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縲€縲€Filter Guide for Various Fluorescent Reagent (Ex/Em)

Fluorescence Reagent Filter Guide (Ex / Em)

This page provides a practical reference for selecting suitable LED wavelengths and emission filters based on excitation (Ex) and emission (Em) spectra of commonly used fluorescent reagents in gel imaging and fluorescence observation.

EXCITATION (EX) EMISSION (EM) LED WAVELENGTH EMISSION FILTER DICHROIC FILTER
Selection Principle:
Choose an LED wavelength that efficiently excites the fluorophore, and combine it with an emission filter that blocks excitation leakage while transmitting fluorescence emission. When excitation and emission spectra are close or overlapping, additional filtering such as a dichroic filter may be required to reduce background glow.

Quick Reference Table

Below is a list of commonly used fluorescent reagents and their excitation / emission characteristics. Click the spectral information button for detailed filter guidance for each reagent.

List of Reagents / Ex-Em Wavelengths / LED Light Source / Filter Combination
Below is a list of commonly used fluorescent reagents and their excitation /
emission characteristics. Click the Spectral Info button for detailed filter guidance for each reagent.
Reagent EX nm EX
(Sub)		 EM nm spectral
info		 Reagent EX
nm		 EX
(Sub)		 EM nm spectral
info
Alexa Fluor 532 532 554 Spectral mCitrine 516 529 Spectral
Cy3 550 570 Spectral Midori Green 490 290 530 Spectral
DsRed 557 579 Spectral SYBR Gold 495 540 Spectral
EtBr 300 518 605 Spectral SYBR Green I 498 522 Spectral
FITC 490 525 Spectral SYPRO Orange 470 570 Spectral
Flamingo 512 271 535 Spectral SYPRO Red 550 300 630 Spectral
Gel Green 250 500 530 Spectral SYPRO Ruby 280 450 620 Spectral
Gel Red 270 510 600 Spectral TagRFP 555 583 Spectral
GFP 488 507 Spectral tdTomato 554 581 Spectral
mCherry 580 610 Spectral YFP 513 527 Spectral
Additional Ex/Em Reagent List Reagent List
Notice: This guide is intended for reference purposes only. It does not guarantee visibility of any specific fluorescence signal. Actual results may vary depending on LED intensity, filter quality, optical configuration and camera specifications.

Symbol Meaning

  • ◎ Most suitable
  • ◎* Most suitable, but an additional long-pass or dichroic filter may be required to reduce background.
  • ○ Suitable
  • △ Usable, but with limited effectiveness compared with ◎ and ○.
  • × Not recommended
Hover over the image to view LED and filter wavelengths
For cellular labeling in fluorescence microscopy and in cell biology.
Retains sulfonic acid which is
directly connected to the fluorescent. Especially bright with rhodamine derivative. The reagent with great
light stability
Thermo Fisher
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Excitation and emission wavelengths are very close, making it difficult to separate fluorescence emission from excitation leakage. A 505 nm LED with an SC-56 emission filter is effective. Additional filtering to block longer
wavelength leakage is recommended to reduce background when used
with transmitted light source.

Hover over the image to view LED and filter wavelengths
In addition to such biological assays as immunocytochemistry, Cyanine
dye fluorescence reagent is often
used for labelling RNA.
Thermo Fisher Scientific
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Excitation and emission wavelengths are close, making it difficult to separate fluorescence emission from excitation leakage. A 505 nm LED with an SC-56 emission filter is effective. Additional filtering to block longer
wavelength leakage is recommended to reduce background when used
with transmitted light source.

Hover over the image to view LED and filter wavelengths
Exhibits strong and stable
expression in bacterial cells
with minimal toxicity.
TAKARA
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Excitation and emission peaks are close, but the spectra are broad enough that separation is relatively easier. Blue, cyan and green LEDs can be used for excitation, and
a red emission filter is effective for separation.

Hover over the image to view LED and filter wavelengths
Ethidium bromide is one of the most widely used reagents for nucleic acid band staining. It is treated as hazardous waste by many organizations and requires appropriate handling and disposal.
Wikipedia
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
EtBr has a strong excitation peak in the UV region around 300 nm and can also be excited in the visible range around 480 to 520 nm.
LED505 / LED530 can be used, but sufficient light intensity and effective filtering are important to reduce background glow.

Hover over the image to view LED and filter wavelengths
A type of fluorochrome widely used for microscopic observation.
Often utilized for bloodstain search, dye tracking in the area of forensic medicine and serology
Wikipedia
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
The excitation peak is around 490 nm, and excitation with a 505 nm LED is possible. However, because excitation and emission wavelengths are close, background-suppression optics may be required. A high-power blue LED with an appropriate emission filter can also be effective.

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Low cost protein detection with simple protocol. High sensitivity and quantitatively enables identification
of protein by mass spectrometry. Compatible also with UV excitation.
BIO-RAD
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Ex 512 nm and LED505 nm can be effective. LED device with background eliminating feature or high power
blue LED & SC52 pair can also be effective since Ex/Em is close.

Hover over the image to view LED and filter wavelengths
Green fluorescent nucleic acid dye designed to replace the highly toxic EtBr. Compare to EtBr, it is ultra-sensitive and very stable, both hydrologically and thermally. Very simple procedures for pre or post staining
Biotium
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm

Hover over the image to view LED and filter wavelengths
Fluorescent nucleic acid dye designed to replace the highly toxic EtBr. Compare to EtBr, it is ultra-sensitive and very stable, both hydrologically and thermally. Very simple procedures for pre or post staining.
Biotium
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Wavelength characteristic is similar to EtBr. Compare to EtBr, it has more abosrption around 520 nm and thus
LED light is effective.

Hover over the image to view LED and filter wavelengths
Enables real time detection without damaging cell. It also demonstrate its function as fusion protein (GFP tag) Widely used as a tool to locate
protein which is taking part in
signal transaction inside the cell.
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Em/Ex peaks are close but Em is spread over to short wavelength
side and Blue LED is effective

Hover over the image to view LED and filter wavelengths
Red fluorescent proteins isolated
from Discosoma sp. Utilizes for fusion expression with target protein, promoter activity monitoring and bacteria expression. Takes short time for expression and stable as well.
TAKARA
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Em/Ex peaks are close but Em is spread over to short wavelength side and Cyan or Green LED can be used. Excitation with Green LED and separation with red filter is effective.

Hover over the image to view LED and filter wavelengths
mCitrine is an empty vector and gene insertion can be achieved by LIC cloning protocol. mCitrine is highly versatile with its compatibility with many other vector as well as usability as single expression vector.
TAKARA
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Ex 510m and 505 nm LED is efficient. However Ex/Em is close and may require LED device with background eliminating features. High power blue LED with SC52 filter is also effective.

Hover over the image to view LED and filter wavelengths
Ideal Nucleic acid staining reagent for prestained method. Low background and high S/N ratio makes safe alternative to the ethidium bromide.
NIPPON Genetics
Ex Effcy Filter evaluation
SC-54 SC-60
470 nm
505 nm
530 nm
Ralatively easy to excite with Blue LED (470m). Easy to separate since Ex/Em peak has some space between them.

Hover over the image to view LED and filter wavelengths
Filter evaluation
Thermo Fisher Scientific
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Relatively easy to excite with Blue
and Cyan LED. Easy to separate since Ex/Em peak has some space between them.

Hover over the image to view LED and filter wavelengths
Asymmetrical cyanine dye used as
a nucleic acid stain in molecular biology. It binds to double helix DNA and can be replacement for EtBr as it is safer to work However any small molecule capable of binding DNA with high affinity is a possible carcinogen, including SYBR Green.
Thermo Fisher Scientific
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Ex is similar to SYBR Gold, however Em is adjacent to Ex and bit harder
to separate. LED 505 with background eliminating features or high power blue LED & SC52 pair is also effective.

Hover over the image to view LED and filter wavelengths
Provides fast, simple, sensitive staining of proteins. Has very little protein-to-protein variation in staining. Enables long lasting
and stable emission. With low background, it requires less time
for cleaning gel. Detection with immunoreagent is also possible
with western blotting.
Thermo Fisher Scientific
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Ex is spread over to wide range
and both Blue and Cyan LED are
highly effective for excitation.
With enough space between Ex&Em,
it is relatively easy to separate too

Hover over the image to view LED and filter wavelengths
For cellular labeling in fluorescence microscopy and in cell biology.
Retains sulfonic acid which is
directly connected to the fluorescent. Especially bright with rhodamine derivative. The reagent with great
light stability

Thermo Fisher Scientific
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm

Hover over the image to view LED and filter wavelengths
For cellular labeling in fluorescence microscopy and in cell biology.
Retains sulfonic acid which is
directly connected to the fluorescent. Especially bright with rhodamine derivative. The reagent with great
light stability
Thermo Fisher Scientific
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
With wide space between the peaks
of Ex & Em, it is considered easy reagent for separation. Blue LED
is most effective for excitation.
Cyan LED is also effective.

Hover over the image to view LED and filter wavelengths
Generated from the wild-type RFP from sea anemone entacmaea quadricolor. It is brightest monomeric red fluorescent protein available. Mammalian cells transiently transfected with TagRFP expression vectors produce bright fluorescence
in 10-12 hrs after transfection.
Evrogen JSC
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Cyan, or Green LED is effective for excitation. Em is too close with Green, and background may appear with SC-56 but sould be able to
separate with SC-60 since Em is
spread over to long wavelength side.

Hover over the image to view LED and filter wavelengths
Exceptionally bright red fluorescent protein (2.5x brighter than EGFP) It is easily detected as deep as 1 cm below the surface, and ideal for live animal imaging studies. High cohesion stability make it suitable to produce protein N-, C- fusion protein too.
Takara Bio
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Wavelength characteristics is similar to Tag RFP and Em/Ex are close.
Cyan LED & shortpass filters pair or Green LED & SC-60 filter pair should be used to eliminate background.

Hover over the image to view LED and filter wavelengths
YFP is synthetic non-aequorea fluorescent proteins that can be amplified/cut by various PCR and cloned into any other expression vector of choice. These vectors can also be used as expression vectors or as positive controls and allow monitoring of inducible protein expression.
Cosmo Bio
Ex Effcy Filter evaluation
SC-56 SC-60
470 nm
505 nm *
530 nm
Ex peak at 512 nm and can be excited with 505 nm LED. However Ex/Em is close and may require LED device with background eliminating features. High power blue LED with SC52 filter can also be effective.
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