ECL Western Blotting Detection Reagents
Bioquochem ECL Western Blotting Detection Reagents detect proteins which are bond indirectly to the secondary antibody – HRP. The Reagent 1 together with the Reagent 2 give rise to a reaction catalyzed by the enzyme HRP resulting in the emission of light. The user can detect the signal through X-ray autoradiography.
Scheme 1. Reaction between Reagent 1 and Reagent 2
Protein detection and quantification is a part of expression levels assessments. Most laboratories use them in a broad spectrum of experiments. Mainly this is achieved by proteins electrophoretic separation, followed by its incubation with antibodies. Then, an X-ray reveals the labeled proteins. In Western blotting procedures, proteins are immobilized on a membrane, which is also referred to as a blot. In order to detect them, the user adds a primary antibody that binds specifically to the protein. Then, the addition of a secondary antibody allows the visualization of the first one. The user detects the secondary antibodies, labeled with horseradish peroxidase (HRP), with the ECL kit. By means of luminol-based substrates, this results in a precise detection of proteins even if they are in very low amounts in the sample.