Troubleshooting

Assay optimization

• • Saturation
• • Initial fluorescence
• • Response amplitude
• • Random changes in initial fluorescence
• • Ligand-induced fluorescence change

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Target labeling optimization

• • RED-Tris-NTA labeling of 10x His-tagged proteins
• • Labeling membrane proteins for binding assays

Buffer optimization

• • Buffer - assay buffer composition
• • Reducing agents
• • Buffer autofluorescence
• • Buffer gradients