Fluorescence Filters to Separate Spectra

As an imaging technique, fluorescence microscopy is widely used in medical, biological medical research and many other fields of work.

The technique is based on the detection of fluorescence signals, which are 1000 times weaker than scattered light.

Since scattered light itself is 1000 times weaker than the light used for illuminating the sample, fluorescence microscopy involves detecting a signal that is a million times weaker than the illumination light. Hence, there are high requirements for the filters and dichroic beam splitters, which are employed in fluorescence microscopes.

A detailed description of fluorescence and fluorescence microscopy is available in Carl Zeiss' excellent Microscopy Online Campus website or Nikon's equally valuable resource MicroscopyU.

Measured blocking of a high performance fluorescence filter set

Measured blocking of a high performance fluorescence filter set

Separating Absorption and Emission Spectra

Absorption spectra and emission spectra of many fluorophores (fluorescent dyes) are proximally located and even frequently overlap.

This means that excitation and emission filters must have very steep edges for effectively separating absorption peaks from emission peaks. High out-of-band rejection is needed to detect the weak fluorescence signal among the strong illumination light. The dichroic beamsplitter, at the heart of an epi-fluorescent set-up, mirrors the illumination light into the microscope lens and onto the sample as well as transmits the fluorescence light into the eyepiece or onto the CMOS or CCD camera fitted with the microscope.

Well-designed dichroic beamsplitters possess a wide range of reflection and transmission wavelengths with a distinct in-between transition. Such beamsplitters help subduing the excitation light by a minimum of two orders of magnitude, and enable contrast-rich imaging of cells and other specimens.

Long Experience with Fluorescence Filters

In the late 1960s, Delta Optical Thin Film was the first to suggest the use of interference filters to Carl Zeiss and Leica. In the early 1970s, the company began mass manufacturing of fluorescence filters.

In the early 1990s, Delta Optical was the first to employ the computer-controlled deposition technique. By employing its own synthesis and deposition control software, the company is able to offer coatings made with plasma-assisted e-beam evaporation technology with a quality matching to that achievable by other technologies such as IBS.

Today, Delta Optical is the largest supplier of high performance fluorescence filters to the market-leading manufacturers of fluorescence microscopes, thanks to its decades-long experience in manufacturing the product.

This information has been sourced, reviewed and adapted from materials provided by DELTA Optical Thin Film.

For more information on this source, please visit DELTA Optical Thin Film.


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