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A homogeneous cell-based assay to measure nuclear translocation using beta-galactosidase enzyme fragment
Assay and Drug Development Technologies
|July 13, 2006
Summary
Positional complementation assays using beta-galactosidase (beta gal) enzyme fragment complementation offer a homogeneous method to detect protein translocation. This technique is suitable for high-throughput screening without imaging or reporter genes.
Area of Science:
- Biochemistry
- Cell Biology
- Assay Development
Background:
- Protein translocation is a critical cellular process.
- Detecting protein translocation often requires complex imaging or reporter gene assays.
- Homogeneous assays are desirable for high-throughput screening.
Purpose of the Study:
- To describe and exemplify positional complementation for detecting cellular protein translocation.
- To demonstrate the utility of this method in a high-throughput screening context.
Main Methods:
- Utilized beta-galactosidase (beta gal) enzyme fragment complementation.
- Developed a homogeneous assay format using a microtiter plate.
- Measured nuclear translocation of the glucocorticoid receptor in Chinese hamster ovary-K1 cells.
Main Results:
- Successfully detected nuclear translocation of the glucocorticoid receptor.
- Established a homogeneous, endpoint assay.
- Demonstrated the absence of need for imaging or reporter gene techniques.
Conclusions:
- Positional complementation provides a robust method for measuring protein translocation.
- The assay is suitable for automated, high-throughput screening.
- This technique can identify modulators of nuclear translocation.
