Imaging molecular order in live cells using fluorescence polarization.
Instantaneous PolScope is a microscope designed to image concentration, alignmnet, and orientation of molecules in a single snapshot. The method directly images molecular alignment and orientation, which remain challenging to measure in live cells with fluorescence super-resolution.
The microscope, its calibration, and its use for imaging actin network and septin filaments are reported here:
Mehta, S.B., McQuilken, M., La Riviere, P., Occhipinti, P., Verma, A., Oldenbourg, R., Gladfelter, A.S., and Tani, T. (2016). Dissection of molecular assembly dynamics by tracking orientation and position of single molecules in live cells. Proceedings of the National Academy of Sciences 113, E6352–E6361.
This repository contains the MATLAB code used for calibration and analysis of data produced by such a microscope.
It includes the following:
- MATLAB class (RTFluorPol) that represents the microscope
- Hardware calibration is stored as properties of the class.
- Calibration and pixel-level analysis procedures are written as methods of the class.
- MATLAB class (instantPolGUI) that provides a GUI to calibrate the microscope (i.e., an RTFluorPol object) and analyze movies.
Representative time-series data (alongwith relevant calibration) on retrograde flow of actin network at a leading edge of a migrating cell can be downloaded here.