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Determining Binding Affinity (KD) of Radiolabeled Antibodies to Immobilized Antigens
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JoVE Journal Medicina
Determining Binding Affinity (KD) of Radiolabeled Antibodies to Immobilized Antigens

Determining Binding Affinity (KD) of Radiolabeled Antibodies to Immobilized Antigens

DOI:
10.3791/63927-v

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07:39 min

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June 23, 2022

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, , ,
1Department of Radiology and Biomedical Imaging,Yale University

Capítulos

  • 00:04Introduction
  • 00:49Buffer Preparation
  • 02:05Antigen Immobilization and Blocking Nonspecific Sites with BSA
  • 03:21Serial Dilutions and Addition of the Radiolabeled Antibody (rAb) Solution
  • 05:08Wash Plates and Assay Radioactivity
  • 06:07Results: Determination of Binding Affinity
  • 07:04Conclusion

Summary

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Here, a method is described to determine the binding affinity (KD) of radiolabeled antibodies to immobilized antigens. KD is the equilibrium dissociation constant that can be determined from a saturation binding experiment by measuring the total, specific, and nonspecific binding of a radiolabeled antibody at various concentrations to its antigen.

Tags

Keywords: Binding AffinityKDRadiolabeled AntibodiesImmobilized AntigensNuclear ImagingNuclear TherapyAntigen ImmobilizationBinding BufferBlocking BufferWashing BufferBSATween 20Serial Dilutions

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