PAB-pHAb IgG Labeling Kit

PAB protein (Protein of Antibody Binding, MW 25 kDa) contains the IgG Fc-binding domain derived from Protein G and includes a cysteine residue at its C-terminus. Under reducing conditions, the C-terminal cysteine of PAB becomes a reduced thiol group. Through appropriate coupling reactions, toxins or dyes, including pH-sensitive dyes, can be specifically conjugated to this reduced thiol group. Such site-specific conjugation does not interfere with the specific binding between PAB and the IgG Fc region. PAB conjugated with toxins or dyes, including pH-sensitive dyes, can serve as a universal reagent for binding human or mouse IgG antibodies, enabling the study of antibody internalization or phagocytosis in relevant cells.

pHAb dyes are pH-sensitive fluorescent dyes that exhibit very low fluorescence at pH > 7, but their fluorescence significantly increases when the solution becomes acidic. The excitation wavelength (Ex) of pHAb dyes is 532 nm, and the emission wavelength (Em) is 560 nm; their fluorescence can be detected using TAMRA or Cy3 filters.

The PAB-pHAb IgG labeling kit includes the PAB-pHAb reagent, which is a complex in which pHAb is specifically conjugated to PAB via a reductive thiol group at the C-terminus of PAB. PAB-pHAb can serve as a universal binding reagent for human or mouse IgG antibodies, and it reflects the endocytic or phagocytic efficiency of antibodies by detecting the fluorescence emitted from the antibody-PAB-pHAb complex under the acidic conditions of endosomes or lysosomes. The PAB-pHAb IgG labeling reagent employs a unique method to remove free, unlabeled pHAb, thereby significantly reducing background caused by free pHAb. It is easy and quick to use, allowing antibody labeling to be completed within 30 minutes; the labeled antibodies do not require purification and can be directly used in subsequent intracellular endocytosis or phagocytosis experiments.