A high throughput immuno-affinity mass spectrometry method for detection and quantitation of SARS-CoV-2 nucleoprotein in human saliva and its comparison with RT-PCR, RT-LAMP, and lateral flow rapid antigen test
A high throughput immuno-affinity mass spectrometry method for detection and quantitation of SARS-CoV-2 nucleoprotein in human saliva and its comparison with RT-PCR, RT-LAMP, and lateral flow rapid antigen test
Objectives: many reverse transcription polymerase chain reaction (RT-PCR) methods exist that can detect SARS-CoV-2 RNA in different matrices. RT-PCR is highly sensitive, although viral RNA may be detected long after active infection has taken place. SARS-CoV-2 proteins have shorter detection windows hence their detection might be more meaningful. Given salivary droplets represent a main source of transmission, we explored the detection of viral RNA and protein using four different detection platforms including SISCAPA peptide immunoaffinity liquid chromatography-mass spectrometry (SISCAPA-LC-MS) using polyclonal capture antibodies.
Methods: the SISCAPA-LC MS method was compared to RT-PCR, RT-loop-mediated isothermal amplification (RT-LAMP), and a lateral flow rapid antigen test (RAT) for the detection of virus material in the drool saliva of 102 patients hospitalised after infection with SARS-CoV-2. Cycle thresholds (Ct) of RT-PCR ( E gene) were compared to RT-LAMP time-to-positive (TTP) ( NE and Orf1a genes), RAT optical densitometry measurements (test line/control line ratio) and to SISCAPA-LC-MS for measurements of viral protein.
Results: SISCAPA-LC-MS showed low sensitivity (37.7 %) but high specificity (89.8 %). RAT showed lower sensitivity (24.5 %) and high specificity (100 %). RT-LAMP had high sensitivity (83.0 %) and specificity (100.0 %). At high initial viral RNA loads (<20 Ct), results obtained using SISCAPA-LC-MS correlated with RT-PCR (R 2 0.57, p-value 0.002).
Conclusions: detection of SARS-CoV-2 nucleoprotein in saliva was less frequent than the detection of viral RNA. The SISCAPA-LC-MS method allowed processing of multiple samples in <150 min and was scalable, enabling high throughput.
immuno-affinity, mass spectrometry, rapid antigen test, RT-LAMP, RT-PCR, SARS-CoV-2
Lane, Dan
63c76b9b-3fc0-49a5-a066-8a07a33fe214
Allsopp, Rebecca
450376b2-75af-445b-879e-afd8072f519f
Holmes, Christopher W.
63e0b32f-df81-4174-819b-c361fcdf7cce
Skipp, Paul
1ba7dcf6-9fe7-4b5c-a9d0-e32ed7f42aa5
Lane, Dan
63c76b9b-3fc0-49a5-a066-8a07a33fe214
Allsopp, Rebecca
450376b2-75af-445b-879e-afd8072f519f
Holmes, Christopher W.
63e0b32f-df81-4174-819b-c361fcdf7cce
Skipp, Paul
1ba7dcf6-9fe7-4b5c-a9d0-e32ed7f42aa5
Lane, Dan, Allsopp, Rebecca and Holmes, Christopher W.
,
et al.
(2024)
A high throughput immuno-affinity mass spectrometry method for detection and quantitation of SARS-CoV-2 nucleoprotein in human saliva and its comparison with RT-PCR, RT-LAMP, and lateral flow rapid antigen test.
Clinical Chemistry and Laboratory Medicine.
(doi:10.1515/cclm-2023-0243).
Abstract
Objectives: many reverse transcription polymerase chain reaction (RT-PCR) methods exist that can detect SARS-CoV-2 RNA in different matrices. RT-PCR is highly sensitive, although viral RNA may be detected long after active infection has taken place. SARS-CoV-2 proteins have shorter detection windows hence their detection might be more meaningful. Given salivary droplets represent a main source of transmission, we explored the detection of viral RNA and protein using four different detection platforms including SISCAPA peptide immunoaffinity liquid chromatography-mass spectrometry (SISCAPA-LC-MS) using polyclonal capture antibodies.
Methods: the SISCAPA-LC MS method was compared to RT-PCR, RT-loop-mediated isothermal amplification (RT-LAMP), and a lateral flow rapid antigen test (RAT) for the detection of virus material in the drool saliva of 102 patients hospitalised after infection with SARS-CoV-2. Cycle thresholds (Ct) of RT-PCR ( E gene) were compared to RT-LAMP time-to-positive (TTP) ( NE and Orf1a genes), RAT optical densitometry measurements (test line/control line ratio) and to SISCAPA-LC-MS for measurements of viral protein.
Results: SISCAPA-LC-MS showed low sensitivity (37.7 %) but high specificity (89.8 %). RAT showed lower sensitivity (24.5 %) and high specificity (100 %). RT-LAMP had high sensitivity (83.0 %) and specificity (100.0 %). At high initial viral RNA loads (<20 Ct), results obtained using SISCAPA-LC-MS correlated with RT-PCR (R 2 0.57, p-value 0.002).
Conclusions: detection of SARS-CoV-2 nucleoprotein in saliva was less frequent than the detection of viral RNA. The SISCAPA-LC-MS method allowed processing of multiple samples in <150 min and was scalable, enabling high throughput.
Text
A high throughput immuno-affinity mass spectrometry method for detection and quantitation of SARS-CoV-2 - accepted
- Accepted Manuscript
More information
Accepted/In Press date: 12 December 2022
e-pub ahead of print date: 23 January 2024
Additional Information:
Funding Information:
This research was funded by the UK Medical Research Council Confidence in Concept (MRC CiC) Funding (Grant Number: MC_PC_18054). We acknowledge the support of the NIHR Leicester Biomedical Research Centre and the John and Lucille van Geest Foundation.
Keywords:
immuno-affinity, mass spectrometry, rapid antigen test, RT-LAMP, RT-PCR, SARS-CoV-2
Identifiers
Local EPrints ID: 487958
URI: http://eprints.soton.ac.uk/id/eprint/487958
ISSN: 1434-6621
PURE UUID: 89faf02f-408c-43fa-8b57-cacaed550f91
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Date deposited: 11 Mar 2024 17:49
Last modified: 01 May 2024 01:34
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Contributors
Author:
Dan Lane
Author:
Rebecca Allsopp
Author:
Christopher W. Holmes
Corporate Author: et al.
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