Abstract 3226: Multiphoton Flow Cytometry for High-Throughput Analysis of Intrinsic Biomarkers to Guide Cellular Transplantation for Cardiac Disease
Cellular therapies for cardiac disease could be enhanced by an instrument capable of determining viability and/or potency of cells without the use of extrinsic markers and sorting cells based on that identification. We hypothesized that a multiphoton fluorescence excitation (MPFE)-based flow instrument could be used to excite endogenous fluorophores as intrinsic biomarkers, penetrate deep into large cellular entities and subsequently purify populations based on these biomarkers. To this end we have merged flow cytometry technology with MPFE to create a proof of concept prototype Multiphoton Flow Cytometer (MPFC). Using MPFE we can detect differences in the intensity of intrinsic fluorescence between dead and live cells and between stem cells and mature cells. Additionally, we have developed and fully validated a flow cell and software that can be used in conjunction with the MPFE for comprehensive flow cytometry control and analysis. In recent studies, we can detect intrinsic fluorescence from NADH of embyroid bodies in flow mode on the MPFC consistent with static mode analyses (average aiu, 91.8±35.0 and 86.8±36.7 respectively, p=0.84). The large error reflects the large variation in intrinsic fluorescence intensities within a given EB population (Figure 1⇓). This distribution supports the notion that viability and differentiation state might be discerned based exclusively on intrinsic fluorescence intensity. Thus, given its capacity for high-throughput, noninvasive characterization and purification of cellular entities, the MPFC system could prove a powerful tool to guide clinical cellular transplantation for cardiovascular disease.