Genomic Profiling of Circulating Plasma RNA for the Analysis of Cancer

the other assayed selectins. A few measurements do not fit any discernable pattern, notably plasma sVCAM-1 on day 3 at 4 and 21 °C; plasma sE-selectin on day 1 at 4 °C and day 3 at all temperatures, whole blood sE-selectin at 4 °C on day 1 and after freeze-thaw cycle 4; and plasma CRP on day 1 at 4 °C and after freeze-thaw cycle 3. These aberrant values were likely analytic artifacts, because removal of values 4 SDs from the mean or adjustment for run-to-run variability did not change the results qualitatively. The large CRP difference observed after freeze-thaw cycle 3 but not after cycles 4 or 5 suggests assay error or possibly CRP release from LDL and complement factors. It has been demonstrated that a portion of systemic CRP is bound to cholesterol in modified LDL particles (17) and to different complement factors (18). Interaction of CRP with cholesterol and complement factors might hamper the detection of CRP in the assay used. No data were available for sE-selectin, but it is possible that some sE-selectin is bound to different circulating leukocyte types or microvesicles derived from endothelial cells. Freeze-thaw cycles might release these bound forms of sE-selectin, leading to an increase in detectable sE-selectin. Possible effects of residual platelets in the plasma on marker values obtained are not addressed in our study. sVCAM-1, sICAM-1, and sE-selectin, however, are derived mainly from endothelial cells and not from platelets. Although the presence of platelets in EDTA plasma might contribute to the absolute amounts of these markers, it does not influence their stability. In contrast, sP-selectin is mainly derived from platelets, a characteristic that may explain the variance observed in sP-selectin at different time and temperature points. Our results show that sP-selectin is unstable under all storage conditions and requires immediate assay. The other cardiovascular risk markers evaluated were stable when stored in whole blood samples for several days at room temperature. sVCAM-1, sICAM-1, and CRP were stable at 4 and 21 °C in plasma or whole blood for 5 days and sE-selectin for 2 days. These findings have important implications for clinical studies measuring these markers, reducing the need for immediate transfer of samples to a laboratory for processing and analysis.