Skip to content
1887

Journal of General Virology header logo Journal of General Virology

Volume 105, Issue 12

Comparison of different T cell assays for the retrospective determination of SARS-CoV-2 infection Open Access

Abstract

It is important to be able to retrospectively determine severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections with high accuracy, both for post-coronavirus disease 2019 (COVID-19) epidemiological studies, and to distinguish between Long COVID and other multi-syndromic diseases that have overlapping symptoms. Although serum antibody levels can be measured to retrospectively diagnose SARS-CoV-2 infections, peptide stimulation of memory T cell responses is a more sensitive approach. This is because robust memory T cells are generated after SARS-CoV-2 infection and persist even after antibodies wane below detectability thresholds. In this study, we compare T cell responses using FluoroSpot-based methods and overnight stimulation of whole blood with SARS-CoV-2 peptides followed by an ELISA. Both approaches have comparable sensitivity and specificity but require different equipment and samples to be used. Furthermore, the elimination of peptides that cross-react with other coronaviruses increases the assay specificity but trades off some sensitivity. Finally, this approach can be used on archival, cryopreserved PBMCs. This work shows comparative advantages for several methods to measure SARS-CoV-2 T cell responses that could be utilized by any laboratory studying the effects of the coronavirus disease 2019 pandemic.

  • Received:
  • Accepted:
  • Published Online:
Keyword(s): COVID-19 , peptide , SARS-CoV-2 and T cell
Funding
This study was supported by the:
  • National Institute for Health and Care Research (Award G112259)
    • Principal Award Recipient: NyarieSithole
  • Addenbrooke's Charitable Trust, Cambridge University Hospitals (Award 900276)
    • Principal Award Recipient: NyarieSithole
  • Wellcome Trust (Award 225023/Z/22/Z)
    • Principal Award Recipient: BenjaminKrishna
© 2024 The Authors
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License. This article was made open access via a Publish and Read agreement between the Microbiology Society and the corresponding author’s institution.
Loading

Article metrics loading...

/content/journal/jgv/10.1099/jgv.0.002055
2024-12-20
2025-12-08

Metrics

Loading full text...

Full text loading...

/deliver/fulltext/jgv/105/12/jgv002055.html?itemId=/content/journal/jgv/10.1099/jgv.0.002055&mimeType=html&fmt=ahah

References

  1. Ayoubkhani D, Bosworth ML, King S, Pouwels KB, Glickman M et al. Risk of long COVID in people infected with SARS-CoV-2 after two doses of a COVID-19 vaccine: community-based, matched cohort study. Epidemiology 2022 [View Article]
     [Google Scholar]
  2. Emary KR, Golubchik T, Aley PK, Ariani CV, Angus B et al. Efficacy of chadox1 ncov-19 (AZD1222) vaccine against SARS-cov-2 variant of concern. The Lancet 2021; 202012/01:1351–1362
     [Google Scholar]
  3. Krishna BA, Metaxaki M, Wills MR, Sithole N. Reduced incidence of long coronavirus disease referrals to the Cambridge University teaching hospital long coronavirus disease clinic. Clin Infect Dis 2023; 76:738–740 [View Article]
     [Google Scholar]
  4. Kuodi P, Gorelik Y, Zayyad H, Wertheim O, Wiegler KB et al. Association between vaccination status and reported incidence of post-acute COVID-19. medRxiv 2022 [View Article]
     [Google Scholar]
  5. Thomas SJ, Moreira ED Jr, Kitchin N, Absalon J, Gurtman A et al. Safety and efficacy of the BNT162b2 mRNA Covid-19 vaccine through 6 months. N Engl J Med 2021; 385:1761–1773 [View Article] [PubMed]
     [Google Scholar]
  6. Cerqueira-Silva T, Oliveira V de A, Boaventura VS, Pescarini JM, Júnior JB et al. Influence of age on the effectiveness and duration of protection of Vaxzevria and CoronaVac vaccines: a population-based study. Lancet Reg Health Am 2022; 6:100154 [View Article] [PubMed]
     [Google Scholar]
  7. Nasreen S, Chung H, He S, Brown KA, Gubbay JB et al. Effectiveness of COVID-19 vaccines against symptomatic SARS-CoV-2 infection and severe outcomes with variants of concern in Ontario. Nat Microbiol 2022; 7:379–385 [View Article]
     [Google Scholar]
  8. Pérez-Alós L, Armenteros JJA, Madsen JR, Hansen CB, Jarlhelt I et al. Modeling of waning immunity after SARS-CoV-2 vaccination and influencing factors. Nat Commun 2022; 13:1614 [View Article] [PubMed]
     [Google Scholar]
  9. Healey Q, Sheikh A, Daines L, Vasileiou E. Symptoms and signs of long COVID: a rapid review and meta-analysis. J Glob Health 2022; 12:05014 [View Article] [PubMed]
     [Google Scholar]
  10. Koelle K, Martin MA, Antia R, Lopman B, Dean NE. The changing epidemiology of SARS-CoV-2. Science 2022; 375:1116–1121 [View Article]
     [Google Scholar]
  11. Krishna BA, Metaxaki M, Sithole N, Landín P, Martín P et al. Cardiovascular disease and covid-19: a systematic review. Int J Cardiol Heart Vasc 2024; 54:101482 [View Article] [PubMed]
     [Google Scholar]
  12. Krishna B, Wills M, Sithole N. Long COVID: what is known and what gaps need to be addressed. Br Med Bull 2023; 147:6–19 [View Article] [PubMed]
     [Google Scholar]
  13. Michelen M, Manoharan L, Elkheir N, Cheng V, Dagens A et al. Characterising long COVID: a living systematic review. BMJ Glob Health 2021; 6:e005427 [View Article] [PubMed]
     [Google Scholar]
  14. Subramanian A, Nirantharakumar K, Hughes S, Myles P, Williams T et al. Symptoms and risk factors for long COVID in non-hospitalized adults. Nat Med 2022; 28:1706–1714 [View Article] [PubMed]
     [Google Scholar]
  15. Davis HE, McCorkell L, Vogel JM, Topol EJ. Long COVID: major findings, mechanisms and recommendations. Nat Rev Microbiol 2023; 21:133–146 [View Article] [PubMed]
     [Google Scholar]
  16. Excellence N. COVID-19 rapid guideline: managing the long-term effects of COVID-19; 2020 https://www.nice.org.uk/guidance/ng188
  17. Kim H, Hong H, Yoon SH. Diagnostic performance of CT and reverse transcriptase polymerase chain reaction for coronavirus disease 2019: a meta-analysis. Radiology 2020; 296:E145–E155 [View Article]
     [Google Scholar]
  18. Woloshin S, Patel N, Kesselheim AS. False negative tests for SARS-CoV-2 infection - challenges and implications. N Engl J Med 2020; 383:e38 [View Article] [PubMed]
     [Google Scholar]
  19. Wang W, Wang C-Y, Wang S-I, Wei JC-C. Long-term cardiovascular outcomes in COVID-19 survivors among non-vaccinated population: a retrospective cohort study from the TriNetX US collaborative networks. EClinicalMedicine 2022; 53:101619 [View Article] [PubMed]
     [Google Scholar]
  20. Neumann G, Kawaoka Y. Which virus will cause the next pandemic?. Viruses 2023; 15:199 [View Article] [PubMed]
     [Google Scholar]
  21. Bonifacius A, Tischer-Zimmermann S, Dragon AC, Gussarow D, Vogel A et al. COVID-19 immune signatures reveal stable antiviral T cell function despite declining humoral responses. Immunity 2021; 54:340–354 [View Article] [PubMed]
     [Google Scholar]
  22. Cox RJ, Brokstad KA. Not just antibodies: B cells and T cells mediate immunity to COVID-19. Nat Rev Immunol 2020; 20:581–582 [View Article] [PubMed]
     [Google Scholar]
  23. Dispinseri S, Secchi M, Pirillo MF, Tolazzi M, Borghi M et al. Neutralizing antibody responses to SARS-CoV-2 in symptomatic COVID-19 is persistent and critical for survival. Nat Commun 2021; 12:2670 [View Article] [PubMed]
     [Google Scholar]
  24. Fergie J, Srivastava A. Immunity to SARS-CoV-2: lessons learned. Front Immunol 2021; 12:654165 [View Article] [PubMed]
     [Google Scholar]
  25. Ibarrondo FJ, Fulcher JA, Goodman-Meza D, Elliott J, Hofmann C et al. Rapid decay of anti–SARS-CoV-2 antibodies in persons with mild Covid-19. N Engl J Med 2020; 383:1085–1087 [View Article]
     [Google Scholar]
  26. Long Q-X, Tang X-J, Shi Q-L, Li Q, Deng H-J et al. Clinical and immunological assessment of asymptomatic SARS-CoV-2 infections. Nat Med 2020; 26:1200–1204 [View Article]
     [Google Scholar]
  27. Seow J, Graham C, Merrick B, Acors S, Pickering S et al. Longitudinal observation and decline of neutralizing antibody responses in the three months following SARS-CoV-2 infection in humans. Nat Microbiol 2020; 5:1598–1607 [View Article] [PubMed]
     [Google Scholar]
  28. Barreiro P, Sanz JC, San Román J, Pérez-Abeledo M, Carretero M et al. A pilot study for the evaluation of an Interferon Gamma Release Assay (IGRA) to measure T-cell immune responses after SARS-CoV-2 infection or vaccination in a unique cloistered cohort. J Clin Microbiol 2022; 60:e0219921 [View Article] [PubMed]
     [Google Scholar]
  29. Fernández-González M, Agulló V, Padilla S, García JA, García-Abellán J et al. Clinical performance of a standardized severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) interferon-γ release assay for simple detection of T-Cell responses after infection or vaccination. Clin Infect Dis 2022; 75:e338–e346 [View Article] [PubMed]
     [Google Scholar]
  30. Krishna BA, Lim EY, Mactavous L, Lyons PA, Doffinger R et al. Evidence of previous SARS-CoV-2 infection in seronegative patients with long COVID. eBioMedicine 2022; 81:104129 [View Article]
     [Google Scholar]
  31. Schwarzkopf S, Krawczyk A, Knop D, Klump H, Heinold A et al. Cellular immunity in COVID-19 convalescents with PCR-confirmed infection but with undetectable SARS-CoV-2–specific IgG. Emerg Infect Dis 2021; 27:122–129 [View Article]
     [Google Scholar]
  32. Petrone L, Picchianti-Diamanti A, Sebastiani GD, Aiello A, Laganà B et al. Humoral and cellular responses to spike of δ SARS-CoV-2 variant in vaccinated patients with immune-mediated inflammatory diseases. Int J Infect Dis 2022; 121:24–30 [View Article] [PubMed]
     [Google Scholar]
  33. Murugesan K, Jagannathan P, Pham TD, Pandey S, Bonilla HF et al. Interferon-γ release assay for accurate detection of severe acute respiratory syndrome coronavirus 2 T-cell response. Clin Infect Dis 2021; 73:e3130–e3132 [View Article] [PubMed]
     [Google Scholar]
  34. Oliver MA, Meredith RT, Smith BR, Bermingham MD, Brackett NF et al. Longitudinal T Cell responses against ancestral, delta, and omicron SARS-CoV-2 variants determined by rapid cytokine release assay in whole blood. Immunohorizons 2022; 6:398–407 [View Article]
     [Google Scholar]
  35. Scurr MJ, Lippiatt G, Capitani L, Bentley K, Lauder SN et al. Magnitude of venous or capillary blood-derived SARS-CoV-2-specific T cell response determines COVID-19 immunity. Nat Commun 2022; 13:5422 [View Article] [PubMed]
     [Google Scholar]
  36. Scurr MJ, Zelek WM, Lippiatt G, Somerville M, Burnell SEA et al. Whole blood-based measurement of SARS-CoV-2-specific T cells reveals asymptomatic infection and vaccine immunogenicity in healthy subjects and patients with solid-organ cancers. Immunology 2022; 165:250–259 [View Article] [PubMed]
     [Google Scholar]
  37. Tan AT, Lim JM, Le Bert N, Kunasegaran K, Chia A et al. Rapid measurement of SARS-CoV-2 spike T cells in whole blood from vaccinated and naturally infected individuals. J Clin Invest 2021; 131:e152379 [View Article] [PubMed]
     [Google Scholar]
  38. Lineburg KE, Grant EJ, Swaminathan S, Chatzileontiadou DSM, Szeto C et al. CD8+ T cells specific for an immunodominant SARS-CoV-2 nucleocapsid epitope cross-react with selective seasonal coronaviruses. Immunity 2021; 54:1055–1065 [View Article] [PubMed]
     [Google Scholar]
  39. Riou C, Schäfer G, du Bruyn E, Goliath RT, Stek C et al. Rapid, simplified whole blood-based multiparameter assay to quantify and phenotype SARS-CoV-2-specific T-cells. Eur Respir J 2022; 59:2100285 [View Article] [PubMed]
     [Google Scholar]
  40. Schwarz M, Torre D, Lozano-Ojalvo D, Tan AT, Tabaglio T et al. Rapid, scalable assessment of SARS-CoV-2 cellular immunity by whole-blood PCR. Nat Biotechnol 2022; 40:1680–1689 [View Article] [PubMed]
     [Google Scholar]
  41. Le Bert N, Tan AT, Kunasegaran K, Tham CYL, Hafezi M et al. SARS-CoV-2-specific T cell immunity in cases of COVID-19 and SARS, and uninfected controls. Nature 2020; 584:457–462 [View Article]
     [Google Scholar]
  42. Bergamaschi L, Mescia F, Turner L, Hanson AL, Kotagiri P et al. Longitudinal analysis reveals that delayed bystander CD8+ T cell activation and early immune pathology distinguish severe COVID-19 from mild disease. Immunity 2021; 54:1257–1275 [View Article] [PubMed]
     [Google Scholar]
  43. Collier DA, Ferreira IATM, Kotagiri P, Datir RP, Lim EY et al. Age-related immune response heterogeneity to SARS-CoV-2 vaccine BNT162b2. Nature 2021; 596:417–422 [View Article] [PubMed]
     [Google Scholar]
  44. Mallone R, Mannering SI, Brooks-Worrell BM, Durinovic-Belló I, Cilio CM et al. Isolation and preservation of peripheral blood mononuclear cells for analysis of islet antigen-reactive T cell responses: position statement of the T-Cell Workshop Committee of the Immunology of Diabetes Society. Clin Exp Immunol 2011; 163:33–49 [View Article] [PubMed]
     [Google Scholar]
  45. Davies EL, Noor M, Lim EY, Houldcroft CJ, Okecha G et al. HCMV carriage in the elderly diminishes anti-viral functionality of the adaptive immune response resulting in virus replication at peripheral sites. Front Immunol 2022; 13:1083230 [View Article] [PubMed]
     [Google Scholar]
  46. Binayke A, Zaheer A, Vishwakarma S, Singh S, Sharma P et al. A quest for universal anti-SARS-CoV-2 T cell assay: systematic review, meta-analysis, and experimental validation. NPJ Vaccines 2024; 9:3 [View Article] [PubMed]
     [Google Scholar]
  47. Tiezzi C, Rossi M, Vecchi A, Doselli S, Penna A et al. FluoroSpot assay to analyze SARS-CoV-2-specific T cell responses. STAR Protoc 2023; 4:102584 [View Article] [PubMed]
     [Google Scholar]
  48. Vallejo A, Vizcarra P, Quereda C, Moreno A, Casado JL et al. IFN‐γ + cell response and IFN‐γ release concordance after in vitro SARS‐CoV‐2 stimulation. Eur J Clin Investigation 2021; 51:e13636 [View Article]
     [Google Scholar]
  49. Guo L, Wang G, Wang Y, Zhang Q, Ren L et al. SARS-CoV-2-specific antibody and T-cell responses 1 year after infection in people recovered from COVID-19: a longitudinal cohort study. Lancet Microbe 2022; 3:e348–e356 [View Article] [PubMed]
     [Google Scholar]
  50. Guo L, Zhang Q, Gu X, Ren L, Huang T et al. Durability and cross-reactive immune memory to SARS-CoV-2 in individuals 2 years after recovery from COVID-19: a longitudinal cohort study. Lancet Microbe 2024; 5:e24–e33 [View Article] [PubMed]
     [Google Scholar]
  51. Hellerstein M. What are the roles of antibodies versus a durable, high quality T-cell response in protective immunity against SARS-CoV-2. In Vaccine X 2020 p 100076 [View Article]
     [Google Scholar]
  52. Martínez-Gallo M, Esperalba J, Pujol-Borrell R, Sandá V, Arrese-Muñoz I et al. Commercialized kits to assess T-cell responses against SARS-CoV-2 S peptides. A pilot study in health care workers. Med Clin 2022; 159:116–123 [View Article] [PubMed]
     [Google Scholar]
  53. Meredith RT, Yarham RAR, Mills H, Oliver MA. Whole blood cytokine release assays reveal disparity between capillary blood sampling methods. Clin Biochem 2023; 120:110648 [View Article] [PubMed]
     [Google Scholar]
  54. De Silva TI, Liu G, Lindsey BB, Dong D, Moore SC et al. The impact of viral mutations on recognition by SARS-CoV-2 specific T cells. iScience 2021; 24:103353 [View Article] [PubMed]
     [Google Scholar]
  55. Tye EXC, Jinks E, Haigh TA, Kaul B, Patel P et al. Mutations in SARS-CoV-2 spike protein impair epitope-specific CD4+ T cell recognition. Nat Immunol 2022; 23:1726–1734 [View Article]
     [Google Scholar]
  56. Grifoni A, Weiskopf D, Ramirez SI, Mateus J, Dan JM et al. Targets of T cell responses to SARS-CoV-2 coronavirus in humans with COVID-19 disease and unexposed individuals. Cell 2020; 181:1489–1501 [View Article] [PubMed]
     [Google Scholar]
  57. Mateus J, Grifoni A, Tarke A, Sidney J, Ramirez SI et al. Selective and cross-reactive SARS-CoV-2 T cell epitopes in unexposed humans. Science 2020; 370:89–94 [View Article] [PubMed]
     [Google Scholar]
  58. Nelde A, Bilich T, Heitmann JS, Maringer Y, Salih HR et al. SARS-CoV-2-derived peptides define heterologous and COVID-19-induced T cell recognition. Nat Immunol 2021; 22:74–85 [View Article]
     [Google Scholar]
  59. Gao Z, Xu Y, Sun C, Wang X, Guo Y et al. A systematic review of asymptomatic infections with COVID-19. J Microbiol Immunol Infect 2021; 54:12–16 [View Article] [PubMed]
     [Google Scholar]
  60. Hopkins G, Gomez N, Tucis D, Bartlett L, Steers G et al. Lower humoral and cellular immunity following asymptomatic SARS-CoV-2 infection compared to symptomatic infection in education (The ACE Cohort). J Clin Immunol 2024; 44:147 [View Article] [PubMed]
     [Google Scholar]
  61. Kundu R, Narean JS, Wang L, Fenn J, Pillay T et al. Cross-reactive memory T cells associate with protection against SARS-CoV-2 infection in COVID-19 contacts. Nat Commun 2022; 13:80 [View Article] [PubMed]
     [Google Scholar]
  62. La Gruta NL, Gras S, Daley SR, Thomas PG, Rossjohn J. Understanding the drivers of MHC restriction of T cell receptors. Nat Rev Immunol 2018; 18:467–478 [View Article]
     [Google Scholar]
  63. Braun J, Loyal L, Frentsch M, Wendisch D, Georg P et al. SARS-CoV-2-reactive T cells in healthy donors and patients with COVID-19. Nature 2020; 587:270–274 [View Article]
     [Google Scholar]
  64. Echeverría G, Guevara Á, Coloma J, Ruiz AM, Vasquez MM et al. Pre-existing T-cell immunity to SARS-CoV-2 in unexposed healthy controls in Ecuador, as detected with a COVID-19 Interferon-Gamma Release Assay. Int J Infect Dis 2021; 105:21–25 [View Article] [PubMed]
     [Google Scholar]
  65. Krishna BA, Lim EY, Metaxaki M, Jackson S, Mactavous L et al. Spontaneous, persistent, T cell–dependent IFN-γ release in patients who progress to long covid. Sci Adv 2024; 10:eadi9379 [View Article]
     [Google Scholar]
/content/journal/jgv/10.1099/jgv.0.002055
Loading
Comparison of different T cell assays for the retrospective determination of SARS-CoV-2 infection
J. Gen. Virol. 105, 002055 (2024); https://doi.org/10.1099/jgv.0.002055
/content/journal/jgv/10.1099/jgv.0.002055
/content/journal/jgv/10.1099/jgv.0.002055
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Most cited Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An error occurred
Approval was partially successful, following selected items could not be processed due to error