Uncover your Binding Kinetics Data Itinerary What data am I missing SPR vs Other Techniques How to Get Binding Kinetics with SPR SPR Applications Optimizing Binding Kinetics Nicoyas SPR Solution ID: 1033504
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1. Webinar: Behind the SPR CurtainUncover your Binding Kinetics Data
2. ItineraryWhat data am I missing?SPR vs. Other TechniquesHow to Get Binding Kinetics with SPRSPR ApplicationsOptimizing Binding KineticsNicoya’s SPR SolutionOpenSPR in 30 SecondsWrap Up2
3. ItineraryWhat data am I missing?SPR vs. Other TechniquesHow to Get Binding Kinetics with SPRSPR ApplicationsOptimizing Binding KineticsNicoya’s SPR SolutionOpenSPR in 30 SecondsWrap Up2
4. ItineraryWhat data am I missing?SPR vs. Other TechniquesHow to Get Binding Kinetics with SPRSPR ApplicationsOptimizing Binding KineticsNicoya’s SPR SolutionOpenSPR in 30 SecondsWrap Up2
5. ItineraryWhat data am I missing?SPR vs. Other TechniquesHow to Get Binding Kinetics with SPRSPR ApplicationsOptimizing Binding KineticsNicoya’s SPR SolutionOpenSPR in 30 SecondsWrap Up2
6. ItineraryWhat data am I missing?SPR vs. Other TechniquesHow to Get Binding Kinetics with SPRSPR ApplicationsOptimizing Binding KineticsNicoya’s SPR SolutionOpenSPR in 30 SecondsWrap Up2
7. ItineraryWhat data am I missing?SPR vs. Other TechniquesHow to Get Binding Kinetics with SPRSPR ApplicationsOptimizing Binding KineticsNicoya’s SPR SolutionOpenSPR in 30 SecondsWrap Up2
8. ItineraryWhat data am I missing?SPR vs. Other TechniquesHow to Get Binding Kinetics with SPRSPR ApplicationsOptimizing Binding KineticsNicoya’s SPR SolutionOpenSPR in 30 SecondsWrap Up2
9. ItineraryWhat data am I missing?SPR vs. Other TechniquesHow to Get Binding Kinetics with SPRSPR ApplicationsOptimizing Binding KineticsNicoya’s SPR SolutionOpenSPR in 30 SecondsWrap Up2
10. Scientific publications involving Surface Plasmon Resonance (SPR) have increased drastically over the years.3
11. What type of data can I get with SPR?On rate (kon) Off rate (koff) Affinity (KD)SpecificityYes/No BindingConcentration4
12. What type of data can I get with SPR?On rate (kon) Off rate (koff) Affinity (KD)SpecificityYes/No BindingConcentration4
13. What type of data can I get with SPR?On rate (kon) Off rate (koff) Affinity (KD)SpecificityYes/No BindingConcentration4
14. What type of data can I get with SPR?On rate (kon) Off rate (koff) Affinity (KD)SpecificityYes/No BindingConcentration4
15. What type of data can I get with SPR?On rate (kon) Off rate (koff) Affinity (KD)SpecificityYes/No BindingConcentration4
16. What type of data can I get with SPR?On rate (kon) Off rate (koff) Affinity (KD)SpecificityYes/No BindingConcentration4
17. Yes/No BindingKoffKonKD5
18. Affinity Doesn’t Tell the Whole Story6
19. Other Binding Techniques/TechnologiesConditionSPRITCMSTBLICoIPReal-time binding YESYESNOProtein-Protein OnlyNOAffinityYESYESYESYESNOSample ConsumptionLowHighIntermediateLowIntermediate/HighSample flexibilityHighLowIntermediateIntermediateLowLabelsNoNoYesNoAntibody-capture7
20. Main SPR BenefitsNo labelling requiredReal-time binding data kon koff KD Low sample volumes neededFlexibility with applications8
21. Main SPR BenefitsNo labelling requiredReal-time binding data kon koff KD Low sample volumes neededFlexibility with applications8
22. Main SPR BenefitsNo labelling requiredReal-time binding data kon koff KD Low sample volumes neededFlexibility with applications8
23. Main SPR BenefitsNo labelling requiredReal-time binding data kon koff KD Low sample volumes neededFlexibility with applications8
24. Surface Plasmon ResonanceLabel-Free Interaction Analysis9
25. How do I get binding kinetics data?ImmobilizationAnalyte PreparationRegenerationFit Sensorgram Curves10
26. How do I get binding kinetics data?ImmobilizationAnalyte PreparationRegenerationFit Sensorgram Curves10
27. How do I get binding kinetics data?ImmobilizationAnalyte PreparationRegenerationFit Sensorgram Curves10
28. How do I get binding kinetics data?ImmobilizationAnalyte PreparationRegenerationFit Sensorgram Curves10
29. Binding Kinetics11
30. ApplicationsBinding Kinetics/Equilibrium Constants:DetectionQuantificationMaterials ScienceProtein-ProteinProtein-AntibodyProtein-Nucleic AcidProtein-LipidProtein-CarbohydratesProtein-Small MoleculeProtein-PeptideProtein-Aptamer12
31. ApplicationsBinding Kinetics/Equilibrium Constants:DetectionQuantificationMaterials ScienceProtein-ProteinProtein-AntibodyProtein-Nucleic AcidProtein-LipidProtein-CarbohydratesProtein-Small MoleculeProtein-PeptideProtein-Aptamer12
32. ApplicationsBinding Kinetics/Equilibrium Constants:DetectionQuantificationMaterials ScienceProtein-ProteinProtein-AntibodyProtein-Nucleic AcidProtein-LipidProtein-CarbohydratesProtein-Small MoleculeProtein-PeptideProtein-Aptamer12
33. ApplicationsBinding Kinetics/Equilibrium Constants:DetectionQuantificationMaterials ScienceProtein-ProteinProtein-AntibodyProtein-Nucleic AcidProtein-LipidProtein-CarbohydratesProtein-Small MoleculeProtein-PeptideProtein-Aptamer12
34. ApplicationsBinding Kinetics/Equilibrium Constants:DetectionQuantificationMaterials ScienceProtein-ProteinProtein-AntibodyProtein-Nucleic AcidProtein-LipidProtein-CarbohydratesProtein-Small MoleculeProtein-PeptideProtein-Aptamer12
35. Protein-ProteinLigand: Biotinylated protein on streptavidin sensor chipAnalyte: Mutant proteinkon = 0.35x106 M-1s-1koff = 2.0x10-3 s-1KD = 5.7 nM13
36. Protein-LipidLigand: Liposomes formed with 2 different phospholipids, immobilized onto LIP-1 Sensor ChipAnalyte: Proprietary proteinLiposome 1kon = 810 M-1s-1koff = 5.53x10-4 s-1KD = 682 nMLiposome 2kon = 1100 M-1s-1koff = 9.24x10-4 s-1KD = 841 nM14
37. Antibody-AntigenLigand: Anti-PSA immobilized onto Gold Sensor Chip with custom chemistryAnalyte: PSAkon = 4.0x104 M-1s-1koff = 1.8x10-6 s-1KD = 4.5 nMCV = 3.2 %15
38. Protein-Small MoleculeLigand: His-tagged Enzyme NTA Sensor ChipAnalyte: ATP (507 Da)kon = 38.0 M-1s-1koff = 1.18x10-2 s-1KD = 303uM16
39. Protein-Nucleic AcidLigand: Aptamer (biotinylated) immobilized onto Streptavidin Sensor ChipAnalyte: Lysozymekon = 1.8x105 M-1s-1koff = 2.6x10-3 s-1KD = 14.6 nM17
40. Large Molecules/Particles/VirusesLigand: CD receptor on COOH sensor chipAnalyte: 500nm drug loaded polymer nanoparticleKD: 0.43µM (+/- 0.03)kon: 1.0e3 1/M*s (+/-67)koff: 4.3e-4 1/s (+/- 0.02e-6)18
41. Optimizing Binding Kinetics
42. Optimizing Binding Kinetics 20
43. Ligand ImmobilizationCovalent CouplingCarboxylAmineThiolCapture CouplingStreptavidinBiotinNTAGST21HydrophobicLIPMEM
44. Non-Specific BindingNon-specific interaction of the analyte with the sensor chipBSA – up to 1%Salt – up to 1 MTween 20 – 0.005-0.5%Adjust pH22
45. Sample PreparationBuffer: 1. pH2. degass3. filter Unstable additives (ex: ATP) – new buffer everydayDegass first, then add additives (detergent/BSA/DMSO)Sample and buffer match23
46. Sample PreparationBuffer: 1. pH2. degass3. filter Unstable additives (ex: ATP) – new buffer everydayDegass first, then add additives (detergent/BSA/DMSO)Sample and buffer match23
47. Sample PreparationBuffer: 1. pH2. degass3. filter Unstable additives (ex: ATP) – new buffer everydayDegass first, then add additives (detergent/BSA/DMSO)Sample and buffer match23
48. Sample PreparationBuffer: 1. pH2. degass3. filter Unstable additives (ex: ATP) – new buffer everydayDegass first, then add additives (detergent/BSA/DMSO)Sample and buffer match23
49. Estimate optimal ligand immobilization levelLowest immobilization level that gives sufficient signal for multiple analyte concentrations 24
50. Binding CurvesPreliminary binding test with low concentrationChoose 2 to 5 concentrations Dilute your samples 3 fold3 repeatsIncrease interaction time to get curvature1234525
51. Limiting Mass Transfer EffectsHigher flow ratesLowest surface density of ligandMass transport corrected modelsAnalyte26
52. Regeneration ConditionsSensor ChipAnalyteLigand27
53. Optimizing Regeneration ConditionsCommon Regeneration Conditions:Start with least hard conditionRun at fast pump flow rate28
54. CleaningMaintenance is Key:Post ExperimentWeekly MaintenanceYearly Maintenance29
55. LimitationsWhat are the biggest limitations with SPR?No access to instrumentationCostly core facilitiesCostly maintenance contractsSteep learning curves30
56. OpenSPR is built for scientists.Our mission: Design cutting-edge sensor technology to unleash the limitless potential of lifesciences researchers so they can make their next big discovery.31
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58. SolutionAvoid costly & inconvenient core facilities with our affordable benchtop solutionBenchtopTrain anyone in your lab to become an SPRexpert with our user-friendly solutionUser-FriendlyPublish faster with label-free bindingkinetics & affinity dataReal-Time DataForget about expensive service contractsso you can focus on your researchLow Maintenance33
59. LSPR vs SPRNicoya Lifesciences: LSPRTraditional SPRSPECTROMETERGold FilmGlass substrateSensor ChipReflected Light34
60. Localized Surface Plasmon Resonance (LSPR)35
61. Advantages of LSPRAffordable sensors & instrumentNo temperature controlSimple and robust opticsMinimal background interferenceUser friendly Evanescent plasmon field36
62. Advantages of LSPRAffordable sensors & instrumentNo temperature controlSimple and robust opticsMinimal background interferenceUser friendly Evanescent plasmon field36
63. Advantages of LSPRAffordable sensors & instrumentNo temperature controlSimple and robust opticsMinimal background interferenceUser friendly Evanescent plasmon field36
64. Advantages of LSPRAffordable sensors & instrumentNo temperature controlSimple and robust opticsMinimal background interferenceUser friendly Evanescent plasmon field36
65. Advantages of LSPRAffordable sensors & instrumentNo temperature controlSimple and robust opticsMinimal background interferenceUser friendly Evanescent plasmon field36
66. OpenSPR Instrument37
67. OpenSPR in 30 Seconds38
68. RecapWhat did I learn today?SPR data and comparisonOptimizing binding kineticsSPR applications30 seconds with OpenSPR39
69. RecapWhat did I learn today?SPR data and comparisonOptimizing binding kineticsSPR applications30 seconds with OpenSPR39
70. RecapWhat did I learn today?SPR data and comparisonOptimizing binding kineticsSPR applications30 seconds with OpenSPR39
71. RecapWhat did I learn today?SPR data and comparisonOptimizing binding kineticsSPR applications30 seconds with OpenSPR39
72. You’re in good company.40
73. Questions or Comments?Donna Nguyen Application Scientistdonna@nicoyalife.com 1 877-673-6777 ext. 708www.nicoyalife.com
74. Thanks for Listening!Let’s Connect:@Nicoya_Life@NicoyaLifesciences@nicoyalifescienceslinkedin.com/company/nicoya-lifesciences