• Identify Guanine nucleotide exchange (GEF) proteins for Ras GTPase proteins: in vitro assay using purified proteins, not suitable for use with cell lysates
• Biochemical characterization of GEF activity
• Identify Ras GEF inhibitors in a high-throughput screen format

Kit contents (60 - 300 assays for 96-well plate, 660 assays for 384-well plate)

• Exchange buffer
• K-Ras4B His-tagged protein
• hSOS1-His-tagged protein
• 384-well plate
• 96-well plate

Equipment & materials required

• Temperature regulated fluorimeter (ex: 485 nm ±20 nm; em 535 nm ±20 nm)

This fluorescence-based assay leverages the spectral difference between free and bound fluorescent guanine nucleotide analogs to monitor small GTPase nucleotide exchange. Upon binding to GTPases, fluorescence intensity increases sharply, enabling detection of GEF activity for known or novel proteins.

Key characteristics

• Detection method: Fluorescence-based assay: (ex: 485 nm em: 535 nm)
• Timeframe: Approximately 2 hours