Combined MT-TIRF

Combining single-molecule techniques with fluorescence microscopy has attracted much interest because it allows the correlation of mechanical measurements with directly visualized DNA:protein interactions. In particular, combination with total internal reflection fluorescence microscopy (TIRF) is advantageous because of the high signal-to-noise ratio this technique achieves. This, however, requires stretching long DNA molecules across the surface of the flow cell to maximize polymer exposure to the excitation light. We have developed a laterally magnetic pulling module to stretch DNA molecules at a constant force and a home-built device to rotate capillaries with mrad precision.

The combination of MT with TIRF microscopy allows us to correlate biological function with the presence of a protein or a component of a protein complex.

We have built two combined lateral MT and TIRF setups equipped with ultra-sensitive EM-CCD cameras and fast CMOS cameras for bead tracking.

MT-TIRF-related papers of the group

Madariaga-Marcos et al. Nanoscale Mar 1;10(9):4579-4590 (2018). doi: 10.1039/c7nr07344e. Force determination in lateral magnetic tweezers combined with TIRF microscopy. LINK