Effects of Sample Processing Procedures on PBMC Gene Expression
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Date
June 30, 2016
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Citation
Ray, Ashley. 2016. Effects of Sample Processing Procedures on PBMC Gene Expression. Master's thesis, Harvard Extension School.
Abstract
This study investigated the effects that sample processing had on PBMC gene expression profiles. Specifically, two vacutainer types were used in this study; sodium heparin CPT vacutainers and “standard” sodium heparin vacutainers as well as two freezing devices; CoolCell and programmable rate-controlled freezer. Biogen’s clinical trials collect whole blood in sodium heparin CPT vacutainers and cryopreserve PBMCs using a CoolCell device because both are more convenient and economical for the site. The results revealed that when RNA from freshly isolated PBMCs from both vacutainer types were compared 21 DEGs were identified. Regardless of the vacutainer type, when comparing RNA from freshly isolated PBMCs to RNA from thawed cryopreserved cells using a CoolCell device ~50 DEGs were identified but only ~30 DEGs were found when RNA from freshly isolated PBMCs were compared to RNA from rate-control frozen PBMCs. Additional testing needs to be performed to better appreciate when to use each vacutainer. My results suggest that a rate-controlled freezer causes less DEGs than a CoolCell and that about 74% of the cryopreservation-related genes (17/23) were common regardless of vacutainer type or freezing device used. The data from this project will assist in the design of scientifically-based “Best Practices” needed for appropriate sample processing of PBMCs.
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