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Receptor Internalization Assays

Imaging-free, quantifiable internalization assays

The rate and extent of receptor internalization can dramatically impact the biological efficacy and mechanism of action of a drug. PathHunter® Internalization assays are simple, imaging-free chemiluminescent assays that quantitatively measure the internalization and recycling patterns of a receptor target. Identify and differentiate between strongly and weakly internalizing biologics!
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PathHunter Internalization Assay Principle

The small enzyme fragment of β-gal called ProLink™ (PK) is appended to the C-terminus of the receptor. The larger, complementing enzyme fragment termed Enzyme Acceptor (EA) is localized exclusively on the surface of early endosomes. Stimulation of the receptor results in receptor activation and subsequent internalization and trafficking to cellular endosomes. These events drive complementation of the two of β-gal enzyme fragments. The resultant increase of β-gal enzyme activity can be easily measured using chemiluminescent substrate. Above, are two examples of a GPCR (Left panel) and a receptor tyrosine kinase (Right panel) internalization assay.

See validation data for internalization assays

Download slides explaining how these assays work, with attached validation data.

View Validation Data

Receptor Internalization with Biologics

OPRM1 Total Internalization


GFP-labeled GPCR


Insulin Receptor Internalization

Cells expressing the mu Opioid receptor (left panel) were incubated with increasing concentrations of a peptide ligand that causes internalization of the receptor. The internalization signal was also verified with a GFP-tagged receptor (middle panel).  Insulin receptor internalization (right panel), denoted by increasing activity, is caused by increasing concentrations of insulin.