis and principal component analysis, chemometric analysis methods were used to provide reference for the quality consistency evaluation of Mistletoe medicinal materials. Methods: The chromatographic column was Waters ACQUITY UPLC HSS T3 chromatographic column (2.1×100 mm, 1.8 μm). The mobile phase was acetonitrile ( A ) -0.5 % acetic acid water ( B ) with gradient elution. The detection wavelength was 254 nm. Column temperature 30℃ ; the injection volume was 3 μL.《The Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine ( 2012 Edition )》was used to establish the fingerprint of Mistletoe, and the common peaks were calibrated and the similarity was evaluated. Cluster analysis and principal component analysis were performed on 14 batches of Mistletoe by SPSS 26.0. Results: The methodological investigation results of the fingerprint of Mistletoe were good, and 14 common peaks were identified. By comparing with the reference substance, it was identified that the No.4 peak was Syringin and the No.9 peak was homoeriodictyol-7-O-β-D-glucoside. The similarity evaluation results showed that the similarity of 11 batches of samples was above 0.950, and the similarity of the other 3 batches was less than 0.950. The samples were clustered into three categories by cluster analysis, S5 was a separate category, S12, S7, S9 were a category, and the rest of the samples were clustered into one category. The cumulative variance contribution rate of the four principal component factors obtained by principal component analysis was 88.732%. The principal component factors of S5, S10, S3 and S11 samples were in the top four of the comprehensive scores, and the overall quality was good. Conclusion: The fingerprint method established in this study has good separation, precision and stability, short analysis time and strong specificity. Combined with similarity evaluation, cluster analysis and principal component analysis, it can provide a theoretical basis for the quality control of Mistletoe medicinal materials.