Liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) has significantly grown over the last 20-plus years to become a routine clinical application [1]. As predicted, it has opened the field of analytes that could be successfully measured by mass spectrometry (MS) techniques, e.g. drugs, hormones, and newborn screening and importantly, this technology has provided opportunities to significantly address unmet clinical needs [1], [2], [3], [4], [5], [6]. However, the application in clinical laboratories is still considered highly technical and not applicable to routine 24/7 workflows. Whilst there have been significant process improvements, gaps remain across the total testing process, especially post-analytical, that have continued to limit instrumentation to the specialised areas of the laboratory.
Currently, there remain many challenges with LC-MS/MS that have limited its full translation to the core/central laboratory. These include:
Throughput – LC-MS/MS is associated with batch mode processing that limits the ability to run assays with high sample numbers all by first tier testing e.g. congenital adrenal hyperplasia screening [7];
Post-analytical processing – which remains for many laboratories based on spread sheets or similar as the “middleware” to review output parameters and import them into the laboratory information system;
Service support – mass spectrometry companies have not adapted to the clinical needs of 24/7 support and continue to base their service support on a 9 to 5 model and slow response time for breakdowns; and
Education, training and competence – the level required can be higher than the capacity and broad capability of teams to operate an LC-MS/MS system as a main frame instrument.
So, whilst the dictum of change to MS if it is better than immunoassay holds for the improved specificity, because of the issues noted above, immunoassay remains the predominant first-tier testing principle for steroid analysis, therapeutic drug monitoring (TDM) and toxicology screening. These challenges may be resolved for many routine tests soon.
Building on its Serum Work Area, Roche Diagnostics is making a leap of faith to incorporate automated LC-MS/MS[1] [8, 9]. This has in part been tried previously by others with limited success [10, 11]. With a larger planned initial test menu, the Roche approach pivots away from batch mode processing to a random access testing model, giving the central laboratory a choice between immunoassay and mass spectrometry for more tests. To anchor these methods, as part of the development of this automated solution, Roche is publishing candidate reference measurement procedures (RMP) in preparation for submission to the Joint Committee for Traceability in Laboratory Medicine (JCTLM) database,[2] and as an example, the RMP for methotrexate was published in 2023 and is now included in the database identified as C19RMP11 [12, 13]. The development of RMP to establish metrological traceability was the topic of the editorial associated with Roche’s first set of publications [13], [14], [15], [16], [17], [18], [19].
In this special issue of the Journal, the second collation of five methodology papers describing RMP is presented [20], [21], [22], [23], [24]. These papers continue to address the need for a greater number of Systeme International (SI)-traceable RMP for small molecules. Each paper presents a single analyte isotope dilution mass spectrometry method for the quantification of 1) carbamazepine [20], 2) its main metabolite carbamazepine 10,11-epoxide [21], 3) phenobarbital [22], 4) primidone (which is bio-transformed to phenobarbital) [23], and 5) zonisamide [24] in human serum and plasma. Of these five analytes, only phenobarbital had a RMP listed in the JCTLM previously and two of the Roche methods are newly listed RMP (i.e. carbamazepine – JCTLM ID C20RMP10 and zonisamide JCTLM ID C20RMP10) [12]. Clinically, each of these therapeutic drugs is used alone or in combination with other medications to control certain types of seizures in people with epilepsy.
Epilepsy affects around 50 million people worldwide, affecting individuals from birth through to adults, and contributes 0.5 % to the global burden of disease, and for 70 % of individuals there is the opportunity to live seizure-free if appropriately diagnosed and treated [25]. Antiepileptic TDM has been used to improve the effectiveness and patient safety since the 1960s. Immunoassay-based TDM replaced HPLC-based methods in the mid-1980s because they offered simplicity and sensitivity, but at the expense of specificity due to cross-reactivity [26], [27], [28]. As an example, carbamazepine 10,11-epoxide can interfere with carbamazepine immunoassays that may confound clinical interpretation, limiting the effectiveness of treatment and the potential for adverse outcomes [29]. Hence, with the introduction of LC-MS/MS into clinical laboratories 20-plus years ago, it seemed appropriate to move away from immunoassay. Despite this knowledge, the move to improve specificity has been hampered by the lack of adequate progression to full automation of LC-MS/MS [30, 31].
In summary, automation is a key component of the smart laboratory and its ability to have random access LC-MS/MS assays formatted for antiepileptic TDM is likely to bridge the chasm and allow its widespread adoption [2]. If this initiative is successful, many of the challenges faced by LC-MS/MS (i.e. throughput, post-analytical processing, service support and training requirements) will be dissipated. Whilst automation can bring its own challenges, this more sophisticated approach has the advantages of lean processing efficiency, greenness (through automation) and safety in handling of reagents [32], [33], [34]. The opportunity to have LC-MS/MS run as part of a laboratory’s 24/7 service delivery is likely to be a game changer, in that it is projected to have a profound impact on the central laboratory and its role in the clinical management of epilepsy and beyond.
Acknowledgments
RG has signed a non-disclosure agreement with Roche Diagnostics concerning their new LC-MS/MS system. The information provided in this editorial is in the public domain and provided in good faith to reflect the proposed mass spectrometry product at the time of writing.
References
1. Seger, C, Salzmann, L. After another decade: LC-MS/MS became routine in clinical diagnostics. Clin Biochem 2020;82:2–11. https://doi.org/10.1016/j.clinbiochem.2020.03.004.Search in Google Scholar PubMed
2. Greaves, RF, Bernardini, S, Ferrari, M, Fortina, P, Gouget, B, Gruson, D, et al.. Key questions about the future of laboratory medicine in the next decade of the 21st century: a report from the IFCC-Emerging Technologies Division. Clin Chim Acta 2019;495:570–89. https://doi.org/10.1016/j.cca.2019.05.021.Search in Google Scholar PubMed
3. Greaves, RF, Ho, CS, Hoad, KE, Joseph, J, McWhinney, B, Gill, JP, et al.. Achievements and future directions of the APFCB mass spectrometry harmonisation project on serum testosterone. Clin Biochem Rev 2016;37:63–84.Search in Google Scholar
4. Shackleton, C. Clinical steroid mass spectrometry: a 45-year history culminating in HPLC-MS/MS becoming an essential tool for patient diagnosis. J Steroid Biochem Mol Biol 2010;121:481–90. https://doi.org/10.1016/j.jsbmb.2010.02.017.Search in Google Scholar PubMed
5. Millington, DS, Kodo, N, Norwood, DL, Roe, CR. Tandem mass spectrometry: a new method for acylcarnitine profiling with potential for neonatal screening for inborn errors of metabolism. J Inherit Metab Dis 1990;13:321–4. https://doi.org/10.1007/bf01799385.Search in Google Scholar PubMed
6. Pope, JD, Black, MJ, Drummer, OH, Schneider, HG. Urine toxicology screening by liquid chromatography time-of-flight mass spectrometry in a quaternary hospital setting. Clin Biochem 2021;95:66–72. https://doi.org/10.1016/j.clinbiochem.2021.05.004.Search in Google Scholar PubMed
7. Greaves, RF, Kumar, M, Mawad, N, Francescon, A, Le, C, O’Connell, M, et al.. Best practice for identification of classical 21-hydroxylase deficiency should include 21 deoxycortisol analysis with appropriate isomeric steroid separation. Int J Neonatal Screen 2023;9:58. https://doi.org/10.3390/ijns9040058.Search in Google Scholar PubMed PubMed Central
8. Bonislawski, A. Roche planning 2024 launch of automated mass spec clinical analyzer. 360Dx. 2023 [Accessed 7th April 2024].Search in Google Scholar
9. F. Hoffmann-La Roche Ltd. The future cobas® Mass Spec analyzer*. https://diagnostics.roche.com/global/en/c/mass-spec-analyzer.html [Accessed 19th April 2024].Search in Google Scholar
10. Junger, S, Hoene, M, Shipkova, M, Danzl, G, Schöberl, C, Peter, A, et al.. Automated LC-MS/MS: ready for the clinical routine laboratory? J Mass Spectrom Adv Clin Lab 2023;30:1–9. https://doi.org/10.1016/j.jmsacl.2023.07.001.Search in Google Scholar PubMed PubMed Central
11. Benton, SC, Tetteh, GK, Needham, SJ, Mücke, J, Sheppard, L, Alderson, S, et al.. Evaluation of the 25-hydroxy vitamin D assay on a fully automated liquid chromatography mass spectrometry system, the Thermo Scientific Cascadion SM Clinical Analyzer with the Cascadion 25-hydroxy vitamin D assay in a routine clinical laboratory. Clin Chem Lab Med 2020;58:1010–17. https://doi.org/10.1515/cclm-2019-0834.Search in Google Scholar PubMed
12. Joint Committee for Traceability in Laboratory Medicine (JCTLM). Database of higher-order reference materials, measurement methods/procedures and services. Sevres Cedex, France: International Bureau of Weights and Measures (BIPM).Search in Google Scholar
13. Engel, A, Ruhe, L, Singh, N, Wright, JA, Liesch, F, Bauland, F, et al.. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure (RMP) for the quantification of methotrexate in human serum and plasma. Clin Chem Lab Med 2023;61:1917–29. https://doi.org/10.1515/cclm-2022-1001.Search in Google Scholar PubMed
14. Greaves, RF, Mackay, LG. The development of reference measurement procedures to establish metrological traceability. Clin Chem Lab Med 2023;61:1887–9. https://doi.org/10.1515/cclm-2023-0753.Search in Google Scholar PubMed
15. Taibon, J, Santner, T, Singh, N, Ibrahim, SC, Babitzki, G, Koppl, D, et al.. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure (RMP) for the quantification of aldosterone in human serum and plasma. Clin Chem Lab Med 2023;61:1902–16. https://doi.org/10.1515/cclm-2022-0996.Search in Google Scholar PubMed
16. Salzmann, L, Spescha, T, Singh, N, Schierscher, T, Bachmann, M, Bauland, F, et al.. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure (RMP) for the quantification of lamotrigine in human serum and plasma. Clin Chem Lab Med 2023;61:1930–41. https://doi.org/10.1515/cclm-2022-0997.Search in Google Scholar PubMed
17. Salzmann, L, Wild, J, Singh, N, Schierscher, T, Liesch, F, Bauland, F, et al.. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure (RMP) for the quantification of gabapentin in human serum and plasma. Clin Chem Lab Med 2023;61:1955–66. https://doi.org/10.1515/cclm-2022-0998.Search in Google Scholar PubMed
18. Kobel, A, Schierscher, T, Singh, N, Salzmann, L, Liesch, F, Bauland, F, et al.. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of levetiracetam in human serum and plasma. Clin Chem Lab Med 2023;61:1967–77. https://doi.org/10.1515/cclm-2022-1038.Search in Google Scholar PubMed
19. Salzmann, L, Spescha, T, Singh, N, Kobel, A, Fischer, V, Schierscher, T, et al.. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of topiramate in human serum and plasma. Clin Chem Lab Med 2023;61:1942–54. https://doi.org/10.1515/cclm-2022-1273.Search in Google Scholar PubMed
20. Schierscher, T, Salzmann, L, Singh, N, Bachmann, M, Bauland, F, Geistanger, A, et al.. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of carbamazepine in human serum and plasma. Clin Chem Lab Med 2023. https://doi.org/10.1515/cclm-2023-0943.Search in Google Scholar PubMed
21. Schierscher, T, Singh, N, Kobel, A, Wild, J, Bauland, F, Geistanger, A, et al.. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of carbamazepine-10,11-epoxide in human serum and plasma. Clin Chem Lab Med 2024. https://doi.org/10.1515/cclm-2023-1045.Search in Google Scholar PubMed
22. Schierscher, T, Salzmann, L, Singh, N, Bachmann, M, Kobel, A, Wild, J, et al.. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of phenobarbital in human serum and plasma. Clin Chem Lab Med 2024. https://doi.org/10.1515/cclm-2023-1104.Search in Google Scholar PubMed
23. Schierscher, T, Salzmann, L, Singh, N, Fischer, V, Kobel, A, Bauland, F, et al.. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure (RMP) for the quantification of primidone in human serum and plasma. Clin Chem Lab Med 2024. https://doi.org/10.1515/cclm-2023-1032.Search in Google Scholar PubMed
24. Schierscher, T, Salzmann, L, Singh, N, Wild, J, Fischer, V, Bauland, F, et al.. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of zonisamide in human serum and plasma. Clin Chem Lab Med 2023. https://doi.org/10.1515/cclm-2023-0736.Search in Google Scholar PubMed
25. WHO. Epilepsy. https://www.who.int/news-room/fact-sheets/detail/epilepsy#:∼:text=The%20estimated%20proportion%20of%20the,diagnosed%20with%20epilepsy%20each%20year [Accessed 19th April 2024].Search in Google Scholar
26. Contin, M, Riva, R, Albani, F, Perucca, E, Baruzzi, A. Determination of total and free plasma carbamazepine concentrations by enzyme multiplied immunoassay: interference with the 10,11-epoxide metabolite. Ther Drug Monit 1985;7:46–50. https://doi.org/10.1097/00007691-198503000-00007.Search in Google Scholar PubMed
27. Eadie, MJ. Therapeutic drug monitoring – antiepileptic drugs. Br J Clin Pharmacol 1998;46:185–93. https://doi.org/10.1046/j.1365-2125.1998.00769.x.Search in Google Scholar PubMed PubMed Central
28. Kang, J, Park, YS, Kim, SH, Kim, SH, Jun, MY. Modern methods for analysis of antiepileptic drugs in the biological fluids for pharmacokinetics, bioequivalence and therapeutic drug monitoring. Korean J Physiol Pharmacol 2011;15:67–81. https://doi.org/10.4196/kjpp.2011.15.2.67.Search in Google Scholar PubMed PubMed Central
29. Dasgupta, A. Chapter 2 – immunoassays and issues with interference in therapeutic drug monitoring. In: Clarke, W, Dasgupta, A, editors. Clinical challenges in therapeutic drug monitoring. San Diego: Elsevier; 2016:17–44 pp.10.1016/B978-0-12-802025-8.00002-7Search in Google Scholar
30. Antonelli, G, Marinova, M, Artusi, C, Plebani, M. Mass spectrometry or immunoassay: est modus in rebus. Clin Chem Lab Med 2017;55:1243–5. https://doi.org/10.1515/cclm-2017-0197.Search in Google Scholar PubMed
31. Cross, TG, Hornshaw, MP. Can LC and LC-MS ever replace immunoassays? J Appl Bioanal 2016;2:108–16. https://doi.org/10.17145/jab.16.015.Search in Google Scholar
32. Liker, JK. The Toyota way: 14 management principles from the World’s greatest manufacturer, 2nd ed. New York: McGraw-Hill; 2021. https://www.accessengineeringlibrary.com/content/book/9781260468519.Search in Google Scholar
33. Pena-Pereira, F, Wojnowski, W, Tobiszewski, M. AGREE – Analytical GREEnness metric approach and software. Anal Chem 2020;92:10076–82. https://doi.org/10.1021/acs.analchem.0c01887.Search in Google Scholar PubMed PubMed Central
34. Lippi, G, Da Rin, G. Advantages and limitations of total laboratory automation: a personal overview. Clin Chem Lab Med 2019;57:802–11. https://doi.org/10.1515/cclm-2018-1323.Search in Google Scholar PubMed
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