Proximity ligation assay

ProximityAssay14TC

Proximity ligation assay

A proximity ligation assay (PLA) is a molecular biology technique used to detect protein interactions and modifications in cells and tissues. It is based on the principle of detecting the close proximity of two target molecules by using pairs of specific DNA probes that can only hybridize when in close proximity.

Principle

In a proximity ligation assay, two target proteins are recognized by primary antibodies that are conjugated to DNA oligonucleotides. If the two proteins are in close proximity, the DNA oligonucleotides can hybridize to connector oligonucleotides, which are then ligated together by a DNA ligase enzyme. The ligated DNA strands can be amplified and detected using techniques such as polymerase chain reaction (PCR) or fluorescence microscopy.

Applications

Proximity ligation assays are commonly used in research settings to study protein-protein interactions, post-translational modifications, and protein localization within cells. They can provide valuable insights into signaling pathways, protein complexes, and disease mechanisms.

Advantages

One of the main advantages of proximity ligation assays is their high specificity and sensitivity in detecting protein interactions. They can also be multiplexed to simultaneously detect multiple protein targets in a single sample. Additionally, PLA can be performed on fixed cells or tissues, making it suitable for studying protein interactions in biological samples.

Limitations

Despite its advantages, proximity ligation assays also have limitations. The technique can be time-consuming and labor-intensive, requiring multiple steps for probe hybridization, ligation, and amplification. False positives can occur if the DNA probes nonspecifically bind to other molecules in the sample. Optimization of experimental conditions is crucial to minimize background noise and ensure reliable results.

See also

• Protein-protein interaction
• Enzyme-linked immunosorbent assay