DSB-X™ Biotin Protein Labeling Kit, 1 kit - FAQs

How can I biotinylate my antibodies if the antibody concentration is low?

Given that the concentration of your antibodies is low, the biotinylation efficiency will be reduced (the lowest concentration given in the protocol is 0.5 mg per mL). In-house, we have not used less than 0.5 µg per mL of antibody, but increasing the amount of DSB-X biotin (Table 1 in manual for DSB-X Biotin Protein Labeling Kit (Cat. No. D-20655) from 2µL to 4µL in the coupling reaction might increase the efficiency).

Biotinylation of 0.5 mg per mL antibody according to the protocol:
0.5 mg per mL x 0.2 mL = 0.1 mg antibody
Yield of biotinylated antibody: (0.1 mg antibody x 0.85%) in 0.2 mL = 0.4 mg per mL
Amount of biotinylated antibody in 25 µL: 0.4 mg per mL x 25 µL = 10 µg antibody in 25 µL, which is in the recommended range (5-50 µg for 5 x 10e7 cells).

Biotinylation of 0.2 mg per mL: 0.2 mg per mL x 0.2 mL = 0.04 mg antibody
Yield of biotinylated antibody: (0.04 mg antibody x 0.85%) in 0.2 mL = 0.17 mg per mL
Amount of biotinylated antibody in 25 µL: 0.17 mg per mL x 25 µL = 4 µg antibody in 25 µL, which is slightly outside the recommended range.

Note: Antibody may be easily concentrated without affecting the biological activity by using centrifuge concentrators (i.e., Centricon filter columns).

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