O. Kofanova, C. Bellora, R. A. Quesada, A. Bulla, S. Panadero-Fajardo, M. Keipes, K. Shea, M. Stone, P. Lescuyer, F. Betsou.
Journal of Immunological Methods. November 2018; https://doi.org/10.1016/j.jim.2018.11.012 [Epub ahead of print]
Uncontrolled preanalytical variables can reduce the accuracy and reproducibility of downstream analytical results from peripheral blood mononuclear cells (PBMCs).
PBMCs were isolated from EDTA and citrate-anticoagulated blood samples, obtained from healthy subjects and patients with inflammatory and infectious conditions. PBMC-derived RNA samples were examined for gene expression changes induced by extended blood pre-centrifugation delays at 4 °C and RT. We used Taqman RTqPCR to evaluate the combination of two target genes for their “diagnostic performance” in identifying EDTA and citrate-anticoagulated PBMC samples with extended pre-centrifugation times.
We established the PBMC preanalytical score, a gene expression metric to assess the PBMC quality related to the pre-centrifugation delay at room temperature for different anticoagulants. The PBMC preanalytical score measurement can identify:
- EDTA PBMC samples or RNA extracted from these PBMCs with RT precentrifugation times >48 h with 98% sensitivity and 87% specificity at a cutoff of 57.
- citrate PBMC samples or RNA extracted from these PBMCs with RT precentrifugation times of >48 h with 92% sensitivity and 84% specificity at a cutoff of 348.
The proposed PBMC preanalytical score may enable objective PBMC sample qualification for downstream applications, which may be influenced by blood precentrifugation delays.