Establishment of reference measurement procedure and reference material for Treponema pallidum

Yanmin Lin ^{1 2}, Jiayi Yang ², Xia Wang ², Jingya Yang ^{1 3}, Lianhua Dong ²

Abstract

Stem cell preparations, as a new type of biotherapeutic product, require strict quality control for cell safety. Screening of stem cell donors and culture-related blood products, including Treponema pallidum detection, is essential to lower the risk of infectious disease transmission via stem cell products. In this study, a reference measurement procedure was established based on digital PCR. Homogeneous reference material carrying the tpp47 gene of Treponema pallidum was prepared and characterized. Two digital PCR assays presented satisfactory linearity across five orders of magnitude. Both assays had a limit of quantification of 57 copies/reaction, while their limits of detection were 9.69 copies/reaction and 9.59 copies/reaction respectively. Consistent quantitative results were obtained using the duplex digital PCR assay. The certified value with expanded uncertainty of the reference material was determined as (2.21 ± 0.22) × 10⁶ copies/μL. Assay validation in simulated stem cell matrix showed no obvious interference on detection accuracy. This reference material provides accurate absolute quantification of target gene copy number, and helps enhance the reliability of Treponema pallidum testing in stem cell preparation manufacture and clinical diagnosis.