Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

You might also read

Related Articles

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

Sort by
Same author

Molecularly Engineered Wing-Shaped Azobenzene Memristors for Logic-in-Memory and Edge Visual Intelligence.

Advanced science (Weinheim, Baden-Wurttemberg, Germany)·2026
Same author

Correction to: Puerarin alleviates myocardial ischemia-reperfusion injury by enhancing FUNDC1-mediated mitophagy.

Molecular and cellular biochemistry·2026
Same author

Mirror-image Bilateral Complete and Incomplete Cleft Lip and Alveolus in Monozygotic Twins: A Case Report.

The Cleft palate-craniofacial journal : official publication of the American Cleft Palate-Craniofacial Association·2026
Same author

Puerarin alleviates myocardial ischemia-reperfusion injury by enhancing FUNDC1-mediated mitophagy.

Molecular and cellular biochemistry·2026
Same author

Double-stranded RNA and ROS scavenging nanoplatform for modulating skin inflammation.

Nature communications·2026
Same author

Curcumin Alleviates Myocardial Ischaemia-Reperfusion Injury by Inhibiting NLRP3 Inflammasome Activation and Pyroptosis via the Nrf2/HO-1 Pathway: An Integrated Network Pharmacology and Experimental Study.

Basic & clinical pharmacology & toxicology·2026

Related Experiment Video

Updated: May 30, 2025

The Isolation, Differentiation, and Quantification of Human Antibody-secreting B Cells from Blood: ELISpot as a Functional Readout of Humoral Immunity
08:26

The Isolation, Differentiation, and Quantification of Human Antibody-secreting B Cells from Blood: ELISpot as a Functional Readout of Humoral Immunity

Published on: December 14, 2016

15.1K

Theoretical and practical considerations for validating antigen-specific B cell ImmunoSpot assays.

Paul V Lehmann1, Alexey Y Karulin1, Noémi Becza1

  • 1Research & Development Department, Cellular Technology Limited, Shaker Heights, OH 44122, USA.

Journal of Immunological Methods
|January 26, 2025
PubMed
Summary

New guidelines are proposed for validating B cell ImmunoSpot assays, crucial for immune monitoring in regulated settings. These validated assays accurately quantify antigen-specific B cells using minimal blood samples.

Keywords:
Antibody-secreting cellAntigen-specific immunityFluoroSpotMemory B cellMultiplexed immunoassaySARS-CoV-2

More Related Videos

Development of an IFN-γ ELISpot Assay to Assess Varicella-Zoster Virus-specific Cell-mediated Immunity Following Umbilical Cord Blood Transplantation
08:04

Development of an IFN-γ ELISpot Assay to Assess Varicella-Zoster Virus-specific Cell-mediated Immunity Following Umbilical Cord Blood Transplantation

Published on: July 9, 2014

15.7K
Use of Interferon-γ Enzyme-linked Immunospot Assay to Characterize Novel T-cell Epitopes of Human Papillomavirus
13:41

Use of Interferon-γ Enzyme-linked Immunospot Assay to Characterize Novel T-cell Epitopes of Human Papillomavirus

Published on: March 8, 2012

12.4K

Related Experiment Videos

Last Updated: May 30, 2025

The Isolation, Differentiation, and Quantification of Human Antibody-secreting B Cells from Blood: ELISpot as a Functional Readout of Humoral Immunity
08:26

The Isolation, Differentiation, and Quantification of Human Antibody-secreting B Cells from Blood: ELISpot as a Functional Readout of Humoral Immunity

Published on: December 14, 2016

15.1K
Development of an IFN-γ ELISpot Assay to Assess Varicella-Zoster Virus-specific Cell-mediated Immunity Following Umbilical Cord Blood Transplantation
08:04

Development of an IFN-γ ELISpot Assay to Assess Varicella-Zoster Virus-specific Cell-mediated Immunity Following Umbilical Cord Blood Transplantation

Published on: July 9, 2014

15.7K
Use of Interferon-γ Enzyme-linked Immunospot Assay to Characterize Novel T-cell Epitopes of Human Papillomavirus
13:41

Use of Interferon-γ Enzyme-linked Immunospot Assay to Characterize Novel T-cell Epitopes of Human Papillomavirus

Published on: March 8, 2012

12.4K

Area of Science:

  • Immunology
  • Assay Development

Background:

  • B cell ELISPOT/FluoroSpot (B cell ImmunoSpot) assays are valuable for detecting rare antigen-specific B cells in peripheral blood mononuclear cells (PBMC).
  • These assays are increasingly used for immune monitoring in regulated environments.
  • Currently, no standardized guidelines exist for qualifying and validating B cell ImmunoSpot assay results.

Purpose of the Study:

  • To propose guidelines for the qualification and validation of B cell ImmunoSpot assay results.
  • To adapt existing T cell ImmunoSpot testing experience for B cell assays within regulatory requirements.
  • To address the unique characteristics of B cell assays in the validation process.

Main Methods:

  • Development of a streamlined protocol for formal validation of antigen-specific B cell ImmunoSpot assays.
  • Utilizing three experiments and 2.2 × 10^7 PBMC per donor for validation.
  • Validation of a multiplexed assay requiring only 1-2 × 10^6 PBMC per sample.

Main Results:

  • A streamlined protocol enables formal validation of B cell ImmunoSpot assays.
  • A validated multiplexed assay accurately quantifies antigen-specific memory B cells secreting various antibodies (IgM, IgG, IgA, IgE).
  • The assay requires a minimal sample volume (1-2 mL of blood).

Conclusions:

  • The proposed guidelines facilitate the validation of B cell ImmunoSpot assays for regulated immune monitoring.
  • Validated multiplexed B cell ImmunoSpot assays provide high-content data with ease of implementation and scalability.
  • These assays offer an economical use of PBMC and broad applicability to diverse antigenic systems.