Storage: 2-8°C
UNSPSC Code: 12352204
Components: Luminol Substrate Solution with enhancer; NaBO3 Starting Solution; Blocking Reagent; Anti-mouse IgG-POD antibody, Fab fragments/Anti-rabbit IgG-POD antibody
RIDADR: NONH for all modes of transport
Features and Benefits:
• High sensitivity: Detection with this kit is as sensitive as radioactive detection.
• Fast: Short exposure times ranging from a few seconds to one hour for obtaining highly sensitive results.
• Nonradioactive: Safe and produces no radioactive waste.
• Stable hard-copy results on film: No fading occurs during storage.
• Quantifiable results: Bands or dots on films can be quantified with a densitometer.
• Economically: Detection of antigen with low amounts of antibody or low affinity antibody.
Application:
BM Chemiluminescence Western Blotting Kit (Mouse/Rabbit) is used to detect proteins with peroxidase-labeled secondary antibodies and the chemiluminescent substrate luminol on western blots. The chemiluminescence system is specifically suited for western and dot blot applications where high sensitivity is required. Chemiluminescence detection of proteins is 1 to 3 orders of magnitude more sensitive than colorimetric methods. Compared with radioactive detection of blotted antigens, the chemiluminescence image is at least as sensitive after much shorter exposure times and without the risk of working with radioactivity.
General description:
Detection of membrane-bound molecules is carried out by a variety of procedures. In protein analysis, the molecules of interest are usually detected with specific first antibodies, enzyme-labeled secondary reagents (antibody- or streptavidin-conjugates) and a suitable chromogenic substrate. These detection systems, however, may be time-consuming or lacking sensitivity. In addition, colors tend to fade with time and exposure to light.
To overcome these problems, attempts have been made to use chemiluminescence for fast and sensitive detection of enzyme conjugates on blots. The BM Chemiluminescence Western Blotting System is designed around peroxidase-labeled secondary antibodies and the substrate luminol. In the presence of hydrogen peroxide (H2O2), horseradish peroxidase (POD, HRP) catalyzes the oxidation of diacylhydrazides such as luminol. An activated intermediate reaction product is formed, which decays to the ground state by emitting light. Strong enhancement of the light emission is produced by 4-iodophenol. This acts as a radical transmitter between the formed oxygen radical and luminol.
Other Notes:
For life science research only. Not for use in diagnostic procedures.
Preparation Note:
Working solution: Preparation of additional reagents and solutions
The table inside the PI describes the preparation of working solutions. For reproducible results equilibrate all solutions to 20 to 25 °C before use.
Storage conditions (working solution): • 1% Blocking solution: -15 to -25 °C
• 0.5% Blocking solution: -15 to -25 °C
• POD-labeled secondary antibody stock solution: 12 month at 2 to 8 °C
• POD-labeled secondary antibody working solution: prepare always fresh
• Detection solution: 1 week at 2 to 8 °C
Reconstitution:
Reconstitute lyophilizate in 100 μl double-dist. water. This solution is stable for 12 months at 2 to 8 °C.
Principle:
Each lot is function tested on a dot blot.