Biotin Conjugation Kit
Biotin is a widely used and powerful tool for research due to the specific and high affinity with streptavidin/avidin. Antibody or protein conjugated with several biotin molecules can also amplify the detection signal through streptavidin-conjugated molecule such as streptavidin-HRP, streptavidin-FITC, etc. Biotinylated antibody or protein can be used in various applications including ELISA, WB, IHC, IFA and FACS. Croyez Biotin Labeling Kit is designed for biotinylation of a small quantity (10 μg-1 mg) of antibody or protein. It provides a rapid and easy process with high efficiency to conjugate biotin to antibody or protein.
Component:
Stability & Storage:
-20°C or -80°C for 12 months under sterile conditions from date of receipt.
Equilibrate kit to room temperature before use.
Note:
Common non-buffering salts (e.g. sodium chloride) have no effect on conjugation efficiency. Avoid buffer component that contains primary amine (e.g. amino acid or ethanolamine) and thiols (e.g. mercaptoethanol or DTT).
Components that have on effect or little effect on labeling reaction:
Blue ice
| C08001-10 | C08001-100 | C08001-1000 | |
| Biotin | 1 vial | 1 vial | 1 vial |
| 10X Modifier | 1 vial | 1 vial | 1 vial |
| 10X Quencher | 1 vial | 1 vial | 1 vial |
Stability & Storage:
-20°C or -80°C for 12 months under sterile conditions from date of receipt.
Equilibrate kit to room temperature before use.
Note:
Common non-buffering salts (e.g. sodium chloride) have no effect on conjugation efficiency. Avoid buffer component that contains primary amine (e.g. amino acid or ethanolamine) and thiols (e.g. mercaptoethanol or DTT).
Components that have on effect or little effect on labeling reaction:
- up to 50 mM Tris
- up to 50 mM HEPES
- up to 10% glycerol
- up to 0.02% sodium azide
Blue ice
Antibody concentrations of 1-4 mg/mL generally give optimal results.
Recommended amount and volume of antibody for optimal results.
| Kit size | Antibody amount | Reaction volume |
| 10 μg | 10-20 μg | 4-20 μL |
| 100 μg | 100-200 μg | 40-200 μL |
| 1 mg | 1-2 mg | 400-2000 μL |
Biotin conjugation protocol
1. Dissolve antibody in PBS or other buffers that do not contain amine, Tris or glycerol. Use 10X Modifier (e.g. Add 1 μL of 10X Modifier for 9 μL of antibody) or dialysis against PBS if pH value of used buffer is out of 7 to 8.
2. Spin down and equilibrate the vial of Biotin at room temperature before opening the cap.
3. Make sure all buffers are well dissolved before using. If not, please vortex the vial to make salts dissolved.
4. Remove the cap of the vial of Biotin and pipette antibody into the vial. Mix gently by pipetting several times.
5. Cover the cap and incubate in the dark at room temperature for 3 hours.
6. After incubating, add 10X Quencher (e.g. 1 μL of 10X Quencher for 9 μL of antibody-biotin mixture) and mix gently by pipetting. The conjugates can be used after 30 minutes.
