With so many enhanced chemiluminescent substrates to choose from, it may seem overwhelming to select the substrate that will best suit your lab’s needs. Start by asking yourself the following questions:
- Is the abundance of target protein in my sample low or high?
- Will I need to detect low and high abundance proteins on the same blot?
- Does my data need to be quantitative?
- Is cost an issue?
- Consider your workflow. Do you require a long signal duration?
How does it work?
Enhanced chemiluminescence (ECL) is a highly sensitive method used to detect proteins in Western blot experiments. In the chemiluminescence reaction, horseradish peroxidase catalyzes the oxidation of luminol in the presence of hydrogen peroxide, a strong oxidizer. The oxidized luminol produces a product that emits light as it decays. Unfortunately, the light emitted has a low intensity and the duration is short. To address these shortcomings, an enhancer is added to the luminol reagent. This addition greatly enhances light intensity as well as alters the reaction kinetics to improve signal duration. Light emission only occurs during the enzyme-substrate reaction, so once the substrate in proximity to the enzyme is exhausted light is no longer emitted. All WesternBright™ substrates are prepared with highly purified raw materials to yield high signal, low background, durable signal and lot-to-lot consistency.
What’s in a name?
WesternBright™ ECL is our flagship product. It has all the benefits that you would expect from an ECL substrate. Its sensitivity, low background, high signal-to-noise ratios, dynamic range and affordability make it a clear leader in its class. This versatile substrate works great with film and CCD cameras.
WesternBright™ Quantum was developed for those who require quantitative data. With its extended dynamic range you will be able to quantitate both high and low abundance proteins on the same blot. Quantum was optimized for those using a CCD camera since the detection of signal covers a much broader range as compared to film. With that said, Quantum will still work with film but its value may be diminished.
WesternBright™ Sirius, as the name implies, is our brightest shining star. This substrate was developed for those who are detecting low abundance proteins. Sirius will deliver maximum sensitivity with low background and a durable signal duration. Because of its high sensitivity, you can decrease the amount of primary and secondary antibody per blot for significant cost savings. Sirius is also compatible with both film and CCD imaging.
AdvanBlock™-Chemi blocking solution:
HRP Conjugated Secondary Antibodies:
AdvanWash washing solution: