Proximity ligation assay

Proximity ligation assay (in situ PLA) is a technology that extends the capabilities of traditional immunoassays to include direct detection of proteins, protein interactions, extracellular vesicles and post translational modifications with high specificity and sensitivity.

[1][2] Protein targets can be readily detected and localized with single molecule resolution and objectively quantified in unmodified cells and tissues.

Two primary antibodies raised in different species recognize the target antigen on the proteins of interest (Figure 1).

[4] In the specific case shown (Figure 5), the nucleus is enlarged because this is a B-cell lymphoma cell.

In situ proximity ligation assays (isPLA) has been applied to antibody validation in human tissues with various advantages over IHC, including increased detection specificity, decreased unspecific staining, and better localization.

Figure 1 : PLA starts with the binding of antibodies from different species to 2 proteins of interest, in this case protein * (star) and protein#
Figure 2 : Binding of PLA probes.
Figure 3 : Rolling circle DNA synthesis starts.
Figure 4 : Fluorescent probes bind to the amplified DNA.
Figure 5 : Fluorescence microscopy image showing interaction of the proteins in the cytoplasm. Nucleus in blue, PLA product in red.