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What is widefield microscopy?

Similar to brightfield microscopy, widefield fluorescence microscopy illuminates the sample from below. The difference is in the light. Instead of a white light, fluorescence microscopy uses light of frequencies that can excite fluorescent molecules associated to your sample. With the widefield fluorescence microscopy technique, you illuminate the whole sample with light of a specific wavelength. You can then visualize or record the emitted light through the microscope’s eyepieces or camera.

Figure: two DNA molecules trapped using four optical traps. DNA bridging proteins XRCC4 (green) and XLF (red) can be seen both individually and as a DNA bridging complex (orange) using correlated widefield fluorescence microscopy. Data courtesy of Prof. Erwin Peterman and Prof. Gijs Wuite at the VU University Amsterdam.

How does a widefield microscope work?

  1. A light source emits an excitation light, which passes through an excitation filter that only allows light of specific wavelengths to pass. The excitation light is reflected off a dichroic mirror, a filter that selectively lets certain colors pass while reflecting the rest. 
  2. Once the excitation light hits the sample, it temporarily excites fluorescent molecules associated to the sample to a higher energy state. Once they relax back to their ground-state, light of a longer wavelength is emitted.
  3. The objective lens collects some of the emitted light. The dichroic mirror transmits the rest through an emission filter. 
  4. The emission filter blocks excitation light and transmits emission light to the eyepiece or camera.

Explore other imaging techniques

Confocal Fluorescence

Multicolor confocal, perfect for visualizing biological processes in solution.

STED Super Resolution

The perfect choice for performing experiments in highly crowded environments, offering unprecedented resolution (< 35 nm).

TIRF Fluorescence

Suitable for visualization on surfaces as it eliminates background fluorescence outside the focal plane.

IRM (label-free)

IRM allows you to visualize microtubules without the need for fluorescence labeling.

Our solution

The C-Trap® Optical Tweezers – Fluorescence & Label-free Microscopy is the world’s first instrument that allows simultaneous manipulation and visualization of single-molecule interactions in real time. It combines high resolution optical tweezers, fluorescence and label-free microscopy and an advanced microfluidics system in a truly integrated and correlated solution.

The C-Trap offers you a fast workflow to seamlessly catch and manipulate single molecules. The instrument measures their structural changes or interactions while you visualize them in teal time with high spatial and temporal resolution, ultimately offering you a complete and detailed picture of biomolecular properties and interactions.

Curious to learn more?
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