UC San Diego

Chronic lymphocytic leukemia (CLL) is a form of blood cancer, in which DNA is mutated to produce abnormal B cell lymphocytes. CLL induces the cells to differentially regulate the expression of specific miRNA, producing miRNA with specific cancer related mutations. These abnormal cells frequently rupture and release the miRNA into the bloodstream, adding to the cell free circulating (CFC) miRNA. Measuring the ratio of miRNA expression between B cell lymphocytes and plasma can determine the extent of CLL within a particular patient. While the goal of the project is to determine how QIAGEN kits and dielectrophoresis (DEP) affect the detected expression levels of CLL-related miRNA biomarkers, the goal of this work is to determine consistency of detected miRNA levels with a spike-in into samples of CLL cells and plasma. This includes determining general RNA loss at the extraction step and subsequent miRNA loss and expression measurements at the qRTPCR step. Another goal is finding the dielectrophoretic parameters that will allow for separation of CFC RNA and B cells from whole blood and from each other. In terms of findings, there was a more significant loss of miRNA at the qRTPCR step than there was loss of RNA at the extraction step. Plasma was also found to have reduced miRNA signals and copy number compared to extractions from cells, due to endogenous RNase activity. With a functional limit of 103 Hz, dielectrophoretic models were able unable to predict frequencies separating any of the analytes with respect to plasma and B cells