pH-sensitive IgG labeling reagents plus

pH sensitive IgG labeling reagents provide an easy solution to measure internalization activities of antibodies. This reagent utilizes a pH-sensitive fluorescently labeled Fc binding protein that binds to IgG antibodies from various species, resulting in the formation of a fluorescently labeled antibody-reagent complex. After antibody internalization, the surrounding pH becomes acidic and significantly enhances fluorescence signal of antibody-reagent complex. The fluorescence intensity can be used as an indicator to determine the internalization activity of antibodies. By measuring the strength of the fluorescence signal, researchers can assess the efficiency of antibody internalization into cells. This information is crucial in understanding the cellular uptake mechanism of antibodies and assessing their efficacy in targeted therapies or diagnostic applications. Additionally, monitoring the fluorescence intensity can also provide insights into the kinetics of antibody internalization, helping researchers optimize experimental condition and improve the design of antibody-based drug delivery systems.

This product can becan be used for human IgG1, IgG2, IgG3, IgG4, rabbit IgG, mouse IgG1, IgG2a, IgG2b and IgG3 and can be detected with  Flow cytometry  FITC or AF488 filter.

1. UA070080 与抗体孵育

 1.1 使用去离子水，将冻干粉形式的 UA070080 根据溶解方法进行复溶。

 1.2 用细胞培养基制备足够体积的 4X待测抗体工作溶液（受试浓度的四倍，受试浓度根据前期流式条件摸索）。例如：2μg/ml可能是抗体的良好起始测试浓度，因此 4X工作溶液为 8μg/ml。

 1.3 用细胞培养基制备足够体积的4X UA070080工作溶液。待测抗体与 UA070080质量比 3.5:1（摩尔比 1:2）。

 1.4 将4X待测抗体工作溶液和4X UA070080工作溶液按 1:1 的体积比混合。室温避光孵育 1h，得到 Ab-UA070080 复合物2X工作液。

2. Ab-UA070080 复合物与细胞孵育

 2.1细胞处理：收集、洗涤细胞，使用完全培养基调整细胞浓度，悬浮细胞 1-2×105 cells/mL 的浓度，贴壁细胞 0.5-1×105cells/mL 的浓度，96 孔板每孔加入 100μL 细胞悬液。

 2.2 孵育：每孔加入 100μL Ab-UA070080 复合物 2X 工作液，37℃ 5% CO2 培养箱中培养。

3. 流式检测

 培养 18-24h 后，收集细胞进行流式检测抗体的内吞效果，使用 FITC 或AF488 通道检测