Liquid Chromatography-Mass Spectrometry (LC-MS) Analysis In Therapeutic .

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Liquid Chromatography-MassSpectrometry (LC-MS)Analysis in TherapeuticDrug Monitoring‘The second eBook of the Hamilton series’

ForewordThe efficacy and clinical utility of therapeutic drug monitoring (TDM) relies onthe specific, sensitive, and reproducible measurement of analytes. Liquidchromatography coupled with mass spectrometry (LC-MS) has emerged asan attractive alternative to classical immunoassays. Currently, hundreds oftherapeutic drugs are analyzed daily in the clinical setting using this techniqueand several in vitro diagnostic (IVD) certified kits have entered the market.We believe that LC-MS has the potential to become the gold standard for TDM,and therefore we have collaborated with leaders in the field to make sure ourautomation solutions are compatible with their kits, workflows and instruments.In this eBook, we discuss the impact of LC-MS in TDM, the drivers behind theneed for automation during sample preparation for LC-MS and the solutions thatHamilton offers to customers working in this field.This eBook is part of a dedicated campaign for LC-MS analysis in TDM, where weaim to provide our readers with interesting educational resources and additionalinsights into the way our customers are using Hamilton solutions to accomplishtheir tasks.I want to thank our Senior Market Segment Leader Analytics and our InternationalCommercial Leader Diagnostics for their valuable contributions to this eBook.We hope you find the content beneficial.Your kind feedback is always highly appreciated.Your sincerely,Dr. Gabriela Boza-MoranScientific Content ManagerHamilton Robotics Direct Salesmboza-moran@hamilton.ch2

Table ofContentsExploring the Role of LC-MSin Therapeutic Drug Monitoring .04LC-MS: Why Do We Need Automation? .08What Can Hamilton Offer?.11Application Note 1: Automation Sample Preparationfor Immunosuppresant Analysis.15Application Note 2: Automated Sample Preparationfor Vitamin D3/D2 Analysis.19MassSTAR:Dual MassSTAReasyPunchCE-IVD solution for LC-MS samplepreparation (Chromsystems’ kits)RUO solution for LC-MS samplepreparationRUO solution for samplepunching and processingSee page 113

LIQUID CHROMATOGRAPHY-MASS SPECTROMETRY (LC-MS)Exploring the Role of LC-MSin Therapeutic Drug MonitoringTherapeutic drug monitoring (TDM) refers to the repeatedmeasurement of an analyte (a therapeutic drug) in a patient’sblood or plasma, with the aim of finding the dosage regimenneeded to maintain therapeutic concentrations. TMD is thefocus of clinical pharmacology, an area that studies theeffects of the body on drugs (pharmacokinetics) and theeffects of drugs on the body (pharmacodynamics).1 TDM isgenerally required for drugs with a narrow therapeutic indexand/or severe side effects such as immunosuppressants.2For drugs with a narrow therapeutic index,small variations in dose or blood concentrationmay cause therapeutic failures or adversedrug reactions.Historically, immunoassays have been the gold standard forthe analysis of molecules in clinical chemistry. These testsare based on antibody–antigen binding, however they aresubject to several limitations3 such as: (1) limited specificity,due to cross-reactions; (2) limited reproducibility, due toantibody lot-to-lot variations (from the same manufacturerand/or discordance between the results obtained fromassays from different manufacturers); and (3) limitedsensitivity, due to the low linear dynamic range. Furthermore,since every analyte requires a specific antibody, many donot have a commercially available immunoassay.Liquid chromatography coupled with mass spectrometry(LC-MS) has emerged as an attractive alternative to classicalimmunoassays and other popular enzyme-subtract tests, asit does not suffer the same limitations found in immunoassays.LC-MS is highly (1) specific and (2) reproducible (as itidentifies compounds based on their unique chemicalstructure without the need to use antibodies), as well as(3) sensitive (due to its high linear dynamic range. Theseunique features enable the detection of steroids and otheranalytes that are difficult to measure at low concentrations.Additionally, LC-MS allows the simultaneous analysis ofseveral (up to hundreds) compounds from the same sampleduring the same run.4,5,6 All these advantages have madeLC-MS very attractive in the clinical setting, where thousandsof samples (blood, plasma, serum, urine, hair, spittle andmilk) are analyzed every year for the assessment for multipleparameters.7The initial adoption of MS in the clinical setting waslimited by the technical complexities of the MS techniqueused in the 1980s–’90s (mainly gas chromatography–mass spectrometry or GC-MS). The new (soft) ionizationtechniques, the high performance of the system oncecoupled with front end liquid chromatography (LC) and asecond mass analyzer (MS/MS), gave MS the conditionsnecessary to be a viable and attractive alternative for routineclinical laboratories.8Initially, the use of LC-MS in TDM was focused onimmunosuppressive drugs (also known as immuno suppressants), however its use has now extended to theassessment of anticonvulsants, antibiotics, antifungaldrugs, antiviral drugs, antidepressants, anti-cancer drugsand cardiac drugs, among others.6,11 LC-MS is particularlysuited for the clinical setting when using tandem massspectrometry (LC-MS/MS) and performing selected/multiple4

reaction monitoring (SRM/MRM), a technique that selects theanalytes to be analyzed and quantified with high precision.9LC-MS/MS (particularly triple quadrupole MS) is currentlyone of the most popular methods for TDM.Laboratories select clinical tests based on their individualrequirements. From a regulatory point of view, it is notuncommon for clinical laboratories to use laboratorydeveloped tests (LDTs, also known as “in-house tests”or “homebrew tests”), which — unlike in vitro diagnostics(IVD) certified tests — do not need US Food and DrugAdministration (FDA) clearance or a European CE-IVD mark.An LDT is an in vitro diagnostic test designed, manufacturedand used within a single laboratory.12 Their use in the clinicis expected to be limited in the future. By contrast, an IVDcertified test is an in vitro diagnostic test “sold as a completekit that a laboratory purchases from a manufacturer, andcomes with all of the procedures and controls to performthe test”.13 Some commercial CE-IVD kits for TDM areMassTox TDM Series A (Chromsystems Instruments &Chemicals GmbH),14 ClinMass Complete Kit, advanced, forimmunosuppressants in whole blood (RECIPE Chemicals Instruments GmbH)15 and MassTrak Immunosuppressantkit (Waters, M., & Watson, I. (2017). Therapeutic Drug Monitoring: Clinical Guide. Fourth Edition. Retrieved 19 June 2020, from e/ADD-00061347 TDM Learning Guide.pdfJunaid, T., Wu, X., Thanukrishnan, H., & Venkataramanan, R. (2019). Therapeutic Drug Monitoring. Clinical Pharmacy Education, Practice andResearch, 425-436. doi: 10.1016/b978-0-12-814276-9.00030-1National Cancer Institute. (2020). NCI Dictionary of Cancer Terms. Retrieved 19 June 2020, from ancer-terms/def/immunoassayJannetto, P. (2015). Liquid Chromatography Tandem Mass Spectrometry. Retrieved 19 June 2020, from romatography-tandem-mass-spectrometry.htmlBazydio, L., & Mullins, G. (2016). Technical Comparison of Immunoassay and Mass Spectrometry : June 2016 - MedicalLab ManagementMagazine. Retrieved 19 June 2020, from der Gugten, J. (2020). Tandem mass spectrometry in the clinical laboratory: A tutorial overview. Clinical Mass Spectrometry, 15, 36-43. doi:10.1016/j.clinms.2019.09.002Keevil, B., Owen, L., & Adaway, J. (2015). Integrating LC-MS/MS into the Clinical Laboratory. Retrieved 19 June 2020, from gyV1EGzMx4Jannetto, P., & Fitzgerald, R. (2016). Effective Use of Mass Spectrometry in the Clinical Laboratory. Clinical Chemistry, 62(1), 92-98. doi:10.1373/clinchem.2015.248146Grebe, S., & Singh, R. (2011). LC-MS/MS in the Clinical Laboratory – Where to From Here?. Retrieved 19 June 2020, from 91/van der Gugten, J. (2020). Tandem mass spectrometry in the clinical laboratory: A tutorial overview. Clinical Mass Spectrometry, 15, 36-43. doi:10.1016/j.clinms.2019.09.002Garg, U., & Zhang, Y. (2016). Mass Spectrometry in Clinical Laboratory: Applications in Therapeutic Drug Monitoring and Toxicology. Methodsin Molecular Biology, 1-10. doi: 10.1007/978-1-4939-3252-8 1FDA. (2018). Laboratory Developed Tests. Retrieved 19 June 2020, from ics/laboratorydeveloped-testsMamuszka, H. (2019). The Neverending LDT vs IVD Debate. Retrieved 19 June 2020, from tems. (2020). Therapeutic Drug Monitoring by LC-MS/MS. Retrieved 19 June 2020, from drug-monitoring/lc-msms.htmlRECIPE. n.d. Immunosuppressants In Whole Blood, Advanced RECIPE. [online] Available at: ced/ [Accessed 19 June 2020].MassTrak Immunosuppressants Kit: Waters. (2020). Retrieved 19 June 2020, from GB/MassTrakImmunosuppressants-Kit/nav.htm?cid 10010989&lset 1&locale en GB5

LC-MS FOR THERAPEUTICDRUG MONITORING IN THECLINICAL SETTINGTherapeutic drug monitoring (TDM) is a clinical practice used to optimize individual dosing regimens by measuring specificdrugs at designated intervals to maintain their concentration within a target range.1,2 The need for accurate, precise, andstandardized measurement of drugs presents a major challenge for clinical laboratories and the diagnostics industry.3To be suitable for TDM, a drug should satisfy certain criteria including:A narrowtherapeutictarget range1SignificantpharmacokineticvariabilityA reasonablerelationshipbetween bloodconcentrationsand clinical effectsEstablishedtherapeutic rangeand toxic thresholdAvailabilityof cost-effectivedrug assayA diagnosis is made and a drug is selected.A dosage schedule is designed to reach atarget therapeutic plasma concentration.The drug is administered to the patient.11122Samples are collected from the patient atrepeated intervals and tested in a clinicallaboratory using liquid chromatographycoupled with mass spectrometry (LC-MS).2*Please note that other analytical methods also exist.2443Drug concentrations in the patient’s blood/plasma is determined. A pharmacokineticmodel is applied and clinical judgement isused to adjust the dosage if necessary.Onsetofeffect4Side effectsPeak effectAdverse reactionTherapeutic WindowDesired responseDuration ofactionSubtherapeutic

WHAT IS LIQUIDCHROMATOGRAPHY-MASSSPECTROMETRY?Liquid chromatography coupled with mass spectrometry (LC-MS) is a highly specific, sensitive and reproducibleanalytical technique that combines the physical separation capabilities of liquid chromatography with the massanalysis capabilities of mass spectrometry (MS).4111Samples are extracted/prepared for analysis andthen separated using ahigh performance liquidchromatography (HPLC)column.5From HPLCsystemsHPLC System2Mass SpectrometerThe separated samplespecies are sprayed into anion source while transitioninginto a gas phase.5 Samplescan be ionized throughseveral methods. The mostcommon for TDM analysis iselectrospray ionization (ESI)3Mass spectrometryseparates gas phase ionsaccording to their m/z (massto charge ratio) value, usingelectrical and/or magneticfields to differentiate ions.5,6There are different typesof mass analysers. Oneof the most common forTDM analysis is the triplequadrupole.22 Ion Source4 Detector3 Mass Analysers4As ions cross the massanalyzer, the detectorcounts them and amplifiestheir signal.54Computer1. Ghiculesco, R., 2008. Abnormal laboratory results: Therapeutic drug monitoring: which drugs,why, when and how to do it. Australian Prescriber, [online] 31(2), pp.42-44. Available at: n-and-how-to-do-it#article [Accessed 24 June 2020].2. Kang, J. and Lee, M., 2009. Overview of Therapeutic Drug Monitoring. The Korean journal ofinternal medicine, [online] 24(1), p.1. Available at: 54/ [Accessed 24 June 2020].3. Brandhorst, G., Oellerich, M., Maine, G., Taylor, P., Veen, G. and Wallemacq, P., 2012. LiquidChromatography–Tandem Mass Spectrometry or Automated Immunoassays: What Are theFuture Trends in Therapeutic Drug Monitoring?. Clinical Chemistry, [online] 58(5), pp.821-825.Available at: 7fce8984d6f496394ce2f887.pdf [Accessed 24 June 2020].4. Technology Networks. 2020. Liquid Chromatography-Mass Spectrometry (LC-MS) For Oligonucleotide Analysis. [online] Available at: ms-for-oligonucleotide-analysis-330452 [Accessed 24 June 2020].5. CHROMacademy. 2020. Mass Spectrometry Fundamental LC-MS Introduction. [online] Available at: emical/Openlit/Chromacademy%20LCMS%20Intro.pdf [Accessed 24 June 2020].

LCMS IN AUTOMATIONLC-MS: Why Do We NeedAutomation?There has been a steep adoption of LC-MS-based testing inthe clinical setting over the past few years. Central laboratories,such as the University Hospital of South Manchester, UKprocess between 2,000 to 3,000 samples per day (analyzingseveral analytes per sample).1,2 Automation, particularlyduring the sample preparation step, can significantly increasethe throughput of clinical laboratories and provide researcherswith more time to focus on more valuable tasks.Sample preparation is a critical step in LC-MS methodology.Due to the specific technology used for detection, massspectrometers are very sensitive to the presence of saltsand charged ions. Additionally, the detection of lowabundance molecules can be masked by the presence ofhigh abundance ones.3 The quality and reproducibility of thesample preparation step therefore has a significant influenceon the final results. This is particularly critical in the clinicalsetting, where the results obtained from the tests have adirect influence on the patient’s treatment.There is no standard or universal method for the preparationof samples for LC-MS analysis in the clinical setting.Depending on the type and concentration of the analyte,the matrix, the goal of the assessment and the analyticalmethod used, samples might require very different treatment(reduction and alkylation, digestion, desalting, proteinprecipitation, solid-phase extraction, etc.).4,5 In the routineclinical setting, the workflows for each type of measurementneed to be standardized to ensure reproducibility. Yet thisis often hindered by manual processing errors. Even withthe availability of experienced, qualified staff (which is animportant source of fixed costs), inter-operator variabilityremains. Whilst high throughput is considered the main sellingpoint of automation, in the clinical practice, the need for highreproducibility is the main reason for transitioning from manualto automated workflows.Current regulatory changes also require a higher level ofcompliance for the diagnostic tests performed in the clinicalsetting (beyond investigational use), including LC-MS-basedtests. At present, most clinical laboratories develop andvalidate their LC-MS assays LDTs. In the US, laboratories witha Clinical Laboratory Improvement Amendments (CLIA) licenseare allowed to develop and use their own assays, without theneed to obtain an IVD certification.6 Even though the FDAhas expressed the intention to better regulate LDTs, no finalguidance has been issued since the initial draft guidance of2014.7 It must be highlighted, however, that despite the lackof final guidance, the FDA can, and already does, exercise“enforcement discretion” when there are concerns about thevalidity of the results provided by an LDT.6In Europe, the conditions for a much higher regulatory over sight were announced back in 2017, and they are expectedto have a significant impact in the way clinical laboratorieswork in the future. Currently, LDTs are developed and used incertified labs, without the need for a CE-IVD mark.8 Yet from2022, only LDTs for which an equivalent CE-IVD test does notexist, will be permitted for use.8

The new EU regulation (EU 2017/746) covers the regulatorycontrol for LDT and now ensures stricter measures for clinicalIVD tests.9 These measures cover the tests, kit, instruments,and the users. Automation will help clinical laboratories complywith these new regulations to ensure: TraceabilityStandardization of workflowsElimination/ minimization of human errorControl of user accessWorkflows for handling errorslaboratories need to seriously consider the instrument provider,as this will impact the complexity of any future LDT validationprocess.“Platform selection is one of the most criticalchoices a diagnostic developer can makebecause it is effectively a partner for life”– Hannah Mamuszka, CEO, Alva 106Reimbursement policies are expected to be based in theuse of IVD tests complying with the new regulations. ManyReferences1., B., Owen, L., & Adaway, J. (2015). Integrating LC-MS/MS into the Clinical Laboratory. Retrieved 19 June 2020, from gyV1EGzMx4Garg, U., & Zhang, Y. (2016). Mass Spectrometry in Clinical Laboratory: Applications in Therapeutic Drug Monitoring and Toxicology. Methodsin Molecular Biology, 1-10. doi: 10.1007/978-1-4939-3252-8 1Reubsaet, L. (2016). Determination of Very Low-Abundance Diagnostic Proteins in Serum Using Immuno-Capture LC–MS–MS. Retrieved19 June 2020, from -immunocapture-lc-ms-ms?pageID 2Thermoscientific. (2020). Sample Preparation for Mass Spectrometry Thermo Fisher Scientific - UK. Retrieved 19 June 2020, from thods/sample-preparation-mass-spectrometry.htmlvan der Gugten, J. (2020). Tandem mass spectrometry in the clinical laboratory: A tutorial overview. Clinical Mass Spectrometry, 15, 36-43. doi:10.1016/j.clinms.2019.09.002Mamuszka, H. (2019). The Neverending LDT vs IVD Debate. Retrieved 19 June 2020, from ontent/uploads/2019/06/jpm219-Mamuszka.pdfFDA. (2018). Laboratory Developed Tests. Retrieved 19 June 2020, from ics/laboratorydeveloped-testsInstitut, J. (2020). Laboratory Developed Tests LDT Game Changer for Medical Labs. Retrieved 19 June 2020, from lation. (2017). REGULATION (EU) 2017/746 OF THE EUROPEAN PARLIAMENT AND OF THE COUNCIL. Retrieved 19 June 2020, /?uri CELEX:02017R0746-201705059

ENSURINGCOMPLIANCE INTDM THROUGHAUTOMATIONCurrently, most clinicallaboratories develop andvalidate their own LC-MSassays.Liquid chromatography-mass spectrometry (LC-MS)based testing is increasing in popularity for clinicallaboratories conducting therapeutic drug monitoring(TDM). In response to an increase in this and other typesof clinical tests, European regulations for diagnostic testsperformed in the clinic will change. Stricter regulationmeasures are also still in discussion in the US.Clinical laboratories need to adapt to stricter regulatorycontrol for laboratory developed tests (LDTs), whichcover the tests, kit, instruments, and the users.In the US, laboratorieswith a Clinical LaboratoryImprovement Amendments(CLIA) license can developand use their assays, withoutthe need to obtain an IVDcertification.In 2017, Europe introducedthe EU 2017/746, whichcame with new, harsherregulatory measures. Thesenew regulations will applyfrom 2022 onwards.Currently in Europe, LDTsare developed and used incertified labs, without theneed for a CE-IVD mark.In 2022, only LDTs for whichan equivalent CE-IVD testdoes not exist, will bepermitted for use in Europe.Automation can be introduced to clinical laboratories to help optimize their testing andincrease compliance with these regulations, providing:Better (end-to-end)traceabilityControlled useraccessStandardization ofworkflowsRobust workflows forhandling errorsElimination/minimizationof human errorIncreased throughputBetter reproducibility

HAMILTON’S SOLUTIONSWhat Can Hamilton Offer?Hamilton is a market leader in precision liquid handling, whereinnovation is not only a key internal value, but also a value thatwe strongly support in the fields we serve. Sample preparationfor LC-MS is one our focus areas. LC-MS has the potential tobecome the gold standard for TDM, and we have collaboratedwith many leaders in this field (Chromsystems, Recipe, Sciex,Waters, Shimazu, ThermoFisher and Agilent, among others) tomake sure our solutions are compatible with their kits, workflowsand instruments.Our solutions are designed with laboratory needs in mind andour philosophy is to provide our customers with the widestrange of options — from turnkey to customized solutions,and from IVD-certified solutions to solutions supporting theautomation of laboratory-developed tests (LDTs). Our mostpopular sample preparation workflows are designed on ourMicrolab STAR line; however, our solutions can be adaptedto any of our other platforms.For customers seeking ready-to-use solutions withfast implementationWe offer an assay ready workstation (ARW) designedspecifically for sample preparation for LC-MS with kits fromChromsystems: the MassSTAR. The MassSTAR is a turnkeysolution with a standardized hardware and software packageand a fixed deck-layout. More importantly, the MassSTARis a CE-IVD certified solution validated for the analysis ofimmunosuppressants and Vitamin D (with the MassTox immunosuppressants and MassChrom 25-OH-VitaminD3/D2 kits, respectively). Hamilton and Chromsystemsspecifically developed this ARW for the diagnostic market.The features of the MassSTAR, including automatic loading,full traceability of the samples, user access control andprotocol set-up for error-handling, comply with regulatorystandards.The MassSTAR can process up to 288 samples per batchin 2 hours and 30 minutes for the immunosuppressantstesting or 4 hours and 30 minutes for the Vitamin D testing.Please note that we do not recommend running more than192 samples at the same time. New kits are currently beingvalidated and certified for the analysis of other molecules.For customers interested in automating their own solutionsstarting from a pre-defined layout, we offer the DualMassSTAR, which is an open platform. Our collaboratorsin Labor Berlin have validated workflows for the analysisof several molecules relevant for TDM (antibiotics,antidepressants, antiepileptics and neuropileptics) usingthis platform. We can replicate or adapt these conditionsand layouts for our customers to allow for more rapidimplementation.For customers looking for flexibilityWe know that each customer is different and we aim toprovide solutions for individual needs. We can providefully customizable deck layouts, including direct sampleinjection into the LC-MS and integration of our variousstandard modules for incubation, shaking, centrifugationand solid-phase extraction (SPE), as well as integrationwith third-party devices. One of our most popular modulesis the Monitored Multi-flow, Positive Pressure EvaporativeExtraction – [MPE]2 – for SPE. The positive pressure systemof our [MPE]2 maintains equal pressure across the filterplate, eliminating the path of least resistance. Furthermore,the [MPE]2 can be ordered with the complementary reagentdispenser to automatically add liquid to the plate, as well aswith the complementary evaporator to dry down sampleswith a heated gas. We also have a vacuum-based SPEmodule for customers preferring this solution.11

HAMILTON’S SOLUTIONSCalibrators &controlsTipsHeater shakerCentrifugeFilter plates (3)Collectionplates (3)Counterweight platesSampleReagents (3)Figure 1: MassSTAR, Deck Layout.TipsReagentcarrierSamplepreparations plateSample loadingpositionsLid parkpositionHeater shakerCentrifugeCounterbalance plateCollection tubes inCobas racksMFX coolingmoduleCVS vacuumstationFigure 2: Dual MassSTAR, Deck Layout.12

HAMILTON’S (optional)LogisticsmoduleAir input fitting96, 48, 24replaceablemanifoldsReagent rack(optional)Evaporatormodule(optional)Labware shuttle/waste receptacle/pipette ntpump(optional)Patented dualstage elevatorACMEmodule(optional)ControlBoxFigure 3: [MPE]2 module. Left: plate logistics Module. Right: components of the full system.For customers working with lower throughputs, we offer ourFlipTube’s solutions. Instead of using plates, samples canbe processed in 1.5 ml FlipTubes specially designed witha unique lid that prevents contamination and evaporation.Our platforms can be equipped with the FlipTube tool-setthat automates the opening and closing of the FlipTubes,as well as with transport tools, racks, adaptors, carriersand dedicated shakers and microcentrifuges that allowfull compatibility with most of the workflows for samplepreparation.For customers looking to process dried blood samples, we offerthe easyPunch STARlet, a specialized platform for automatedsample punching and processing. The easyPunch recognizespositively stained samples (e.g. blood on a white card) andnegatively stained sample (e.g. saliva, plasma or urine onindicating cards) and makes punches of 1.2, 2, 3 and 6 mmdirectly into collection plates. The process is fully traceable.The final decision about which instrument/ARW to use willdepend on the specific needs of the customer.FlipTube toolFlipTube rackFlipTube erFigure 4: FlipTube’s solutions.13

HAMILTON’S SOLUTIONSPunchingmoduleCard magazinesPlate slideCard & plategripperAuxiliary cardrecovery magazineMicrotubesTips & 50mltroughsMTPsFigure 5: easyPunch, Deck Layout.Independent of the selection, all our solutions ensure: Precision, accurate pipetting achieved through ourvarious proprietary technologies:1. Compression-Induced O-ring Expansion (CO-RE), fortip attachment and positioning2. Liquid Level Detection (LLD), to detect the exact levelof liquids in tubes or plates3. Anti-droplet Control (ADC), for correctly pipettingvolatile organic solvents Reproducibility,consistencyrepeated workflow testing. Traceability, automated barcode verification of sam ples, reagents, plates and tips, as well as dynamictracking of each aspiration and dispensation step usingour monitored air displacement (MAD) and total aspirationand dispense monitoring (TADM) technologies.ensuredthrough14

Application NoteDiagnosticsAutomated Sample Preparationfor Immunosuppressant AnalysisAuthors: Frank Hasenäcker 1, Richard Lukacin 1and Stefan Mauch 2Chromsystems Instruments & Chemicals GmbH, Am Haag 12, 82166 Gräfelfing, Germany2Hamilton Bonaduz AG, Via Crusch 8, 7402 Bonaduz, Switzerland1Therapeutic drug monitoring requires solutions that complywith IVD guidelines, enable a full audit trail and are easy to use.Chromsystems together with Hamilton has developedMassSTAR as a solution that meets these requirements. Complete CE-IVD certified workflow Easy to use Full audit trailFigure 1. Hamilton MassSTARIntroductionImmunosuppressive drugs are applied to prevent the body from rejecting an organ transplant or to treat autoimmune diseases.As each patient’s absorption and metabolism of the drugs varies, correct dosing is crucial to avoid toxic reactions but still keepthe drug at effective levels and therefore ensure the wellbeing of the patient.Therapeutic drug monitoring of the regularly prescribed immunosuppressive drugs, cyclosporin A, everolimus, sirolimusand tacrolimus, is common in clinical laboratories. The gold standard for analysis of those drugs is LC-MS/MS.One of the challenges laboratories have been facing is the sample preparation. Monitoring of the processing and a full audit trailare crucial to ensure quality and integrity of results. Current customized automation solutions focus on the processing partbut tend to neglect the monitoring and audit trail.System DescriptionMassSTAR is based on a Hamilton Microlab STARlet IVD with4 channels, CO-RE Gripper and centrifuge integration to theright of the system. The deck consists of carriers for samples,one carrier for reagents and one for calibrators and controls.In addition, there are two carriers for filter plates, collectionplates and pipetting tips as well as a Hamilton Heater Shakerused for incubation. The system has a capacity of up to 288samples per run including controls and calibrators.The application software is based on Hamilton’s Vector 4.3 IVD.The method has been optimized to enable best performance.A user friendly GUI makes the system easy to use and generatesoutput files ready to use for all common LC-MS/MS system.Figure 2. The deck layout of the MassSTAR15

CE-IVD validated optimized workflowKit DescriptionThe Chromsystems ONEMINUTE MassTox r

used in the 1980s-'90s (mainly gas chromatography- mass spectrometry or GC-MS). The new (soft) ionization techniques, the high performance of the system once coupled with front end liquid chromatography (LC) and a second mass analyzer (MS/MS), gave MS the conditions necessary to be a viable and attractive alternative for routine

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