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Anita Pottekat et al.
Liquid biopsy continues to gain traction as a minimally invasive method to monitor biomarkers associated with malignancy and metastasis. A simple blood draw can reveal key biomarkers such as circulating tumor cells (CTCs) and cell-free DNA (cfDNA). However, these analytes are present in low quantities and are prone to rapid degradation and contamination, which presents challenges for the detection of rare CTCs and cfDNA targets.
Higher molecular weight genomic DNA from poorly stabilized leukocytes can contaminate plasma, compromising liquid biopsy assay results by increasing the total amount of extracted DNA while diluting rare cfDNA targets. Preservation of CTCs and cfDNA over time is thus essential for the accuracy of downstream tests, and preanalytical variables that impact plasma volume, cfDNA yield and quality, and cell recovery in ex vivo blood must be understood.
Here, we identified several preanalytical variables, including type of blood collection tube, storage time and conditions post-phlebotomy, and centrifugation conditions for plasma fractionation. We then carried out a series of studies to better understand the interplay between these preanalytical variables and plasma recovery, cell recovery, and cfDNA preservation in healthy donor blood, contrived samples, and patient samples.
We found that choice of blood collection tube, in particular, has a significant impact on preservation of plasma volume, CTCs, cfDNA over time, particularly under real-world stress conditions including temperature, excursion, sample shipping, and storage time post-draw.
The work presented here demonstrates the importance of assessing preanalytical variables for the preservation of cancer biomarkers in whole blood without compromising sample integrity. This will help researchers make informed decisions on how best to stabilize their analytes of interest without introducing un-intended variables due to blood tube selection, shipping and storage conditions, and plasma isolation conditions.