Search research articles

ABOUT JoVE

Overview Leadership Blog JoVE Help Center

AUTHORS

Publishing Process Editorial Board Scope & Policies Peer Review FAQ Submit

LIBRARIANS

Testimonials Subscriptions Access Resources Library Advisory Board FAQ

RESEARCH

JoVE Journal Methods Collections JoVE Encyclopedia of Experiments Archive

EDUCATION

JoVE Core JoVE Business JoVE Science Education JoVE Lab Manual Faculty Resource Center Faculty Site

Terms & Conditions of Use

Related Concept Videos

The Equilibrium Binding Constant and Binding Strength

The Equilibrium Binding Constant and Binding Strength

The equilibrium binding constant (Kb) quantifies the strength of a protein-ligand interaction. Kb can be calculated as follows when the reaction is at equilibrium:

Ligand Binding Sites

Ligand Binding Sites

Proteins are dynamic macromolecules that carry out a wide variety of essential processes; however, the activities of most proteins depend on their interactions with other molecules or ions, known as ligands.
Protein-ligand interactions are quite specific; even though numerous potential ligands surround a cellular protein at any given time, only a particular ligand can bind to that protein. Moreover, a ligand binds only to a dedicated area on the surface of the protein, known as the...

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by

Same author

Antisense Oligonucleotide Tofersen Distribution in the Central Nervous System of SOD1-ALS Autopsy Tissue Donors.

JAMA neurology·2026

Same author

Visualization of sonication in high-viscosity polymer melts.

Ultrasonics·2026

Same author

Immunogenicity assays are biomarker assays: is the 3-tiered paradigm fit-for-purpose? An illustrative case study.

Bioanalysis·2026

Same author

From tiers to truth - a biomarker-based framework for clinically relevant immunogenicity assessment.

Bioanalysis·2026

Same author

Rethinking immunogenicity: an integrated approach reveals the PK/PD impact of pre-existing and drug-sustaining ADA.

Bioanalysis·2026

Same author

Lateral Cervical Ectopic Thyroid: A Seven-Step Diagnostic Algorithm and Dual-Axis Treatment Strategy Derived From Case Analysis and Literature Review.

Head & neck·2026

Same journal

A simple, sensitive microsample LC-MS assay for quercetin and isorhamnetin in mouse and human plasma: application to EMIQ treatment in myotonic dystrophy type 1.

Bioanalysis·2026

Same journal

ADA assays for high-dose biologics: redefining drug tolerance through clinical insights.

Bioanalysis·2026

Same journal

Comparison of SERS spectral data sets of blood serum samples of hypopharyngeal cancer using silver and gold nanoparticles as substrates.

Bioanalysis·2026

Same journal

The Gyrolab platform for immunogenicity assessment and biotherapeutic and biomarker analysis: technical advances and bioanalytical applications.

Bioanalysis·2026

Same journal

Simultaneous quantification of D-penicillamine, D-penicillamine disulfide, and L-cysteine-D-penicillamine disulfide in human plasma: optimization of sample preparation and mass spectrometry procedures to support bioequivalence studies.

Bioanalysis·2026

Same journal

Development and preliminary clinical application of a time-resolved fluoroimmunoassay for anti-rituximab antibodies in membranous nephropathy.

Bioanalysis·2026

See all related articles

Search research articles

Related Experiment Video

Updated: Jan 2, 2026

Analyzing DNA-Protein Interactions with Streptavidin-Based Biolayer Interferometry

Analyzing DNA-Protein Interactions with Streptavidin-Based Biolayer Interferometry

Published on: January 17, 2025

Well-developed ligand-binding assays demonstrate robust performance using singlet analysis.

Douglas Donaldson¹, Shobha Purushothama^1,2, Eric David¹

¹Biogen, Cambridge, MA 02142, USA.

|December 13, 2019

Summary

This summary is machine-generated.

Replicate sample testing in ligand-binding assays (LBAs) may no longer be necessary. Modern LBAs provide accurate results with singlet testing, reducing workload and costs.

Keywords:

AUC Cmax LBA MRD assay validation precision singlet variance component analysis

More Related Videos

An ELISA Based Binding and Competition Method to Rapidly Determine Ligand-receptor Interactions

An ELISA Based Binding and Competition Method to Rapidly Determine Ligand-receptor Interactions

Published on: March 14, 2016

Bio-layer Interferometry for Measuring Kinetics of Protein-protein Interactions and Allosteric Ligand Effects

Bio-layer Interferometry for Measuring Kinetics of Protein-protein Interactions and Allosteric Ligand Effects

Published on: February 18, 2014

Related Experiment Videos

Last Updated: Jan 2, 2026

Analyzing DNA-Protein Interactions with Streptavidin-Based Biolayer Interferometry

Analyzing DNA-Protein Interactions with Streptavidin-Based Biolayer Interferometry

Published on: January 17, 2025

An ELISA Based Binding and Competition Method to Rapidly Determine Ligand-receptor Interactions

An ELISA Based Binding and Competition Method to Rapidly Determine Ligand-receptor Interactions

Published on: March 14, 2016

Bio-layer Interferometry for Measuring Kinetics of Protein-protein Interactions and Allosteric Ligand Effects

Bio-layer Interferometry for Measuring Kinetics of Protein-protein Interactions and Allosteric Ligand Effects

Published on: February 18, 2014

Area of Science:

Bioanalytical Chemistry
Pharmacokinetics
Assay Development

Background:

Historically, replicate testing was essential for bioanalytical laboratory testing, particularly for ligand-binding assays (LBAs), due to the imprecision of older assay technologies.
While regulatory guidelines permit singlet testing with demonstrated assay robustness, duplicate testing remains a common practice in many laboratories.

Purpose of the Study:

To re-evaluate the necessity of replicate sample testing in modern bioanalytical assays, specifically ligand-binding assays (LBAs).
To determine if singlet analysis yields comparable results to duplicate analysis in LBAs, even without automation.

Main Methods:

Re-analysis of data from five pharmacokinetic assay validations and five clinical/preclinical studies.
Original assays were performed in duplicate; re-analysis was conducted using singlet testing methodology.

Main Results:

Data re-evaluated using singlet testing showed results nearly identical to the original duplicate testing results.
The findings indicate that well-developed LBAs produce comparable data regardless of whether singlet or duplicate testing is employed.

Conclusions:

The practice of replicate testing for LBAs should be re-evaluated in light of modern assay precision and accuracy.
Automation is not a prerequisite for successful singlet testing in LBAs, challenging the notion that it is required.