Immunoglobulin G genetic variation can confound assessment of antibody levels via altered binding to detection reagents

Ruth A Purcell; L Carissa Aurelia; Robyn Esterbauer; Lilith F Allen; Katherine A Bond; Deborah A Williamson; Janine M Trevillyan; Jason A Trubiano; Jennifer J Juno; Adam K Wheatley; Miles P Davenport; Thi Ho Nguyen; Katherine Kedzierska; Stephen J Kent; Kevin John Selva; Amy W Chung

doi:10.1002/cti2.1494

Immunoglobulin G genetic variation can confound assessment of antibody levels via altered binding to detection reagents

Ruth A Purcell, L Carissa Aurelia, Robyn Esterbauer, Lilith F Allen, Katherine A Bond, Deborah A Williamson, Janine M Trevillyan, Jason A Trubiano, Jennifer J Juno, Adam K Wheatley, Miles P Davenport, Thi Ho Nguyen, Katherine Kedzierska, Stephen J Kent, Kevin John Selva, Amy W Chung^*

^*Corresponding author for this work

School of Medicine

Research output: Contribution to journal › Article › peer-review

Abstract

Objectives. Amino acid variations across more than 30 immunoglobulin (Ig) allotypes may introduce structural changes that influence recognition by anti-Ig detection reagents, consequently confounding interpretation of antibody responses, particularly in genetically diverse cohorts. Here, we assessed a panel of commercial monoclonal anti-IgG1 clones for capacity to universally recognise two dominant IgG1 haplotypes (G1m-1,3 and G1m1,17).

Methods. Four commercial monoclonal anti-human IgG1 clones were assessed via ELISAs and multiplex bead-based assays for their ability to bind G1m-1,3 and G1m1,17 IgG1 variants. Detection antibodies were validated against monoclonal IgG1 allotype standards and tested for capacity to recognise antigen-specific plasma IgG1 from G1m-1,3 and G1m1,17 homozygous and heterozygous SARS-CoV-2 BNT162b2 vaccinated ( n = 28) and COVID-19 convalescent ( n = 44) individuals. An Fc-specific pan-IgG detection antibody corroborated differences between hinge- and Fc-specific anti-IgG1 responses.

Results. Hinge-specific anti-IgG1 clone 4E3 preferentially bound G1m1,17 compared to G1m-1,3 IgG1. Consequently, SARS-CoV-2 Spike-specific IgG1 levels detected in G1m1,17/G1m1,17 BNT162b2 vaccinees appeared 9- to 17-fold higher than in G1m-1,3/G1m-1,3 vaccinees. Fc-specific IgG1 and pan-IgG detection antibodies equivalently bound G1m-1,3 and G1m1,17 IgG1 variants, and detected comparable Spike-specific IgG1 levels between haplotypes. IgG1 responses against other human coronaviruses and influenza were similarly poorly detected by 4E3 anti-IgG1 in G1m-1,3/G1m-1,3 subjects.

Conclusion. Anti-IgG1 clone 4E3 confounds assessment of antibody responses in clinical cohorts owing to bias towards detection of G1m1,17 IgG1 variants. Validation of anti-Ig clones should include evaluation of binding to relevant antibody variants, particularly as the role of immunogenetics upon humoral immunity is increasingly explored in diverse populations.

Original language	English
Article number	e1494
Pages (from-to)	1-12
Number of pages	12
Journal	Clinical and Translational Immunology
Volume	13
Issue number	3
DOIs	https://doi.org/10.1002/cti2.1494
Publication status	Published - 29 Feb 2024

Keywords

Allotype
Anti-immunoglobulin
IgG
Polymorphisms
Reproducibility
Serology

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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Copyright © 2024 The Authors. Clinical & Translational Immunology published by John Wiley & Sons Australia, Ltd on behalf of Australian and New Zealand Society for Immunology, Inc. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
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Purcell, R. A., Aurelia, L. C., Esterbauer, R., Allen, L. F., Bond, K. A., Williamson, D. A., Trevillyan, J. M., Trubiano, J. A., Juno, J. J., Wheatley, A. K., Davenport, M. P., Nguyen, T. H., Kedzierska, K., Kent, S. J., Selva, K. J., & Chung, A. W. (2024). Immunoglobulin G genetic variation can confound assessment of antibody levels via altered binding to detection reagents. Clinical and Translational Immunology, 13(3), 1-12. Article e1494. https://doi.org/10.1002/cti2.1494

@article{be0ca9f654744d43a16c1ec02c10b56d,

title = "Immunoglobulin G genetic variation can confound assessment of antibody levels via altered binding to detection reagents",

abstract = "Objectives. Amino acid variations across more than 30 immunoglobulin (Ig) allotypes may introduce structural changes that influence recognition by anti-Ig detection reagents, consequently confounding interpretation of antibody responses, particularly in genetically diverse cohorts. Here, we assessed a panel of commercial monoclonal anti-IgG1 clones for capacity to universally recognise two dominant IgG1 haplotypes (G1m-1,3 and G1m1,17).Methods. Four commercial monoclonal anti-human IgG1 clones were assessed via ELISAs and multiplex bead-based assays for their ability to bind G1m-1,3 and G1m1,17 IgG1 variants. Detection antibodies were validated against monoclonal IgG1 allotype standards and tested for capacity to recognise antigen-specific plasma IgG1 from G1m-1,3 and G1m1,17 homozygous and heterozygous SARS-CoV-2 BNT162b2 vaccinated ( n = 28) and COVID-19 convalescent ( n = 44) individuals. An Fc-specific pan-IgG detection antibody corroborated differences between hinge- and Fc-specific anti-IgG1 responses. Results. Hinge-specific anti-IgG1 clone 4E3 preferentially bound G1m1,17 compared to G1m-1,3 IgG1. Consequently, SARS-CoV-2 Spike-specific IgG1 levels detected in G1m1,17/G1m1,17 BNT162b2 vaccinees appeared 9- to 17-fold higher than in G1m-1,3/G1m-1,3 vaccinees. Fc-specific IgG1 and pan-IgG detection antibodies equivalently bound G1m-1,3 and G1m1,17 IgG1 variants, and detected comparable Spike-specific IgG1 levels between haplotypes. IgG1 responses against other human coronaviruses and influenza were similarly poorly detected by 4E3 anti-IgG1 in G1m-1,3/G1m-1,3 subjects. Conclusion. Anti-IgG1 clone 4E3 confounds assessment of antibody responses in clinical cohorts owing to bias towards detection of G1m1,17 IgG1 variants. Validation of anti-Ig clones should include evaluation of binding to relevant antibody variants, particularly as the role of immunogenetics upon humoral immunity is increasingly explored in diverse populations.",

keywords = "Allotype, Anti-immunoglobulin, IgG, Polymorphisms, Reproducibility, Serology",

author = "Purcell, {Ruth A} and Aurelia, {L Carissa} and Robyn Esterbauer and Allen, {Lilith F} and Bond, {Katherine A} and Williamson, {Deborah A} and Trevillyan, {Janine M} and Trubiano, {Jason A} and Juno, {Jennifer J} and Wheatley, {Adam K} and Davenport, {Miles P} and Nguyen, {Thi Ho} and Katherine Kedzierska and Kent, {Stephen J} and Selva, {Kevin John} and Chung, {Amy W}",

note = "Funding: This study was supported by a Medical Research Future Fund (MRFF) GNT #2016062 to JAT, JAJ, AKW, MPD, THON, KK, SJK and AWC, and a National Health and Medical Research Council (NHMRC) Investigator grant #2008092 to AWC. JAJ, AKW, MPD, THON, KK and SJK are also supported by NHMRC Investigator grants.",

year = "2024",

month = feb,

day = "29",

doi = "10.1002/cti2.1494",

language = "English",

volume = "13",

pages = "1--12",

journal = "Clinical and Translational Immunology",

issn = "2050-0068",

number = "3",

}

Purcell, RA, Aurelia, LC, Esterbauer, R, Allen, LF, Bond, KA, Williamson, DA, Trevillyan, JM, Trubiano, JA, Juno, JJ, Wheatley, AK, Davenport, MP, Nguyen, TH, Kedzierska, K, Kent, SJ, Selva, KJ & Chung, AW 2024, 'Immunoglobulin G genetic variation can confound assessment of antibody levels via altered binding to detection reagents', Clinical and Translational Immunology, vol. 13, no. 3, e1494, pp. 1-12. https://doi.org/10.1002/cti2.1494

TY - JOUR

T1 - Immunoglobulin G genetic variation can confound assessment of antibody levels via altered binding to detection reagents

AU - Purcell, Ruth A

AU - Aurelia, L Carissa

AU - Esterbauer, Robyn

AU - Allen, Lilith F

AU - Bond, Katherine A

AU - Williamson, Deborah A

AU - Trevillyan, Janine M

AU - Trubiano, Jason A

AU - Juno, Jennifer J

AU - Wheatley, Adam K

AU - Davenport, Miles P

AU - Nguyen, Thi Ho

AU - Kedzierska, Katherine

AU - Kent, Stephen J

AU - Selva, Kevin John

AU - Chung, Amy W

N1 - Funding: This study was supported by a Medical Research Future Fund (MRFF) GNT #2016062 to JAT, JAJ, AKW, MPD, THON, KK, SJK and AWC, and a National Health and Medical Research Council (NHMRC) Investigator grant #2008092 to AWC. JAJ, AKW, MPD, THON, KK and SJK are also supported by NHMRC Investigator grants.

PY - 2024/2/29

Y1 - 2024/2/29

N2 - Objectives. Amino acid variations across more than 30 immunoglobulin (Ig) allotypes may introduce structural changes that influence recognition by anti-Ig detection reagents, consequently confounding interpretation of antibody responses, particularly in genetically diverse cohorts. Here, we assessed a panel of commercial monoclonal anti-IgG1 clones for capacity to universally recognise two dominant IgG1 haplotypes (G1m-1,3 and G1m1,17).Methods. Four commercial monoclonal anti-human IgG1 clones were assessed via ELISAs and multiplex bead-based assays for their ability to bind G1m-1,3 and G1m1,17 IgG1 variants. Detection antibodies were validated against monoclonal IgG1 allotype standards and tested for capacity to recognise antigen-specific plasma IgG1 from G1m-1,3 and G1m1,17 homozygous and heterozygous SARS-CoV-2 BNT162b2 vaccinated ( n = 28) and COVID-19 convalescent ( n = 44) individuals. An Fc-specific pan-IgG detection antibody corroborated differences between hinge- and Fc-specific anti-IgG1 responses. Results. Hinge-specific anti-IgG1 clone 4E3 preferentially bound G1m1,17 compared to G1m-1,3 IgG1. Consequently, SARS-CoV-2 Spike-specific IgG1 levels detected in G1m1,17/G1m1,17 BNT162b2 vaccinees appeared 9- to 17-fold higher than in G1m-1,3/G1m-1,3 vaccinees. Fc-specific IgG1 and pan-IgG detection antibodies equivalently bound G1m-1,3 and G1m1,17 IgG1 variants, and detected comparable Spike-specific IgG1 levels between haplotypes. IgG1 responses against other human coronaviruses and influenza were similarly poorly detected by 4E3 anti-IgG1 in G1m-1,3/G1m-1,3 subjects. Conclusion. Anti-IgG1 clone 4E3 confounds assessment of antibody responses in clinical cohorts owing to bias towards detection of G1m1,17 IgG1 variants. Validation of anti-Ig clones should include evaluation of binding to relevant antibody variants, particularly as the role of immunogenetics upon humoral immunity is increasingly explored in diverse populations.

AB - Objectives. Amino acid variations across more than 30 immunoglobulin (Ig) allotypes may introduce structural changes that influence recognition by anti-Ig detection reagents, consequently confounding interpretation of antibody responses, particularly in genetically diverse cohorts. Here, we assessed a panel of commercial monoclonal anti-IgG1 clones for capacity to universally recognise two dominant IgG1 haplotypes (G1m-1,3 and G1m1,17).Methods. Four commercial monoclonal anti-human IgG1 clones were assessed via ELISAs and multiplex bead-based assays for their ability to bind G1m-1,3 and G1m1,17 IgG1 variants. Detection antibodies were validated against monoclonal IgG1 allotype standards and tested for capacity to recognise antigen-specific plasma IgG1 from G1m-1,3 and G1m1,17 homozygous and heterozygous SARS-CoV-2 BNT162b2 vaccinated ( n = 28) and COVID-19 convalescent ( n = 44) individuals. An Fc-specific pan-IgG detection antibody corroborated differences between hinge- and Fc-specific anti-IgG1 responses. Results. Hinge-specific anti-IgG1 clone 4E3 preferentially bound G1m1,17 compared to G1m-1,3 IgG1. Consequently, SARS-CoV-2 Spike-specific IgG1 levels detected in G1m1,17/G1m1,17 BNT162b2 vaccinees appeared 9- to 17-fold higher than in G1m-1,3/G1m-1,3 vaccinees. Fc-specific IgG1 and pan-IgG detection antibodies equivalently bound G1m-1,3 and G1m1,17 IgG1 variants, and detected comparable Spike-specific IgG1 levels between haplotypes. IgG1 responses against other human coronaviruses and influenza were similarly poorly detected by 4E3 anti-IgG1 in G1m-1,3/G1m-1,3 subjects. Conclusion. Anti-IgG1 clone 4E3 confounds assessment of antibody responses in clinical cohorts owing to bias towards detection of G1m1,17 IgG1 variants. Validation of anti-Ig clones should include evaluation of binding to relevant antibody variants, particularly as the role of immunogenetics upon humoral immunity is increasingly explored in diverse populations.

KW - Allotype

KW - Anti-immunoglobulin

KW - IgG

KW - Polymorphisms

KW - Reproducibility

KW - Serology

U2 - 10.1002/cti2.1494

DO - 10.1002/cti2.1494

M3 - Article

C2 - 38433763

SN - 2050-0068

VL - 13

SP - 1

EP - 12

JO - Clinical and Translational Immunology

JF - Clinical and Translational Immunology

IS - 3

M1 - e1494

ER -

Immunoglobulin G genetic variation can confound assessment of antibody levels via altered binding to detection reagents

Abstract

Keywords

UN SDGs

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