Regulated protein expression, degradation and trafficking to the cell surface are fundamental elements of normal cellular physiology, and protein dysregulation underlies many disease states. Monitoring changes in protein abundance usually involves SDS-PAGE followed by immunoblotting, a labor-intensive process that requires high-quality antibodies. We have used NanoLuc® Binary Technology (NanoBiT), a two-part complementation system based on NanoLuc® luciferase, to develop a novel peptide tag for sensitively quantifying bioluminescent proteins with no antibody requirement. The peptide tag, designated High BiT (HiBiT), is only 11 amino acids in length. HiBiT-tagged proteins are measured using detection reagents containing its complementing polypeptide Large BiT (LgBiT), which binds with high affinity to HiBiT (KD~1nM) reconstituting a bright, luminescent enzyme. The HiBiT tag uses a simple bioluminescent method to quantify total or surface expression of proteins while offering a wide dynamic range and sensitivity compatible with detecting endogenously expressed proteins when combined with CRISPR gene editing.
Christopher Eggers, PhD
Senior Research Scientist
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