New England Biolabs Canada

Product Pathways - Companion Products

SignalFire™ Plus ECL Reagent #12630

Item# Description List Price Web Price Qty
12630P SignalFire™ Plus ECL Reagent - 20 ml $120.00
12630S SignalFire™ Plus ECL Reagent - 100 ml $519.00
*On-line ordering is for Canadian customers only. Web pricing is applicable only to orders placed online at
Petite Kit Includes Quantity
SignalFire™ Plus ECL Reagent A 1 x 10 ml
SignalFire™ Plus ECL Reagent B 1 x 10 ml
Small Kit Includes Quantity
SignalFire™ Plus ECL Reagent A 1 x 50 ml
SignalFire™ Plus ECL Reagent B 1 x 50 ml


SignalFire™ Plus ECL Reagent from Cell Signaling Technology (CST) is a highly sensitive chemiluminescent substrate capable of detecting low picogram amounts of protein by western blot analysis. SignalFire™ Plus ECL Reagent has an extended duration of signal output lasting several hours following blot exposure, allowing for multiple exposures with either film or a digital imaging system. The strong signal output allows detection of low abundance proteins, conservation of reagents, and short exposure times.

SignalFire™ Plus ECL Reagent requires approximately five-fold less Anti-rabbit IgG, HRP-linked Antibody #7074 or Anti-mouse IgG, HRP-linked Antibody #7076 than traditional ECL reagents. Limiting the amount of HRP exposed to the membrane prevents high background, oversaturation of the target protein signal, or false negative results. Other HRP-conjugated antibodies, including HRP-conjugated primary and anti-biotin-HRP antibodies, should be diluted similarly. Dilution of secondary antibody from alternative vendors may need to be optimized. Titration of lysate and primary antibody concentration is recommended to achieve optimal signal-to-noise ratio.

Western Blotting

Western Blotting

SignalFire™ Plus ECL Reagent is sensitive enough to detect low picogram amounts of protein. Western blot analysis of diluted recombinant human Akt-1 protein (hAkt-1) using Akt (pan) (C67E7) Rabbit mAb #4691.

Western Blotting

Western Blotting

SignalFire™ Plus ECL Reagent has extended signal duration. Western blot analysis of diluted recombinant human Akt-1 protein (hAkt-1) using Akt (pan) (C67E7) Rabbit mAb #4691.

Method Overview

(a) Wash membrane-bound HRP (antibody conjugate) at least three times for 5 minutes in TBS/T. It is very important to thoroughly wash the membrane prior to substrate incubation.

(b) Prepare 1X SignalFire™ Plus ECL Reagent by diluting one part 2X Reagent A and one part 2X Reagent B (e.g. for 10 ml, add 5 ml Reagent A and 5 ml Reagent B). Mix well.

(c) Incubate substrate with membrane for 1 minute, remove excess solution (membrane remains wet), wrap in plastic and expose to X-ray film.

*Avoid repeated exposure to skin (see enclosed Material Safety Data Sheet or refer to our website for further information).


Chemiluminescence systems have emerged as the best all-around method for western blot detection. They eliminate the hazards associated with radioactive materials and toxic chromogenic substrates. The speed and sensitivity of these methods are unequalled by traditional alternatives, and because results are generated on film, it is possible to record and store data permanently. Blots detected with chemiluminescent methods are easily stripped for subsequent reprobing with additional antibodies. HRP-conjugated secondary antibodies are utilized in conjunction with specific chemiluminescent substrates to generate the light signal. HRP conjugates have a very high turnover rate, yielding good sensitivity with short reaction times.

Application References

Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!


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