Jesse Sambrano will be giving a PhD student research seminar on Thursday May 4 3:00-4:00pm in the Jerry Shaw Conference Room (Goddard Hall Room 200).
“Evaluating integrin activation with time-resolved flow cytometry”
Abstract: Flow cytometers that measure fluorescence excited state decay times are emerging with capabilities that permit the resolution of Förster resonance energy transfer (FRET) events in cells. In this work, we optimize time-resolved flow cytometry to measure FRET during activation of integrin dimers expressed on the surface of human leukocyte cells. Such integrins exhibit switchblade-like movements that alter proximal distances between donor and acceptor fluorophores. Moreover, conformational extension adjusts to a bent conformation when resting, thereby permitting quantification by FRET. With laser modulation and phase-sensitive flow cytometry both fluorescence intensity and fluorescence lifetime are measured across large cell populations (n = 10,000) in order to quantify integrin conformational changes. Results indicate FRET donor fluorophores bound to the end of resting integrins had shortened fluorescence lifetimes (~2 ns) owing to energy transfer with nearby acceptor fluorophores bound to the cell membrane, coinciding with an inactive integrin state. Longer fluorescence lifetimes (3 ns) were measured when integrins lengthened indicating activation, or loss of FRET. The ability to quantify activation states of integrins is important for a range of cell adhesion studies which affect cancer research and drug discovery. Measurement of fluorescence lifetimes aids in FRET quantification and is a significant step towards the interpretation of distributions of integrin states on a single cell basis.
Bio: Jesse Sambrano began his fifth year at NMSU in the department of Chemical & Materials PhD program in August under the direction of Dr. Jessica Houston. He received his Bachelor Science Degree from the University of Texas at Paso in Chemistry and Physics in 2011. Jesse is a former RISE to the Postdoctorate Scholar and a recent recipient of the Ruth L. Kirschstein National Research Service F31 Award, funded by the National Institutes of Health. Jesse has presented research on a national and international setting, winning numerous travel awards and outstanding poster presentation awards. Jesse’s research interests include flow cytometry instrumentation and mechanobiology. Jesse will transition to a Post-Doc position at the University of New Mexico in 2018 and ultimately seek a faculty position to continue his work in academia.