Long-arm Biotin Labeling Kit
First Edition (Revised on April, 2016)
As the most common signal amplification systems, biotin and avidin are widely used in the biological detection. The binding efficiency of biotin and avidin will be affected by steric effects. Cloud-Clone Corp. developed Long-arm Biotin Labeling Kit to solve this problem. On one hand, long-arm Biotin retains the characteristics of biotin, on the other hand, it reduces the steric effects and makes higher binding efficiency of biotin and avidin system.
The kits provides filter tubes and related reagents for labeling experiments of 0.1-1mg protein and antibody for six times. Labeling process is simple and quick, and it doesn’t require dialysis.
REAGENTS AND MATERIALS PROVIDED
|biotin 6 tubes
STORAGE AND PERIOD OF VALIDITY
Store at 2-8℃. The period of validity is 1 year.
1. Add proteins or antibodies sample and Reagent A (V:V=1~2) into the inner tube of the filter tube until reaching 2mL. Centrifuge at 7,500 *g for 10min, remove the liquid at the bottom of the outer tube. Add Reagent A to the inner tube until reaching 2mL. Centrifuge at 7,500 g for 10min and collect liquid at the bottom of the inner tube.
2. Dilute the concentrated liquid to 1mg/ml by Reagent A (if the concentration is too low, the concentrated liquid need to be further concentrated ), and aspirate 1ml of diluted sample into the filter tube.
3. Add 150ul of reagent B to NH2-long-arm biotin. Pipette to mix it thoroughly and aspirate 120ul mixture to the filter tube.
4. Incubate and vibrate the tube for 3-4 hours at RT.
5. Add 9.6ul of reagent C into the filter tube, incubate and vibrate for 10 minutes at RT.
6. Centrifuge at 7,500g for 10 minutes, remove the liquid at the bottom of the outer tube. Remaining liquid at the bottom of the inner tube is collected for later use.
1. The Filtration tubes provided in the kit have a molecular weight cutoff (MWCO) of 10KD. So please be careful of the molecular weight of the substances to be labeled.
2. The kit also can be used for labeling of other substances, if the substances can react with -NH2 and have a suitable molecular weight.
3. When NH2-long-arm biotin bind with active center or active center area of proteins, the protein will lose activity. For this kind of protein, other labeling method is recommended.
4. Centrifuge NH2-long-arm biotin before used, prevent the powder adsorbing on the pipe wall and the lid.
5. In order to reduce the samples loss while the amount of sample is very few, the method of dialysis is recommended.