Overview

Excitation filters are always recommended when using pE-Series LED illumination systems for fluorescence microscopy. Successful fluorescence microscopy depends on matching filters with the source of illumination and the stain to be excited. These filters are commonly included within filter cubes in the microscope.

optical_imgBenefits of excitation filters

CoolLED specialize in LED illumination systems for microscopy. We know that to get the best results from your microscope you need to match the right filters to your illumination system and samples.

Although LEDs produce a narrow spectral peak of light, there are often regions of shorter and longer wavelength illumination which may lead to increased background in the image or interfere with the excitation of another stain if no excitation filter is used.

When selecting an LED illumination system which is based on a single, or few wavelength peaks, it is essential that care is taken to match these with the excitation filter in the microscope. Alternatively, a new filter can be fitted which is matched to the illumination system and the stain.

Broad spectrum illumination systems are less sensitive to this optimisation requirement but it is still beneficial to check the filters. With modular LED systems, CoolLED’s wide range of LED wavelengths (LAMs) means that an LED wavelength can be specified which matches with the existing filters in the microscope.

Our products will work with most filter sets out there, but for optimum results we have made some suggestions below. There is never a definite right answer as depending on the demands of your setup you may want more brightness, more contrast or to satisfy some other requirement.

Below are suggested filter sets from Chroma and Semrock for popular fluorophores. In most cases performance is similar, but those looking for particular brightness or contrast are urged to examine the manufacturer’s specifications further or ask CoolLED for help.

 

Single Band Sets pE-300 Series pE-4000 pE-340fura
Chroma Semrock LED Chroma Semrock Chroma Semrock
DAPI 39000 SB (UV @ 365) LED- DAPI-
B- SB (UV@365)
385 (Ch.A) 49028 LED-DAPI-B 49028 LED-DAPI-B
DAPI 49028 MB (UV @ 400) LED-DAPI-
B-MB (UV@400)
CFP 49013 LED-CFP-A 435 (Ch.A) 49001 LED-CFP-A 49013 LED-CFP-A
GFP/FITCS 49002 GFP-4050B 460 (Ch.B) 49002 GFP-4050B 49002 GFP-4050B
YFP 49023 LED-YFP-A 500 (Ch.C) 49003 LED-YFP-A 49023 LED-YFP-A
TRITC/Cy3 49004 Cy3-4040C 550 (Ch.C) 49004 Cy3-4040C 49004 Cy3-4040C
TxRed/mCherry 49008 mCherry-C 550 (Ch.C) 49008 mCherry-C 49008 mCherry-C
Cy5 49006 Cy5-4040C 635 (Ch.D) 49009 Cy5-4040C 49006 Cy5-4040C
Cy7/IrDye800 N/A N/A 740 (Ch.D) 49007 LED-Cy7-A N/A N/A

 

Full Multi Band Sets pE-300 Series pE-4000 pE-340fura
LED Chroma Semrock LED Chroma Semrock Chroma Semrock
DAPI/FITC/TRITC 69013v2 DA/FI/TR-A 385/470/550
(Ch.A.B.C)
69013v2 DA/FI/TR-A 69013v2**** DA/FI/TR-A****
DAPI/FITC/TxRED 69013v2 DA/FI/TR-A 385/490/580
(Ch.A.B.C)
69015** DA/FI/Tx-B 69013v2**** DA/FI/TR-A****
DAPI/FITC/TRITC/Cy5 89402* LED-DA/FI/TR/
Cy-5B*
385/460/550/635
(Ch.A.B.C.D)
89402 LED-DA/FI/TR/Cy5-B 89402**** LED-DA/FI/TR/
Cy5-B****
GFP/mCherry 59022 GFP/DsRed-A 460/550(Ch.B.C) 59022 GFP/DsRed-A 59022***** GFP/DsRed-A******
CFP/YFP 59017*** LED-CFP/YFP/
mCherry-A
435/500(Ch.A.B) 59017 CFP/YFP-A 59017*** LED-CFP/YFP/
mCherry-A*****
CFP/YFP/mCherry 69008*** LED-CFP-YFP/
mCherry-A
435/500/580
(Ch.A.B.C)
69008 LED-CFP/YFP/mCherry-A 69008***** LED-CFP/#YFP/
mCherry-A*****
CFP/YFP/RFP/Cy7 N/A N/A N/A 89403 N/A N/A N/A

 

Pinkel Sets pE-300ultra MB (UV@400) pE-4000 pE-340fura
Chroma Semrock LED Chroma LED Semrock Chroma Semrock
FURA-2 N/A N/A N/A N/A N/A N/A 79001 FURA2-C
FURA-2/GFP N/A N/A N/A N/A N/A N/A 79004 N/A
FURA-2/TRITC N/A N/A N/A N/A N/A N/A 79003 N/A
DAPI/FITC/TRITC 63902 DA/FI/
TR-3X-A
405/470/580
(Ch.A.B.C)
63902

385/470/550

(Ch.A.B.C)

DA/FI/
TR-3X-A
N/A N/A
DAPI/FITC/TxRED 63902 LED-DA/
FI/TC-3X-A
405/470/580
(Ch.A.B.C)
63902

405/460/580

(Ch.A.B.C)

LED-DA/FI/
TX-3X-A
N/A N/A

BFP/GFP/
HcRed-3X-A

(SB UV@365)

N/A N/A
DAPI/FITC/TRITC/Cy5 N/A N/A 405/470/550/635
(Ch.A.B.C.D)
89400

385/460/550/635

(Ch.A.B.C.D)

LED-DA/FI-TR/
Cy5-4x-B
N/A N/A
GFP/mCherry 59222 GFP/
HcRed-2X-A
460/580/
(Ch.B.C)
59222

460/580

(Ch.B.C)

GFP/
HcRed-2X-A
N/A N/A
CFP/YFP 59217*** CFP/
YFP-2X-A***
435/500
(Ch.A.B)
59217

460/580

(Ch.B.C)

GFP/
HcRed-2X-A
N/A N/A
CFP/YFP/mCherry 69308*** LED-mTFP/Venus/
mCherry-3X-A
435/500/580
(Ch.A.B.C)
69308

435/500/580

(Ch.A.B.C)

LED-CFP/YFP/
mCherry- 3XA
N/A N/A

 

* pE-300lite will allow vis. Of all 4. pE-300white & pE-300ultra will allow manual and software
control of DAvsFIvsTR&Cy5
** Very specific to TxRed/mCherry/Red FISH and but not recomended for TRITC/Cy3 etc..  
*** Only recommended for stronger YFP signals        
**** All visable, only independent control of DAPI – others via single source in the pE-340fura  
***** All visiable but via same source -so no switching or balancing