Fluorescence Microscopy in vivo

Fluorescence microscopy is an essential tool for modern biological experimentation. It allows for non-ambiguous detection for molecules of interest while significantly improving the effective resolving power of diffraction limited optics. The power of fluorescence microscopy is most pronounced in live cells in which live molecular interactions in the physiological conditions can be revealed. We are interested in looking at DNA mismatch repair in vivo to complement our single molecule in vitro techniques such as smFRET and AFM, to understand how DNA mismatch repair works in a live cell context. We constructed a four color fluorescence microscope setup that allows us to extract and overlay four colors simultaneously).

IN VIVO FLUORESCENCE.

Schematic of our microscope set up using a quad-view beam splitter.

(left) Quad-view split images of four fluorescence channels. (right) Example overlay of four images taken simultaneously on the quadview.

(left) Examples of in vivo imaging of MMR foci using stochastic photobleaching.  An example image obtained via epifluorescence is shown. (right). Two separate stochastic photobleaching profiles of the individual foci circled in yellow.