Measurement Of Multiple Analytes Research Articles

Evaluation and establishment of reference intervals using the i-STAT1 blood chemistry analyzer in turkeys

In veterinary medicine, point-of-care testing techniques have become popular, since they provide immediate results and only small amounts of blood are needed. The handheld i-STAT1 blood analyzer is used by poultry researchers and veterinarians; however, no studies have evaluated the accuracy of this analyzer determined reference intervals in turkey blood. The objectives of this study were to 1) investigate the effect of storage time on turkey blood analytes, 2) compare the results obtained by the i-STAT1 analyzer to those obtained by the GEM Premier 3000, a conventional laboratory analyzer, and 3) establish reference intervals for blood gases and chemistry analytes in growing turkeys using the i-Stat. For the first and second objectives, we used the CG8+ i-STAT1 cartridges to test blood from 30 healthy turkeys in triplicate and once with the conventional analyzer. To establish the reference intervals, we tested a total 330 blood samples from healthy turkeys from 6 independent flocks during a 3-yr period. Blood samples were then divided into brooder (<1 wk) and growing (1-12 wk of age). Friedman's test demonstrated significant time-dependent changes in blood gas analytes, but not for electrolytes. Bland-Altman analysis revealed that there was agreement between the i-STAT1 and the GEM Premier 300 for most of the analytes. However, Passing-Bablok regression analysis identified constant and proportional biases in the measurement of multiple analytes. Tukey's test revealed significant differences in the whole blood analytes between the means of brooding and growing birds. The data presented in the present study provide a basis for measuring and interpreting blood analytes in the brooding and growing stages of the turkey lifecycle, offering a new approach to health monitoring in growing turkeys.

Potential Biomarkers for Asymptomatic Visceral Leishmaniasis among Iraq-Deployed U.S. Military Personnel.

Visceral leishmaniasis (VL) is a chronic infection caused by Leishmania (L.) donovani or L. infantum parasites. Despite having the infection, most individuals never develop the clinical disease and are able to control the parasite and remain asymptomatic. However, some progress to symptomatic VL, leading to death if untreated. The host immune response has a major role in determining the progression and severity of the clinical manifestations in VL; several immune biomarkers of symptomatic VL have been described with interferon-gamma release as a surrogate biomarker of host cellular immunity. However, new biomarkers to identify asymptomatic VL (AVL) are needed for the identification of people at risk for VL activation. In our study, levels of chemokine/cytokine in the supernatants of peripheral mononuclear blood cells (PBMC) from 35 AVL+ Iraq-deployed participants, stimulated in vitro with soluble Leishmania antigen for 72 h, were assessed by a bead-based assay that allows the measurement of multiple analytes. PBMC of AVL-negative military beneficiaries were used as controls. Monocyte Chemoattractant Protein-1, Monokine Induced by Gamma Interferon and Interleukin-8, were detected at high levels in AVL+ stimulated cultures from Iraq deployers compared to uninfected controls. Measurement of chemokine/cytokine levels can identify cellular immune responses in AVL+ asymptomatic individuals.

Designing electrochemical microfluidic multiplexed biosensors for on-site applications

Clinical assessment based on a single biomarker is in many circumstances not sufficient for adequate diagnosis of a disease or for monitoring its therapy. Multiplexing, the measurement of multiple analytes from one sample and/or of the same target from different samples simultaneously, could enhance the accuracy of the diagnosis of diseases and their therapy success. Thus, there is a great and urgent demand for multiplexed biosensors allowing a low-cost, easy-to-use, and rapid on-site testing. In this work, we present a simple, flexible, and highly scalable strategy for implementing microfluidic multiplexed electrochemical biosensors (BiosensorX). Our technology is able to detect 4, 6, or 8 (different) analytes or samples simultaneously using a sequential design concept: multiple immobilization areas, where the assay components are adsorbed, followed by their individual electrochemical cells, where the amperometric signal readout takes place, within a single microfluidic channel. Here, first we compare vertical and horizontal designs of BiosensorX chips using a model assay. Owing to its easier handling and superior fluidic behavior, the vertical format is chosen as the final multiplexed chip design. Consequently, the feasibility of the BiosensorX for multiplexed on-site testing is successfully demonstrated by measuring meropenem antibiotics via an antibody-free β-lactam assay. The multiplexed biosensor platform introduced can be further extended for the simultaneous detection of other anti-infective agents and/or biomarkers (such as renal or inflammation biomarkers) as well as different (invasive and non-invasive) sample types, which would be a major step towards sepsis management and beyond.Graphical

Design of a methotrexate-controlled chemical dimerization system and its use in bio-electronic devices

Natural evolution produced polypeptides that selectively recognize chemical entities and their polymers, ranging from ions to proteins and nucleic acids. Such selective interactions serve as entry points to biological signaling and metabolic pathways. The ability to engineer artificial versions of such entry points is a key goal of synthetic biology, bioengineering and bioelectronics. We set out to map the optimal strategy for developing artificial small molecule:protein complexes that function as chemically induced dimerization (CID) systems. Using several starting points, we evolved CID systems controlled by a therapeutic drug methotrexate. Biophysical and structural analysis of methotrexate-controlled CID system reveals the critical role played by drug-induced conformational change in ligand-controlled protein complex assembly. We demonstrate utility of the developed CID by constructing electrochemical biosensors of methotrexate that enable quantification of methotrexate in human serum. Furthermore, using the methotrexate and functionally related biosensor of rapamycin we developed a multiplexed bioelectronic system that can perform repeated measurements of multiple analytes. The presented results open the door for construction of genetically encoded signaling systems for use in bioelectronics and diagnostics, as well as metabolic and signaling network engineering.

Adaptation of ELISA detection of Plasmodium falciparum and Plasmodium vivax circumsporozoite proteins in mosquitoes to a multiplex bead-based immunoassay

BackgroundPlasmodium spp. sporozoite rates in mosquitoes are used to better understand malaria transmission intensity, the relative importance of vector species and the impact of interventions. These rates are typically estimated using an enzyme-linked immunosorbent assay (ELISA) utilizing antibodies against the circumsporozoite protein of Plasmodium falciparum, Plasmodium vivax VK210 (P. vivax210) or P. vivax VK247 (P. vivax247), employing assays that were developed over three decades ago. The ELISA method requires a separate assay plate for each analyte tested and can be time consuming as well as requiring sample volumes not always available. The bead-based multiplex platform allows simultaneous measurement of multiple analytes and may improve the lower limit of detection for sporozoites.MethodsRecombinant positive controls for P. falciparum, P. vivax210 and P. vivax247 and previously developed circumsporozoite (cs) ELISA antibodies were used to optimize conditions for the circumsporozoite multiplex bead assay (csMBA) and to determine the detection range of the csMBA. After optimizing assay conditions, known amounts of sporozoites were used to determine the lower limit of detection for the csELISA and csMBA and alternate cut-off measures were applied to demonstrate how cut-off criteria can impact lower limits of detection. Sporozoite rates from 1275 mosquitoes collected in Madagascar and 255 mosquitoes collected in Guinea were estimated and compared using the established csELISA and newly optimized csMBA. All mosquitoes were tested (initial test), and those that were positive were retested (retest). When sufficient sample volume remained, an aliquot of homogenate was boiled and retested (boiled retest), to denature any heat-unstable cross-reactive proteins.ResultsFollowing optimization of the csMBA, the lower limit of detection was 25 sporozoites per mosquito equivalent for P. falciparum, P. vivax210 and P. vivax247 whereas the lower limits of detection for csELISA were found to be 1400 sporozoites for P. falciparum, 425 for P. vivax210 and 1650 for P. vivax247. Combined sporozoite rates after re-testing of samples that initially tested positive for Madagascar mosquitoes by csELISA and csMBA were 1.4 and 10.3%, respectively, and for Guinea mosquitoes 2% by both assays. Boiling of samples followed by csMBA resulted in a decrease in the Madagascar sporozoite rate to 2.8–4.4% while the Guinea csMBA sporozoite rate remained at 2.0%. Using an alternative csMBA cut-off value of median fluorescence intensity (MFI) of 100 yielded a sporozoite rate after confirmational testing of 3.7% for Madagascar samples and 2.0% for Guinea samples. Whether using csMBA or csELISA, the following steps may help minimize false positives: specimens are appropriately stored and bisected anterior to the thorax-abdomen junction, aliquots of homogenate are boiled and retested following initial testing, and an appropriate cut-off value is determined.ConclusionsThe csMBA is a cost-comparable and time saving alternative to the csELISA and may help eliminate false negatives due to a lower limit of detection, thus increasing sensitivity over the csELISA. The csMBA expands the potential analyses that can be done with a small volume of sample by allowing multiplex testing where analytes in addition to P. falciparum, P. vivax210 and P. vivax247 can be added following optimization.

Comparison of portable and conventional laboratory analyzers for biochemical tests in chickens

Antemortem blood biochemical and blood gas analyses are routinely used in health screening and diagnosis of disease in domestic veterinary species. These testing modalities are not routinely performed in poultry, in part, due to the distance from the diagnostic laboratory. Portable blood analyzers such as the i-STAT and VetScan (VS2) can be used to obtain results on the farm without delay, potentially offering a more practical option for poultry practitioners. We investigated the time effect on blood chemistry values and compared the results obtained using the i-STAT and VS2 with those obtained using conventional laboratory analyzers (GEM Premier 3000 and Cobas c501, respectively). We tested blood from 60 healthy chickens. Each sample was tested in triplicate using each of the portable analyzers and once using conventional analyzers. All samples were analyzed within 60 minutes of collection. The concentrations of some analytes were outside the limit of detection of the portable analyzers (i.e., bile acids). Although statistically significant differences were found for some biochemical analytes over time, the actual mean or median differences were too small to be considered of clinical importance. As observed in mammals, significant time-dependent changes in blood gas analytes were observed in whole blood samples exposed to ambient air. Correlation coefficients between portable and conventional analyzers were moderate to high for most of the analytes. For the most part, there was an agreement between the portable and conventional analyzers. We identified constant and proportional biases in the measurement of multiple analytes by both the i-STAT and VS2. Future studies are warranted to establish analyzer-specific reference intervals for poultry.

Multiplexed Detection of Analytes on Single Test Strips with Antibody-Gated Indicator-Releasing Mesoporous Nanoparticles.

Rapid testing methods for the use directly at a point of need are expected to unfold their true potential especially when offering adequate capabilities for the simultaneous measurement of multiple analytes of interest. Considering the unique modularity, high sensitivity, and selectivity of antibody‐gated indicator delivery (gAID) systems, a multiplexed assay for three small‐molecule explosives (TATP, TNT, PETN) was thus developed, allowing to detect the analytes simultaneously with a single test strip at lower ppb concentrations in the liquid phase in <5 min using a fluorescence reader or a smartphone for readout. While the TNT and PETN systems were newly developed here, all the three systems also tolerated harsher matrices than buffered aqueous model solutions. Besides a single‐track strip, the outstanding modularity of the hybrid biosensor materials in combination with strip‐patterning technologies allowed us to obtain a multichannel strip in a straightforward manner, offering comparable analytical performance while allowing to be tailored even more to the user's need.

Fast and Efficient Measurement of Clinical and Biological Samples Using Immunoassay-Based Multiplexing Systems.

Immunoassay is one of the most commonly used biomedical techniques to detect the expression of an antibody or an antigen in a test sample. Enzyme-linked immunosorbent assay (ELISA) has been used for a variety of applications including diagnostic tools and quality controls. However, one of the main limitations of ELISA is its lack of multiplexing ability, so ELISA may not be an efficient diagnostic tool when a measurement of multiple determinants is needed for samples with limited quantity such as blood or biological samples from newborns or babies. Although similar to ELISA in assay measurement, multiplex platforms such as bead-based Luminex and multi-array-based MSD (Meso Scale Discovery) are widely used to measure multiple biomarkers from a single analysis. Luminex is a xMAP-based technology that combines several different technologies to provide an efficient and accurate measurement of multiple analytes from a single sample. The multiplexing can be achieved because up to 100 distinct Luminex color-coded microsphere bead sets can be coated with a reagent specific to a particular bioassay, allowing the capture and detection of specific analytes from a sample. Using Multi-array and electrochemiluminescence technologies, the MSD platform provides the multiplex capability with similar consistence as observed in ELISA. Various biological samples that can be analyzed by both Luminex and MSD systems include serum, plasma, tissue and cell lysate, saliva, sputum, and bronchoalveolar Lavage (BAL). The most common Luminex and MSD-based assays are to detect a combined set of cytokines to provide a measurement of cytokine expression profiling for a diagnostic purpose.

Enhanced DGT capability for measurements of multiple types of analytes using synergistic effects among different binding agents

There is a requirement for simultaneous measurements of diverse analytes to reveal their biogeochemical couplings in the environment. In this study, we first realized simultaneous measurements of three types of analytes, including sulfide [S(-II)], metallic cations, and oxyanions by diffusive gradients in thin films (DGT) using a new mixed ZrO-CA binding gel. The ZrO-CA gel exhibited faster absorption rates for oxyanions than previously reported ZrO-Chelex gel, and faster absorption rates for cations when the gel was saturated with S(-II). It implies that there were synergistic effects among the three binding agents which should facilitate the DGT uptake. The technique's performance was validated under laboratory conditions in mixtures of dissolved compounds. It was shown to be independent of the solution's pH (5–9) and ionic strength (from 1 to 3 mM to 750 mM). The capacities of the ZrO-CA DGTs without or with [S(-II)] saturation to measure As(III), As(V), Cd(II), Fe(II), and P(V) individually were significantly greater than those of other DGTs except for Zr-oxide based DGTs. The measurements in sediment revealed similar distributions of three types of analytes in the vertical profile and confirmed the feasibility and advantage of the ZrO-CA DGT. This study provides a new perspective to enhance the DGT capability in measurement of multiple analytes through utilizing the synergistic effects among different binding agents.

Opportunities and Challenges of Multiplex Assays: A Machine Learning Perspective.

Multiplex assays that allow the simultaneous measurement of multiple analytes in small sample quantities have developed into a widely used technology. Their implementation spans across multiple assay systems and can provide readouts of similar quality as the respective single-plex measures, albeit at far higher throughput. Multiplex assay systems are therefore an important element for biomarker discovery and development strategies but analysis of the derived data can face substantial challenges that may limit the possibility of identifying meaningful biological markers. This chapter gives an overview of opportunities and challenges of multiplexed biomarker analysis, in particular from the perspective of machine learning aimed at identification of predictive biological signatures.

Development and validation of the simultaneous measurement of four vitamin D metabolites in serum by LC-MS/MS for clinical laboratory applications.

The quantification of serum 25-hydroxyvitamin D [25(OH)D] as an indicator of vitaminD status is currently primarily conducted by immunoassays, yet LC-MS/MS would allow more accurate determination. Furthermore, LC-MS/MS would allow simultaneous measurement of multiple analytes. The aim of this study was to develop and validate an LC-MS/MS method to simultaneously measure four vitaminD metabolites (25(OH)D3, 3-epi-25(OH)D3, 25(OH)D2, and 24,25(OH)2D3) in serum for clinical laboratory applications. Serum samples were first prepared in a 96-well supported liquid extraction plate and the eluate was derivatized using the Cookson-type reagent 4-(4'-dimethylaminophenyl)-1,2,4-triazoline-3,5-dione (DAPTAD), which rapidly and quantitatively reacts with the s-cis-diene structure of vitaminD metabolites. The derivatized samples were subjected to LC-MS/MS, ionized by electrospray ionization (positive-ion mode), and detected by selected reaction monitoring. The lower limits of quantification for 25(OH)D3, 3-epi-25(OH)D3, 25(OH)D2, and 24,25(OH)2D3 were 0.091, 0.020, 0.013, and 0.024ng/mL, respectively. The accuracy values and the extraction recoveries for these four metabolites were satisfactory. Serum 25(OH)D levels determined by our LC-MS/MS were compared with those obtained by conventional radioimmunoassay (RIA) that cannot distinguish 25(OH)D3 and 25(OH)D2. The values obtained by the RIA method exhibited a mean bias of about 8.35ng/mL, most likely as a result of cross reaction of the antibody with low-abundance metabolites, including 24,25(OH)2D3. Various preanalytical factors, such as long sample sitting prior to serum separation, repeated freeze-thaw cycles, and the presence of anticoagulants, had no significant effects on these determinations. This high-throughput LC-MS/MS simultaneous assay of the four vitaminD metabolites 25(OH)D3, 3-epi-25(OH)D3, 25(OH)D2, and 24,25(OH)2D3 required as little as 20μL serum. This method will aid further understanding of low-abundance vitaminD metabolites, as well as the accurate determination of 25(OH)D3 and 25(OH)D2.

Chromogranin A

The review summarizes the utility and limitations of chromogranin A (CgA) as a circulating biomarker for neuroendocrine tumors (NETs). Blood CgA measurement has numerous clinical limitations including poor assay reproducibility, low sensitivity (meta-analysis: 73%, 95% confidence interval: 0.71-0.76), and a paucity of prospective validation studies. A recent study noted elevation in 27% of NETs with a predictive value of 50% for metastases. These findings are consistent with its efficacy primarily as a monoanalyte secretory rather than multidimensional neoplastic marker. An automated CgA assay (KRYPTOR) exhibits similar metrics to the DAKO assay but is only useful in serum and routine storage diminishes its accuracy. Current studies indicate that CgA is more effective as a biomarker for cardiac disease. Given the diverse limitations of CgA, NET biomarker focus has evolved toward measurement of multiple analytes, for example, transcripts. Multianalyte algorithmic analyses perform significantly better as diagnostic (>95%) and prognostic markers (>90%) than CgA (30-74 and ∼50%, respectively) since they delineate different aspects of the biological behavior of NETs, (e.g., proliferome and metabolome). CgA is neither a reliable nor robust NET biomarker. As a monoanalyte, it is restricted by poor metrics and has limited predictive value. Its current clinical utility appears optimal in cardiovascular disease. The significance of CgA in NET disease is diminishing as other analytical approaches, particularly transcript multianalyte assays or other strategies, evolve to supersede it.

Abstract B01: Profiling of serum and plasma angiogenic biomarkers in breast cancer and colon cancer using Luminex xMAP technology-based human angiogenesis multiplex immunoassay panels

Abstract Tumors require neovascularization for growth. Consequently, the angiogenesis process is key to understanding tumor development and progression, as well as diagnosis and treatment. Measuring the levels of pro- and anti-angiogenic factors in plasma or serum in cancer patients may provide prognostic information concerning disease processes and mechanisms and the elucidation of potential biomarkers for epidemiological studies or clinical trials. The Luminex xMAP technology, a bead-based multiplex immunoassay for the simultaneous measurement of multiple analytes in a small sample size, has been widely used in cancer biomarker discoveries. We report here the use of two newly developed multiplex human angiogenesis panels to profile the circulating biomarkers commonly used in the angiogenesis pathway analysis. The two panels are 1) MILLIPLEX MAP Human Angiogenesis Panel 1, a 17-plex multiplex immunoassay for the simultaneous quantification of the following 17 angiogenesis biomarkers: Angiopoietin-2, BMP-9, EGF, Endoglin, Endothelin-1, FGF-1, FGF-2, Follistatin, G-CSF, HB-EGF, HGF, IL-8, Leptin, PLGF, VEGF-A, VEGF-C, and VEGF-D, and 2) MILLIPLX MAP Human Angiogenesis Panel 2, a 20-plex multiplex immunoassay for simultaneous quantitation of the following 20 human angiogenesis biomarkers: Angiostatin, soluble E-Selectin, Osteopontin, PDGF-AB/BB, PECAM-1/sCD31, Tenascin C, Thrombospondin-2 (TSP-2), soluble AXL, soluble c-Kit, soluble EGFR, soluble Her2/ErbB2, soluble Her3/ErbB3, soluble HGFR/c-Met, soluble IL-6Rα, soluble Neuropilin-1, soluble Tie-2, soluble urokinase receptor (suPAR), soluble VEGFR1, soluble VEGFR2, and soluble VEGFR3. In this study, we report the use of these 37 angiogenic, growth factor and soluble receptor biomarkers to profile serum samples and plasma samples from a panel of breast cancer and colon cancer patients, with a similar number of normal serum and plasma samples, as a corresponding set, to generate a tumor profile of angiogenic factors using the Luminex xMAP platform. The results of this study demonstrated the utility of these human angiogenesis panels in studying cancer development and cancer biomarker discovery, as well as its potential application in translational research. Citation Format: Wen-Rong Lie, Deborah Droll, Jehangir Mistry. Profiling of serum and plasma angiogenic biomarkers in breast cancer and colon cancer using Luminex xMAP technology-based human angiogenesis multiplex immunoassay panels. [abstract]. In: Proceedings of the AACR Special Conference: Tumor Angiogenesis and Vascular Normalization: Bench to Bedside to Biomarkers; Mar 5-8, 2015; Orlando, FL. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl):Abstract nr B01.

Forecasting Pregnancy Outcomes in Women with Systemic Lupus Erythematosus

Most data about determinants of pregnancy outcomes in women with lupus come from retrospective studies. Now, in this prospective study, 385 pregnant women with lupus were followed with measurement of multiple analytes throughout pregnancy to determine risk factors for adverse outcomes (fetal or neonatal death; birth before 36 weeks' gestation because of placental insufficiency, …

Out-of-plane valve for blood separation and metering: towards lab-in-a-tube applications for blood diagnostics

Diagnostics plays a critical role in human health, while blood analysis is one of the most common clinical diagnoses. Biomedical engineers have been developing technologies that can robotically perform accurate and rapid blood analysis for field application. The simplified fabrication processes and low cost materials become the tendency of point-of-care (POC) devices for blood analysis. Recently, lab-in-a-tube technique comes up as a simple and robust method for POC measurements of multiple analytes. This paper proposed a tubing system with out-of-plane valves using tygon tubing assembled on a circular disc (CD) for blood separation. Pure plasma could be achieved within minutes after extracting blood from human body. The out-of-plane valve showed excellent capability in preventing backflow and reverse mixing due to diffusion. Therefore, the lab-in-a-tube platform has promising applications for the automation of POC analysis, especially important for the rural areas without enough medical resources.

Analytical Performance Comparisons of Multiplex Technologies and Single ELISAs on Seven Serum-based Immunoassays

Throughput and specimen volume requirement limitations to singleplex enzyme-linked immunosorbent assay (ELISA) technologies have inspired development of multiplexed technologies for biomarker discovery. Yet, simultaneous measurement of multiple analytes has been suggested to have its own tradeoffs, most notably an increased propensity for antibody cross-reactivity leading to compromised sensitivity and precision. We compare the analytic performance of 7 serum-based ELISAs (R&D Systems, Minneapolis, …

Potential of a simple lab-on-a-tube for point-of-care measurements of multiple analytes

This technical note presents a simple and disposable lab-on-a-tube (LOT) for point-of-care measurements of multiple analytes. LOT is a one-step device that can perform both sample collection and multi-sensing on-site. Sample collection is conducted by taking advantage of its inherent micro/macro channel structure while multi-sensing is conducted by integrated microsensors. This approach ensures reliable transportation of various samples into the testing area by either passive capillary force or active suction force, thus avoiding the need for a pump or injection components as used in lab-on-a-chip systems. The developed LOT (Diameter = 1 mm, Sensing length = 4.5 mm, Required sample volume = 3.5 microl) is capable of simultaneously quantifying the concentrations of glucose, lactate and oxygen in human serum samples. The result suggests the LOT hold great potential for many point-of-care applications.

Identification and Quantitation of Amphetamines, Cocaine, Opiates, and Phencyclidine in Oral Fluid by Liquid Chromatography—Tandem Mass Spectrometry

Analytical methods for measuring multiple licit and illicit drugs and metabolites in oral fluid require high sensitivity, specificity, and accuracy. With the limited volume available for testing, comprehensive methodology is needed for simultaneous measurement of multiple analytes in a single aliquot. This report describes the validation of a semi-automated method for the simultaneous extraction, identification, and quantitation of 21 analytes in a single oral fluid aliquot. The target compounds included are amphetamine, methamphetamine, 3,4-methylenedioxymethamphetamine, 3,4-methylenedioxy-amphetamine, 3,4-methylenedioxyethylamphetamine, pseudoephedrine, cocaine, benzoylecgonine, codeine, norcodeine, 6-acetylcodeine, morphine, 6-acetylmorphine, hydrocodone, norhydrocodone, dihydrocodeine, hydromorphone, oxycodone, noroxycodone, oxymorphone, and phencyclidine. Oral fluid specimens were collected with the Intercept device and extracted by solid-phase extraction (SPE). Drug recovery from the Intercept device averaged 84.3%, and SPE extraction efficiency averaged 91.2% for the 21 analytes. Drug analysis was performed by liquid chromatography-tandem mass spectrometry in the positive electrospray mode using ratios of qualifying product ions within +/-25% of calibration standards. Matrix ion suppression ranged from -57 to 8%. The limit of quantitation ranged from 0.4 to 5 ng/mL using 0.2 mL of diluted oral fluid sample. Application of the method was demonstrated by testing oral fluid specimens from drug abuse treatment patients. Thirty-nine patients tested positive for various combinations of licit and illicit drugs and metabolites. In conclusion, this validated method is suitable for simultaneous measurement of 21 licit and illicit drugs and metabolites in oral fluid.

The new method for simultaneous measurement of multiple analytes: Multiplex test system and method performances in systemic autoimmune disease diagnosis

The new method for simultaneous measurement of multiple analytes: Multiplex test system and method performances in systemic autoimmune disease diagnosis

Validation and comparison of two multiplex technologies, Luminex® and Mesoscale Discovery, for human cytokine profiling

Biomarker research has rapidly expanded over recent years aided by the progressive development of research tools, in particular the different multiplex technologies allowing simultaneous measurement of multiple analytes. It is foreseeable that such technology will have an integral role in clinical studies for establishing biomarker profiles of disease status, but validation of the tools is essential to confirm the reliability of their application. More comparable studies between multiplex platforms are required to enable users to determine which of these are best for a particular clinical study, as different platforms will have varying levels of performance for the validation parameters. Comparison of two multiplex platforms, the Luminex® and the Mesoscale Discovery, has been performed to determine their performance for the validation parameters of sensitivity, precision and accuracy for the cytokines IL-2, IL-4, IL-8, IL-10, IL-12, IFNγ and TNFα. When measuring high concentrations both platforms show good accuracy (within +/−25% recovery) with all cytokines except IL-12 for the MSD. At low concentrations, +/−25% recovery was seen with all cytokines except IL-2 and IL-8 for the Luminex and IL-2 and IL-12 for the MSD. Although quantitative differences are found, relative differences are comparable, and consequently both platforms have been shown to be suitable for analyzing trends in multiple cytokine profiles, with the Luminex having better precision and the Mesoscale Discovery having greater sensitivity.