Assessment of MpoxPlex, a high-throughput and multiplexed immunoassay: a diagnostic accuracy study

Scott Jones; Bethany Hicks; Helen Callaby; Daniel Bailey; N Claire Gordon; Tommy Rampling; Catherine Houlihan; Rachael Jones; Marcus Pond; Ravi Mehta; Deborah Wright; Clarissa Oeser; Simon Tonge; Ezra Linley; Cathy Rowe; Bassam Hallis; Ashley Otter

doi:10.1016/j.lanmic.2024.100987

Assessment of MpoxPlex, a high-throughput and multiplexed immunoassay: a diagnostic accuracy study

Lancet Microbe. 2025 Jan 17:100987. doi: 10.1016/j.lanmic.2024.100987. Online ahead of print.

Authors

Scott Jones¹, Bethany Hicks², Helen Callaby³, Daniel Bailey⁴, N Claire Gordon⁴, Tommy Rampling⁴, Catherine Houlihan⁴, Rachael Jones⁵, Marcus Pond⁶, Ravi Mehta⁶, Deborah Wright⁷, Clarissa Oeser⁸, Simon Tonge⁹, Ezra Linley⁹, Cathy Rowe², Bassam Hallis², Ashley Otter²

Affiliations

¹ Emerging Pathogen Serology Group, Vaccine Development Evaluation and Preparedness Centre, UK Health Security Agency, Wiltshire, UK. Electronic address: [email protected].
² Emerging Pathogen Serology Group, Vaccine Development Evaluation and Preparedness Centre, UK Health Security Agency, Wiltshire, UK.
³ Rare and Imported Pathogens Laboratory, UK Health Security Agency, Wiltshire, UK; Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen, UK.
⁴ Rare and Imported Pathogens Laboratory, UK Health Security Agency, Wiltshire, UK.
⁵ Chelsea and Westminster Hospital NHS Foundation Trust, London, UK.
⁶ Imperial College Healthcare NHS Trust, London, UK.
⁷ Research and Development, Porton Down, UK Health Security Agency, Wiltshire, UK.
⁸ Immunisation and Vaccine Preventable Diseases Division, UK Health Security Agency, Colindale, London, UK.
⁹ Seroepidemiology Unit, UK Health Security Agency, Manchester, UK.

PMID: 39832516
DOI: 10.1016/j.lanmic.2024.100987

Abstract

Background: In May, 2022, the first global outbreak of mpox (formerly known as monkeypox) occurred. In response, public health agencies in the UK have made smallpox vaccines available to individuals at the highest risk of infection. With mpox cases still being detected globally, novel tools are required to aid with diagnosis, serosurveillance, and the evaluation of immune responses following infection and immunisation with current and new vaccine candidates. Here, we describe the development of a multiplexed immunoassay, MpoxPlex, able to measure IgG responses to 12 Orthopoxvirus antigens concurrently and distinguish between responses to infection and vaccination.

Methods: Using the Luminex (DiaSorin, Saluggia, Italy) platform, antibody responses to vaccinia virus (VACV) antigens B5, A27, and A33 and monkeypox virus (MPXV) antigens E8, B6, B2, M1, A27, A35, H3, A29, and A5 were assessed in serum from individuals after MPXV infection (n=24) and after vaccination (n=75) with modified VACV Ankara-Bavarian Nordic. Assay characteristics and cutoffs were calculated by fitting receiver operating characteristic curves to the median fluorescence intensities of these positive samples and negative samples that were run alongside (n=435). P values were calculated using non-parametric Mann-Whitney, Kruskal-Wallis, and Dunn's multiple comparisons tests.

Findings: Using the results from a combination of eight antigens, we were able to distinguish samples as either post-vaccination or post-infection from negative samples with a sensitivity of 98% and a specificity of 95%. IgG responses to MPXV antigen A27 were able to distinguish post-MPXV infection with a sensitivity of 88% and a specificity of 97%. VACV antigen A27 and MPXV antigens A29 and A5 provided little diagnostic advantage.

Interpretation: With additional benefits over current serological assays, we believe this assay will provide substantial insight into the current global outbreak of mpox. MpoxPlex shows use for both serosurveillance and immunological studies of vaccination and infection.

Funding: Grant-in-aid funding to the Emerging Pathogen Serology Group at Porton Down, UK Health Security Agency.