The video above follows the simple and straightforward procedure that allows you to detect, quantify and obtain cell localization of protein interactions and their modifications in a single experiment.
- Cells or tissue deposited on slides or in microplates are fixed to preserve activation status and transient interactions.
- Validated primary antibodies for the targets are added followed by binding of the PLA probes.
- Hybridization of the oligonucleotide arms of the PLA probes will create a template for rolling circle amplification (RCA) only when the epitopes of the target proteins are in close proximity (<40 nm)
- Amplification and labeling of the RCA product by detection probes.
- The resulting spot can be detected and visualized by standard microscopy.
- Images can easily be quantitatively analyzed using the Duolink Image Tool software, which facilitates both average and single cell data analysis.