Revvity, 6FR2BPAF, HTRF CD32b (FcgRIIb) Cellular Binding Kit, 1,000 Assay Points

The CD32b (FcgRIIb) cellular binding assay is a simple and robust test to monitor binding of the IgG Fc region to the human CD32b receptor in live cells. The CD32b receptor negatively modulates the Antibody-Dependent Cell-Phagocytosis (ADCP) pathway. This assay, based on our innovative Tag-lite technology, can easily be automated to create a high-throughput solution for predicting ADCP. HTRF assays offer many advantages over other technologies: Homogeneous add-and-read format No wash steps Low background Straightforward miniaturization from 96- or 384-well microplates to high density assay formats such as 384-well low volume and 1536-well plates Stable signal, providing flexibility in time of readout or size of assays
Assay principle The CD32b receptor cellular binding assay is a competitive assay involving HEK293 cells expressing a specific CD32b variant conjugated to Lumi4-terbium Cryptate, and human IgG1 labelled with d2. The unlabeled antibody or Fc tagged drug competes with the d2 labelled IgG1 for binding to the receptor, resulting in a decrease of FRET signal. Assay protocol The CD32b receptor cellular binding assays (FcgIIb) present only 1 incubation step after reagents distribution. Lumi4-terbium Cryptate pre-labelled cells are dispensed, followed by the addition of IgG1 labelled with d2 and the sample. After 2h incubation at RT, the signal can be read.