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Preparation :

Use of a 10 mM biotinylation solution is recommended. In order to efficiently biotinylate a sample, biotinylation solution should be used at a 15-fold molar excess over the amount of amine-containing protein. Make sure to calculate the 10 mM biotinylation solution amount (see example below).

Calculate :

A µL of 10 mM biotinylation solution for biotinylation 2 mg IgG (150,000 M.W.)

2 [mg IgG] x 10^-3 [g/mg] x 1/150,000 [mol/g] x 15 [fold] = A [µL of 10 mM biotinylation solution] x 10^-6 [L/µL] x 10 [mmol/L] x 10^-3 [mol/mmol]
A = 20 [µL of 10 mM biotinylation solution]

Direction for Use :

1. Bring each product to room temperature.
2. Dissolve 2 mg of Biotin-LC-LC-NHS [B6096] in 350 µL of DMSO or DMF or 2 mg of Biotin-PEG₂-NHS [B6097] in 400 µL of PBS to prepare a 10 mM biotinylation solution.
3. Dissolve the sample (1-10 mg/mL) in an appropriate buffer such as PBS. Do not use buffers including amines (such as Tris).
4. Add A µL of 10 mM biotinylation solution to the sample solution and incubate the mixed solution for 30 min at room temperature.
5. Remove unreacted and hydrolyzed reagent using desalting column or dialysis methods.

Example for use of B6096

The following is an example of the process used for the biotinylation of Mouse Anti-Human IgG Antibody [M2977] using Biotin-LC-LC-NHS [B6096].

Direction for Use :

1. Bring 2 mg of B6096 to room temperature and dissolve it in 350 µL of DMSO to prepare 10 mM Biotin-LC-LC-NHS solution.
2. Add 10 µL of 10 mM Biotin-LC-LC-NHS solution to 500 µL of M2977 (Conc.: 1 mg/mL Vol.: 0.5 mg).
3. Incubate the mixed solution at room temperature for 30 minutes .
4. Remove unreacted and hydrolyzed reagent by dialysis methods.