A Simple Flow-Cytometric Method Measuring B Cell Surface Immunoglobulin Avidity Enables Characterization of Affinity Maturation to Influenza A Virus

ABSTRACT Antibody (Ab) affinity maturation enables an individual to maintain immunity to an increasing number of pathogens within the limits of a total Ig production threshold. A better understanding of this process is critical for designing vaccines that generate optimal Ab responses to pathogens....

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Gregory M. Frank, Davide Angeletti, William L. Ince, James S. Gibbs, Surender Khurana, Adam K. Wheatley, Edward E. Max, Adrian B. McDermott, Hana Golding, James Stevens, Jack R. Bennink, Jonathan W. Yewdell
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2015
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/b5878bacd8b24a17bf9b60102e41b1e6
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:b5878bacd8b24a17bf9b60102e41b1e6
record_format	dspace
spelling	oai:doaj.org-article:b5878bacd8b24a17bf9b60102e41b1e62021-11-15T15:41:26ZA Simple Flow-Cytometric Method Measuring B Cell Surface Immunoglobulin Avidity Enables Characterization of Affinity Maturation to Influenza A Virus10.1128/mBio.01156-152150-7511https://doaj.org/article/b5878bacd8b24a17bf9b60102e41b1e62015-09-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01156-15https://doaj.org/toc/2150-7511ABSTRACT Antibody (Ab) affinity maturation enables an individual to maintain immunity to an increasing number of pathogens within the limits of a total Ig production threshold. A better understanding of this process is critical for designing vaccines that generate optimal Ab responses to pathogens. Our study describes a simple flow-cytometric method that enumerates virus-specific germinal center (GC) B cells as well as their AC50, a measure of Ab avidity, defined as the antigen concentration required to detect 50% of specific B cells. Using a model of mouse Ab responses to the influenza A virus hemagglutinin (IAV HA), we obtained data indicating that AC50 decreases with time postinfection in an affinity maturation-dependent process. As proof of principle of the utility of the method, our data clearly show that relative to intranasal IAV infection, intramuscular immunization against inactivated IAV in adjuvant results in a diminished GC HA B cell response, with increased AC50 correlating with an increased serum Ab off-rate. Enabling simultaneous interrogation of both GC HA B cell quantity and quality, this technique should facilitate study of affinity maturation and rational vaccine design. IMPORTANCE Though it was first described 50 years ago, little is known about how antibody affinity maturation contributes to immunity. This question is particularly relevant to developing more effective vaccines for influenza A virus (IAV) and other viruses that are difficult vaccine targets. Limitations in methods for characterizing antigen-specific B cells have impeded progress in characterizing the quality of immune responses to vaccine and natural immunogens. In this work, we describe a simple flow cytometry-based approach that measures both the number and affinity of IAV-binding germinal center B cells specific for the IAV HA, the major target of IAV-neutralizing antibodies. Using this method, we showed that the route and form of immunization significantly impacts the quality and quantity of B cell antibody responses. This method provides a relatively simple yet powerful tool for better understanding the contribution of affinity maturation to viral immunity.Gregory M. FrankDavide AngelettiWilliam L. InceJames S. GibbsSurender KhuranaAdam K. WheatleyEdward E. MaxAdrian B. McDermottHana GoldingJames StevensJack R. BenninkJonathan W. YewdellAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 6, Iss 4 (2015)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Gregory M. Frank Davide Angeletti William L. Ince James S. Gibbs Surender Khurana Adam K. Wheatley Edward E. Max Adrian B. McDermott Hana Golding James Stevens Jack R. Bennink Jonathan W. Yewdell A Simple Flow-Cytometric Method Measuring B Cell Surface Immunoglobulin Avidity Enables Characterization of Affinity Maturation to Influenza A Virus
description	ABSTRACT Antibody (Ab) affinity maturation enables an individual to maintain immunity to an increasing number of pathogens within the limits of a total Ig production threshold. A better understanding of this process is critical for designing vaccines that generate optimal Ab responses to pathogens. Our study describes a simple flow-cytometric method that enumerates virus-specific germinal center (GC) B cells as well as their AC50, a measure of Ab avidity, defined as the antigen concentration required to detect 50% of specific B cells. Using a model of mouse Ab responses to the influenza A virus hemagglutinin (IAV HA), we obtained data indicating that AC50 decreases with time postinfection in an affinity maturation-dependent process. As proof of principle of the utility of the method, our data clearly show that relative to intranasal IAV infection, intramuscular immunization against inactivated IAV in adjuvant results in a diminished GC HA B cell response, with increased AC50 correlating with an increased serum Ab off-rate. Enabling simultaneous interrogation of both GC HA B cell quantity and quality, this technique should facilitate study of affinity maturation and rational vaccine design. IMPORTANCE Though it was first described 50 years ago, little is known about how antibody affinity maturation contributes to immunity. This question is particularly relevant to developing more effective vaccines for influenza A virus (IAV) and other viruses that are difficult vaccine targets. Limitations in methods for characterizing antigen-specific B cells have impeded progress in characterizing the quality of immune responses to vaccine and natural immunogens. In this work, we describe a simple flow cytometry-based approach that measures both the number and affinity of IAV-binding germinal center B cells specific for the IAV HA, the major target of IAV-neutralizing antibodies. Using this method, we showed that the route and form of immunization significantly impacts the quality and quantity of B cell antibody responses. This method provides a relatively simple yet powerful tool for better understanding the contribution of affinity maturation to viral immunity.
format	article
author	Gregory M. Frank Davide Angeletti William L. Ince James S. Gibbs Surender Khurana Adam K. Wheatley Edward E. Max Adrian B. McDermott Hana Golding James Stevens Jack R. Bennink Jonathan W. Yewdell
author_facet	Gregory M. Frank Davide Angeletti William L. Ince James S. Gibbs Surender Khurana Adam K. Wheatley Edward E. Max Adrian B. McDermott Hana Golding James Stevens Jack R. Bennink Jonathan W. Yewdell
author_sort	Gregory M. Frank
title	A Simple Flow-Cytometric Method Measuring B Cell Surface Immunoglobulin Avidity Enables Characterization of Affinity Maturation to Influenza A Virus
title_short	A Simple Flow-Cytometric Method Measuring B Cell Surface Immunoglobulin Avidity Enables Characterization of Affinity Maturation to Influenza A Virus
title_full	A Simple Flow-Cytometric Method Measuring B Cell Surface Immunoglobulin Avidity Enables Characterization of Affinity Maturation to Influenza A Virus
title_fullStr	A Simple Flow-Cytometric Method Measuring B Cell Surface Immunoglobulin Avidity Enables Characterization of Affinity Maturation to Influenza A Virus
title_full_unstemmed	A Simple Flow-Cytometric Method Measuring B Cell Surface Immunoglobulin Avidity Enables Characterization of Affinity Maturation to Influenza A Virus
title_sort	simple flow-cytometric method measuring b cell surface immunoglobulin avidity enables characterization of affinity maturation to influenza a virus
publisher	American Society for Microbiology
publishDate	2015
url	https://doaj.org/article/b5878bacd8b24a17bf9b60102e41b1e6
work_keys_str_mv	AT gregorymfrank asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT davideangeletti asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT williamlince asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT jamessgibbs asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT surenderkhurana asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT adamkwheatley asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT edwardemax asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT adrianbmcdermott asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT hanagolding asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT jamesstevens asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT jackrbennink asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT jonathanwyewdell asimpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT gregorymfrank simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT davideangeletti simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT williamlince simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT jamessgibbs simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT surenderkhurana simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT adamkwheatley simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT edwardemax simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT adrianbmcdermott simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT hanagolding simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT jamesstevens simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT jackrbennink simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus AT jonathanwyewdell simpleflowcytometricmethodmeasuringbcellsurfaceimmunoglobulinavidityenablescharacterizationofaffinitymaturationtoinfluenzaavirus
_version_	1718427724607913984

A Simple Flow-Cytometric Method Measuring B Cell Surface Immunoglobulin Avidity Enables Characterization of Affinity Maturation to Influenza A Virus

Ejemplares similares