Abstract: Objective · To establish a quality control system for serum antibodies and provide a quantitative standard for the quality control of serum autoantibody-based biomarker study. Methods · Sera from 50 healthy people were taken to prepare the samples and aliquoted. The aliquoted sera were stored at -80 ℃, -20 ℃, 4 ℃, 37 ℃, and 95 ℃ for a period of time. Samples were collected at different time points. The samples were incubated with Ph.D.-12, a phage display random peptide library. The phage-IgG complex was enriched through immunoprecipitation with protein G-coated magnetic beads. By sequencing the coding sequences of the displayed peptides, the sequence and frequency of the serum antibody-enriched peptides were determined. Using the frequencies of each enriched peptide, Shannon entropies were calculated for each sample. Shannon entropy was applied as an indicator to evaluate the quality of the serum autoantibodies. Results · With the increase of the treating temperature, the Shannon entropies of the sera gradually decreased, while there was no significant difference between the Shannon entropies of -20 ℃ and -80 ℃ . The Shannon entropy reached the lowest value after being treated at 95 ℃ for 10 min. At the same temperature, the Shannon entropies of the sera were inversely proportional to the length of the treating periods. Conclusion · It is practically applicable to decipher the profile of the serum antibody binding peptides, through the combination of a phage display random peptide library and next-generation sequencing. Shannon entropy can be calculated using the frequencies of each enriched peptide, and applied as an indicator to judge the overall quality of the serum antibodies.

Key words: serum, quality control, antibody, phage display, next generation sequencing, Shannon entropy