Lateral flow assay for simultaneous detection of cellular- and humoral immune responses

Department of Molecular Cell Biology Leiden University Medical Center, Einthovenweg 20, 2333 ZC Leiden, The Netherlands.
Clinical Biochemistry (Impact Factor: 2.28). 07/2011; 44(14-15):1241-6. DOI: 10.1016/j.clinbiochem.2011.06.983
Source: PubMed


The development of a cytokine detection assay suitable for detection of multiple biomarkers for improved diagnosis of mycobacterial diseases.
A lateral flow (LF) assay to detect IL-10 was developed utilizing the up-converting phosphor (UCP) reporter-technology. The assay was evaluated using blood samples of leprosy patients. Multiplex applications were explored targeting: 1) IL-10 and IFN-γ in assay buffer; 2) IL-10 and anti-phenolic glycolipid (PGL-I) antibodies in serum from leprosy patients.
Detection of IL-10 below the targeted level of 100pg/mL in serum was shown. Comparison with ELISA showed a quantitative correlation with R(2) value of 0.92. Multiplexing of cytokines and simultaneous detection of cytokine and antibody was demonstrated.
The UCP-LF IL-10 assay is a user-friendly, rapid alternative for IL-10 ELISAs, suitable for multiplex detection of different cytokines and can be merged with antibody-detection assays to simultaneously detect cellular- and humoral immunity.

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Available from: Paul Corstjens,
    • "Since then, UCP reporter particles have also been applied for immunochromatography (lateral flow assays) and, opportunities for development and implementation of UCP-based bioanalytical (POC) systems have increased with recent involvement of commercial suppliers of UCP materials and equipment [19]. This study describes the adaptation of a previously described UCP-LF assay-format [20] [21] to determine IP-10 and CCL4 levels in Mtb antigenstimulated whole-blood samples, following a multi-center evaluation by seven research institutes in six African countries. The assay-format applies sandwich-based immunochromatography with a complementary pair of chemokine-specific antibodies. "
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    ABSTRACT: Objective: Multi-center evaluation of a user-friendly lateral flow test for detection of IP-10 and CCL4 levels in Mycobacterium tuberculosis (Mtb) antigen-stimulated whole blood samples from tuberculosis (TB) suspects. Design and methods: A quantitative lateral flow (LF)-based assay platform was applied to detect chemokines IP-10 and CCL4. Chemokine quantitation was achieved using interference-free, fluorescent up-converting phosphor (UCP) labels. The new assays allowed worldwide shipping and storage without requiring a cold chain and were tested at seven institutes (including Ethiopia, Malawi, The Gambia, South Africa, Uganda and Namibia) employing portable lightweight readers for detection of the UCP label. At each site, clinical samples, confirmed TB and non-TB (i.e. other respiratory diseases (ORD)) cases, were collected and analyzed simultaneously with quality control (QC) human IP-10 or CCL4 standards. Results: Performance of the UCP-LF assay in Africa using QC standards indicated high robustness allowing quantitative detection between 100 and 100,000pg/mL. The optimized assays allowed successful determination of chemokine levels using 1μL whole blood sample from the locally recruited subjects with TB or ORD. Conclusion: This African multi-center trial further demonstrated the applicability of the low-tech and robust UCP-LF platform as a convenient quantitative assay for chemokine detection in whole blood. Ambient shipping and storage of all assay reagents and the availability of lightweight standalone readers were acknowledged as essential requirement for test implementation in particular in remote and resource-limited settings.
    Clinical biochemistry 09/2015; DOI:10.1016/j.clinbiochem.2015.08.013 · 2.28 Impact Factor
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    • "Each tube will be marked with a colored cap specific for one of these stimuli. After 24 hour incubation at 37°C, tubes will be frozen and stored for analysis of cellular markers [20] and/or analysis in field-friendly lateral flow assays for detection of Th1/Th2 cytokines as well as anti-PGL-I Ab [21]. "
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    ABSTRACT: Despite almost 30 years of effective chemotherapy with MDT, the global new case detection rate of leprosy has remained quite constant over the past years. New tools and methodologies are necessary to interrupt the transmission of M. leprae. Single-dose rifampicin (SDR) has been shown to prevent 57% of incident cases of leprosy in the first two years, when given to contacts of newly diagnosed cases. Immunization of contacts with BCG has been less well documented, but appears to have a preventive effect lasting up to 9 years. However, one major disadvantage is the occurrence of excess cases within the first year after immunization. The objective of this study is to examine the effect of chemoprophylaxis with SDR and immunoprophylaxis with BCG on the clinical outcome as well as on host immune responses and gene expression profiles in contacts of newly diagnosed leprosy patients. We hypothesize that the effects of both interventions may be complementary, causing the combined preventive outcome to be significant and long-lasting.Methods/design: Through a cluster randomized controlled trial we compare immunization with BCG alone with BCG plus SDR in contacts of new leprosy cases. Contact groups of around 15 persons will be established for each of the 1300 leprosy patients included in the trial, resulting in approximately 20,000 contacts in total. BCG will be administered to the intervention group followed by SDR, 2 months later. The control group will receive BCG only. In total 10,000 contacts will be included in both intervention arms over a 2-year period. Follow-up will take place one year as well as two years after intake. The primary outcome is the occurrence of clinical leprosy within two years. Simultaneously with vaccination and SDR, blood samples for in vitro analyses will be obtained from 300 contacts participating in the trial to determine the effect of these chemo- and immunoprophylactic interventions on immune and genetic host parameters. Combined chemoprophylaxis and immunoprophylaxis is potentially a very powerful and innovative tool aimed at contacts of leprosy patients that could reduce the transmission of M. leprae markedly. The trial intends to substantiate this potential preventive effect. Evaluation of immune and genetic biomarker profiles will allow identification of pathogenic versus (BCG-induced) protective host biomarkers and could lead to effective prophylactic interventions for leprosy using optimized tools for identification of individuals who are most at risk of developing disease.Trial registration: Netherlands Trial Register: NTR3087.
    BMC Infectious Diseases 10/2013; 13(1):456. DOI:10.1186/1471-2334-13-456 · 2.61 Impact Factor
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    • "The combination of the UCP reporter technology with the user-friendly and rapid LF-based immunochromatography has shown better sensitivity than ELISA-based assays, and several UCP-LF applications to detect different types of biomolecules have already been described (references recapped in Corstjens et al. 2011 [18]). In the current study, the focus is on IFX and IBD, but the results indicate that the assay in its current form is directly applicable to other biologicals (immunotherapeutics) and diseases as e.g. "
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    ABSTRACT: Monitoring levels of biologicals against tumor necrosis factor (TNF) has been suggested to improve therapeutic outcomes in inflammatory bowel diseases (IBDs). This pilot study describes a rapid lateral flow (LF)-based assay for on-site monitoring of serum trough levels of humanized monoclonal antibody infliximab (IFX). The applied chromatographic method utilizes sequential flows of diluted serum, wash buffer, and an immunoglobulin generic label on LF strips with a Test line comprised of TNF-α. The successive flows permitted enrichment of IFX at the Test line before the label was applied. The label, luminescent upconverting phosphor (UCP) particles coated with protein-A, emits a 550-nm visible light upon excitation with 980-nm infrared light. IFX concentrations were determined through measurement of UCP fluorescence at the Test line. The assay was optimized to detect IFX levels as low as 0.17 μg/mL in serum. For patients with IBD, this limit is appropriate to detect levels associated with loss of response (0.5 μg IFX/mL). The assay was evaluated with clinical samples from patients with Crohn’s disease and correlated well within the physiologically relevant range from 0.17 to 10 μg/mL with an IFX-specific ELISA. Performance of the assay was further successfully validated with samples from blood donors, IFX negative IBD patients, and rheumatoid arthritis patients that had developed anti-IFX antibodies. Because of its generic nature, the assay is suited for detecting most therapeutic anti-TNF-α monoclonal antibodies. Figure A rapid lateral flow based assay to determine trough levels of infliximab and other anti‐TNF‐α antibodies. The rapid format showed excellent and quantitative correlation with ELISA. Accurate quantitation was achieved utilizing the up‐converting phosphor reporter technology and a portable lightweight ESEQuant LFR reader adapted with an infrared LED
    Analytical and Bioanalytical Chemistry 07/2013; DOI:10.1007/s00216-013-7154-0 · 3.44 Impact Factor
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