Abstract: Objective To investigate the influence of blood collection tubes with different additives on serum or plasma polypeptide detection and confirm the optimal vacuum tube for the test of serum polypeptides. Methods Vein blood samples were taken from 6 volunteers, including 2 males with age of(28±2) years and 4 females with age of(30±3) years. And the samples from each case were divided and transferred to 6 types of blood collection tubes. The polypeptides were extracted with weak cationic magnetic beads and detected with clin-tof platform. The polypeptide peaks were analyzed and compared by BioExplorer and SPSS 22.0. Results In total, 108 out of 117 peptide peaks showed differences among the six type of tubes(P<0.05), and 95 showed significant differences(P<0.01). The number of different peptide peaks among plasma tubes was significantly higher than that among serum tubes. The highest proportion of normality peptide peaks(110) appeared in the sixth group(coagulation promoting tube containing separation glue), where the mean of intensive peaks number(peak intensity>300) was the largest and the standard deviation was the smallest(27.7±1.97). Four factors(the whole peak intensity, the peak numbers with normal distribution, the average of intensive peak numbers, and intensive peaks' intensity) were investigated with factor analysis to rank tube types, and the sixth group got the highest score. Conclusion The coagulation promoting tube containing separation glue is determined as the most optimal blood collection tube for MALDI-TOF MS.