*See the Specs tab for additional, product-specific information.
These excitation, emission, and dichroic filters are designed specifically for use in fluorescence imaging applications. The Ø25 mm excitation and emission filters are mounted in black anodized housings that are 5 mm and 3.5 mm thick, respectively, while the 25.2 mm x 35.6 mm dichroic filters are unmounted. Nine types of filters are available to accommodate key wavelength ranges for many common fluorophores (or their alternatives): BFP, CFP, WGFP, GFP, FITC, YFP, TRITC, CY3.5, and TXRED.
Filter Design These filters are manufactured to high-performance optical specifications and designed for durability. They are produced with multiple dielectric layers deposited on a high-precision, fused silica substrate. The substrate is ground and polished to ensure that the highest possible image quality is maintained. These hard-coated optics produce filter layers that are more dense than those obtained from electron beam deposition techniques. The dense filter layers reduce water absorption and greatly enhance durability, stability, and performance of the filter. Each filter layer is monitored during growth to ensure minimal deviation from design specification thickness, ensuring overall high-quality filter performance. All filters conform to MIL-STD-810F and MIL-C-48497A environmental standards.
Specification
Excitation Filters
Emission Filters
Dichroic Filters
Substrate Material
Fused Silica
Size
Ø25 mm (Ø21 mm Clear Aperture)
25.2 mm x 35.6 mm
Dimension Tolerance
±0.1 mm
Angle of Incidence
0o ± 5o
45o ± 1.5o
Thickness
5.0 ± 0.1 mm
3.5 ± 0.1 mm
1.05 ± 0.05 mm
Surface Quality
60-40 Scratch-Dig
Fluorophore
Item #
CWL*
FWHM
Item #
CWL*
FWHM
Item #
Refl. Band
Trans. Band
BFP
MF390-18
390 nm
18 nm
MF460-60
460 nm
60 nm
MD416
360-407 nm
425-575 nm
CFP
MF434-17
434 nm
17 nm
MF479-40
479 nm
40 nm
MD453
423-445 nm
460-610 nm
WGFP
MF445-45
445 nm
45 nm
MF510-42
510 nm
42 nm
MD480
415-470 nm
490-720 nm
GFP
MF469-35
469 nm
35 nm
MF525-39
525 nm
39 nm
MD498
452-490 nm
505-800 nm
FITC
MF475-35
475 nm
35 nm
MF530-43
530 nm
43 nm
MD499
470-490 nm
508-675 nm
YFP
MF497-16
497 nm
16 nm
MF535-22
535 nm
22 nm
MD515
490-510 nm
520-700 nm
TRITC
MF542-20
542 nm
20 nm
MF620-52
620 nm
52 nm
MD568
525-556 nm
580-650 nm
CY3.5
MF565-24
565 nm
24 nm
MD588
533-580 nm
595-800 nm
TXRED
MF559-34
559 nm
34 nm
MF630-69
630 nm
69 nm
*Center Wavelength
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Filters for Fluorescence Microscopy
Fluorophores A fluorophore is a molecule or portion of a molecule that is capable of producing fluorescence. When light of the appropriate frequency necessary to excite a molecule from its ground state to an excited state is present, excitation will occur. However, once in an excited state, the molecule will be unstable. After some short period of time (typically 10-15 to 10-9 s), a photon will be released, thereby enabling the molecule to return to a lower energy state. The emitted radiation will be at a longer wavelength (lower energy) than the absorbed radiation due to the loss of energy through various mechanisms such as vibrations, sound, and thermal energy.
A single fluorophore can be continually excited unless it is destroyed by photobleaching (i.e. the nonreversible destruction of a fluorophore due to photon-induced chemical damage or covalent modification). The average number of excitation and emission cycles that a particular fluorophore can undergo prior to photobleaching depends on its molecular structure and the local environment; some fluorophores bleach quickly after emitting only a few photons while others are far more robust and can undergo thousands or even millions of cycles before bleaching occurs. Fluorophores used for confocal applications are specifically chosen because they are resistant to photobleaching.
Filters for Fluorescence Microscopy The experimental setup to the right shows the typical filters used for epi-fluorescence microscopy, a form of microscopy in which both the excitation and emission light travel through the microscope objective. By carefully choosing the appropriate filters and mirrors for a given application, the signal to noise ratio can be maximized. As shown in the schematic to the right, three types of filters are used to maximize the fluorescence signal while minimizing the unwanted radiation. Each optical element is discussed below.
Excitation Filter The excitation filter only allows a narrow band of wavlengths to pass through it. For example, as stated in the table under the Specs Tab and shown in the figure below, the bandpass region corresponding to greater than 85% transmission for Alexa 488 is 430 – 470 nm; incident radiation outside of this range is either partially (for regions near the transmission region) or totally (for regions further from the bandpass region) blocked by the filter.
Dichroic Mirror As shown in the experimental setup near the top of this page, the dichroic mirror of a confocal microscope is used to separate the excitation and emission light paths. The transition wavelength value associated with each mirror indicates the wavelength that corresponds to 50% transmission. For example, as shown in the transmission versus wavelength plot below, the transition wavelength for Alexa 488 is ~510 nm.
Dichroic mirrors are designed to reflect light whose wavelength is below a specific value (i.e. the transition wavelength) while permitting all other wavelengths to pass through it unaltered. The Specs Tab provides information on wavelength ranges corresponding to more than 85% reflectance and transmission for each type of dichroic mirror.
By placing one of these mirrors into the experimental setup at 45o with respect to the incident radiation, the excitation radiation (shown in blue in the experimental setup) is reflected off of the surface of the dichroic mirror and directed towards the specimen and microscope objective, while the fluorescence emanating from the specimen (shown in red in the experimental setup) passes through the mirror to the detection system.
Although the dichroic mirror plays a crucial role in the fluorescence microscope, they are not perfect when it comes to blocking unwanted light; typically, ~90% of the light at wavelengths below the transition wavelength value are reflected and ~90% of the light at wavelengths above this value are transmitted by the dichroic mirror. Hence, some of the excitation light can be transmitted through the dichroic mirror along with the longer wavelength fluorescence emitted by the sample. To prevent this unwanted light from reaching the detection system, an emission filter is used.
Emission Filter An emission filter serves the purpose of allowing the desirable fluorescence from the sample to reach the detector while blocking unwanted traces of excitation light. Like the excitation filter, this filter only allows a narrow band of wavelengths to pass through it. For example, as stated in the table under the Specs Tab and shown in the figure to the right, the bandpass region corresponding to greater than 85% transmission for Alexa 488 is 515-535 nm; incident radiation outside of this range is either partially (for regions near the transmission region) or totally (for regions further from the bandpass region) blocked by the filter.
These excitation fluorescence imaging filters, which can be used with a fast filter wheel to construct a custom microscopy setup, are specifically designed to be used in microscopy and imaging applications. Each Ø25 mm filter is deposited on a 5 mm thick round substrate and mounted in a black anodized housing. These filters provide excellent transmission of the desired excitation wavelength (>90%), with a sharp spectral cutoff and low transmission at other wavelengths (<0.001%). These filters may be mounted into our FW103 Fast Filter Wheel to allow rapid wavelength changes.
These emission fluorescence imaging filters, which can be used with a fast filter wheel to construct a custom microscopy setup, are specifically designed to be used in microscopy and imaging applications. Each Ø25 mm filter is deposited on a 5 mm thick round substrate and mounted in a black anodized housing. These filters provide excellent transmission of the desired emission wavelength (>90%), with a sharp spectral cutoff and low transmission at other wavelengths (<0.001%). These filters may be mounted into our FW103 Fast Filter Wheel to allow rapid wavelength changes.
Thorlabs' 25.2 x 35.6 mm Dichroic Filters are designed to separate light of different wavelengths. When light is incident on the filter at a 45° angle with respect to the normal, the excitation light and its associated back reflection are reflected while the longer wavelength fluorescence signal is transmitted. These unmounted filters have a small mark on the edge to indicate the suface with the dichroic coating. If your application would benefit from a round, mounted dichroic filter, consider our round Dichroic Filters.
Includes One Excitation, One Emission, and One Dichroic Filter
A 10% Savings Over Buying the Individual Filters
Since standard fluorescence imaging applications generally incorporate three different filters (i.e., one excitation, one emission, and one dichroic filter) to maximize the signal-to-noise ratio, Thorlabs offers these filters as a set at a savings over purchasing them separately.
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Includes One Excitation, One Emission, and One Dichroic Filter
