Introduction to TIRF Microscopy
Total Internal Reflection Fluorescence (TIRF) microscopy restricts excitation to a thin layer near the sample surface, often less than 100 nanometers, dramatically improving signal-to-noise ratio for surface-proximal imaging. That performance depends on filters and dichroics that can handle excitation energy re-entering the emission path at high angles, something standard fluorescence optics aren't built for.
This app note covers how evanescent field excitation works, what through-the-lens TIRF demands of optical filters, and how Chroma's substrates, coatings, and cube hardware are built to meet those demands.
Get The App Note
Read it straight through for the full picture of how TIRF optics work, or keep it as a reference when specifying filters for a TIRF setup.
Download the PDF to save, print, or share with your team.
What's Inside
How evanescent wave excitation restricts fluorescence to the sample surface
The shift from prism-based TIRF to through-the-lens systems
Why through-the-lens TIRF places extreme blocking demands on emission filters, exceeding OD 8
Reflected wavefront distortion requirements for TIRF dichroics, and why they're tighter than confocal
How Chroma's substrates, coatings, and TIRF-optimized filter cubes are built to meet those requirements
Representative TIRF filter set configurations and their spectral performance
Who This Is For
Written for researchers and imaging core staff working with TIRF, single-molecule imaging, or other surface-proximal fluorescence techniques, particularly those specifying or troubleshooting filter sets for a through-the-lens TIRF system.
Explore Further
Spectra Viewer
Plot the transmission curves for representative TIRF filter sets against your fluorochrome and laser lines.
TIRF Filter Sets
Browse Chroma's catalog of TIRF-optimized filter sets and cube assemblies.
Guide to Fluorescence and Optical Filters
For the underlying fundamentals, including excitation, emission, and how filter sets work as a system, before diving into TIRF-specific requirements.
If you don’t see what you are looking for, please contact us. We have a large inventory of filters and a knowledgeable staff ready to help you design and build a filter to meet your specifications.
