📋 Workflow
Full workflow, products add-to-cart directly1
Ligand immobilization
1 hImmobilize ligand protein on CM5 or other sensor chip by amine coupling or capture
💡 Tip Moderate immobilization avoids mass transport limitation
⚠️ Caution Ligand must be high-purity and stable
🧪 Materials for this step (click to shop by spec/brand)
2
Analyte gradient
30 minPrepare series concentration of analyte (flow) for binding kinetics
💡 Tip Concentration range spans KD
⚠️ Caution Dilute analyte in matching buffer
🧪 Materials for this step (click to shop by spec/brand)
3
Binding-dissociation measurement
2–4 hInject analytes sequentially to record association, then buffer to observe dissociation
💡 Tip Add regeneration for multi-cycle measurements
⚠️ Caution Constant temperature for reliable kinetics
🧪 Materials for this step (click to shop by spec/brand)
4
Kinetic fitting
1 hFit sensorgrams to a 1:1 model with software, calculate ka, kd and equilibrium KD
💡 Tip Assess residuals to validate model
⚠️ Caution Global fitting across concentrations is more reliable
🧪 Materials for this step (click to shop by spec/brand)
⚠️ 5 Common Beginner Mistakes
- ① Over-immobilization causes mass transport limitation
- ② Buffer mismatch causes non-specific binding
- ③ Air bubbles distort sensorgrams
- ④ Insufficient concentration range hampers fitting
- ⑤ Temperature instability drifts kinetics
❓ FAQ
+Can SPR measure KD? Precision?
Yes, SPR measures ka/kd in real time to compute KD, a gold standard for interaction quantification.
+How to choose immobilization?
Capture/directed for small molecules, amine coupling for proteins; control active site carefully.
+What's needed for small molecules?
Need high-sensitivity SPR system capable of small molecules and high-quality ligand.