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

查看更多产品信息 Antibody Labeling Kits for 1 mg - FAQs (A10235, A10236, A10237, A10238, A20170, A10239, A20173, P30012, A20174, A20171, A20172, D20655, F10240, O10241, A10170)

2 个常见问题解答

当抗体浓度较低时，我应如何对我的抗体进行生物素化操作？

在抗体浓度较低的情况下，生物素化效率会降低(操作手册中给出的最低浓度为0.5 mg / mL)。在公司内部，我们并没有使用低于0.5mg/mL的抗体，而是增加了DSB-X生物素的含量（从《DSB-X生物素蛋白标记试剂盒手册》(货号D-20655）中的表1可见在偶联反应中把DSB-X生物素含量从2 μL提高到4 μL可以提高反应效率)。< br / > < br / > 根据操作步骤生物素化0.5毫克/毫升的抗体：< br / > 0.5毫克/毫升x 0.2毫升 = 0.1毫克抗体< br / >生物素化的抗体的产量：在0.2毫升中的产量(0.1毫克抗体x 0.85%)= 0.4毫克/毫升< br / >在25μL中的生物素化的抗体量：0.4毫克/毫升x 25μL = 10μg，在推荐范围(每 5 x 10 e7细胞推荐5-50μg)。< br / > < br / > 生物素化0.2毫克/毫升的抗体：0.2毫克/毫升x 0.2毫升 = 0.04毫克抗体< br / >生物素化抗体的产量：在0.2毫升中的产量(0.04毫克抗体x 0.85%)= 0.17毫克/毫升< br / >在25μL中的生物素化的抗体量：0.17毫克/毫升x 25μL = 4μg，稍低于推荐范围。注意：可简单通过离心浓缩器（Centricon滤柱）对抗体进行浓缩，而不影响抗体的生物学活性。

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).

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.