Abstract: Protein related tumor biomarkers have been widely used in clinically assisted diagnosis and disease surveillance because they can indicate the physiological state, pathological process and post-drug reaction of cancer patients. Therefore, it is of great significance to accurately measure tumor biomarkers. However, due to the wide dynamic range of body fluid, the existence of high abundant proteins and diverse post-translation modifications pose a great challenge to the accurate quantification of protein related tumor biomarkers. Liquid chromatography-mass spectrometry (LC-MS) combines the high-efficiency separation capability of chromatography technology and the high sensitivity and high throughput detection capability of mass spectrometry technology, which has become a powerful tool for the discovery, verification and analysis of biomarkers. Currently, LC-MS has been widely applied in the field of quantitative research on protein related tumor biomarkers. In this paper, research progress in the quantitative methods for protein related tumor biomarkers in recent years was summarized from the sample pretreatment approach and mass spectrometry scanning strategy. In complex biological samples, protein related tumor biomarkers are usually at ultra-low expression level, which make it difficult to directly analyze protein related tumor biomarkers by mass spectrometry. Therefore, the development of appropriate sample pretreatment methods (e.g. high abundance protein removal, target protein enrichment, quantitative peptide collection, isotope labelling, etc.) to reduce the complexity of biological samples before quantification analysis is a key step to ensure accurate measurement results. In addition, primary MS analysis can only provide limited biological information, which can not effectively distinguish the substances with similar mass-to-charge ratio, and can not meet the needs of protein related tumor biomarkers quantification. Thus, multi-stage mass spectrometry scanning strategy (such as multiple reaction monitoring, parallel reaction monitoring, etc.) has become an effective method for protein quantitative analysis. Although the rapid development of mass spectrometry technology in resolution, mass accuracy and analysis speed provides opportunities for accurate protein analysis, it is still difficult to accurately quantify low-abundant proteins for the high complexity of biological samples in body fluid. Moreover, due to the limitation of the automation for both mass spectrometer device and sample pretreatment, quantitative analysis of protein related tumor biomarkers based on LC-MS is still not applied to the clinical detection on a large scale. Therefore, it is necessary to continuously develop efficient and rapid sample pretreatment approaches, accurate and reliable mass spectrometry scanning strategies and the easy automation technology. In the future, more reliable detection methods for protein biomarkers may make disease screening, diagnosis and treatment more precise and more efficient.