Development of an immunochromatographic lateral-flow device for rapid serodiagnosis of invasive aspergillosis.

Hybridoma Laboratory, School of Biosciences, Geoffrey Pope Building, University of Exeter, Stocker Road, Exeter, Devon EX4 4QD, United Kingdom.
Clinical and vaccine Immunology: CVI (Impact Factor: 2.37). 07/2008; 15(7):1095-105. DOI: 10.1128/CVI.00068-08
Source: PubMed

ABSTRACT Aspergillus fumigatus is a cosmopolitan saprotrophic fungus that is second only to Candida species as a cause of invasive fungal infections in immunocompromised humans. Current immunodiagnostic tests for invasive aspergillosis (IA) are based on the detection of circulating galactomannan (GM) in a patient's serum by using a rat monoclonal antibody (MAb), EB-A2, that binds to tetra (1-->5)-beta-D-galactofuranoside, the immunodominant epitope in GM. The potential cross-reactivity of MAb EB-A2 with non-Aspergillus fungi, with contaminating GM in beta-lactam antibiotics and foodstuffs, and with bacterial lipoteichoic acids has prompted efforts to discover non-GM antigens that can act as surrogate markers for the diagnosis of IA. This paper describes the development of a mouse MAb, JF5, that binds to a protein epitope present on an extracellular glycoprotein antigen secreted constitutively during the active growth of A. fumigatus. The MAb was used to develop an immunochromatographic lateral-flow device (LFD) for the rapid (15-min) detection of Aspergillus antigens in human serum. The test is highly specific, reacting with antigens from Aspergillus species but not with antigens from a large number of clinically important fungi, including Candida species, Cryptococcus neoformans, Fusarium solani, Penicillium marneffei, Pseudallescheria boydii, and Rhizopus oryzae. The LFD was able to detect circulating antigen in serum samples from patients suspected of having or shown to have IA on the basis of their clinical symptoms and results from tests for GM and fungal (1-->3)-beta-D-glucan. The ease of use of the LFD provides a diagnostic platform for the routine testing of vulnerable patients who have an elevated risk of IA.

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    ABSTRACT: Objectives Invasive pulmonary aspergillosis has been increasingly reported in non-neutropenic patients including those with underlying respiratory diseases. We compared the diagnostic performances of Galactomannan, 1,3-beta-D-glucan, and Aspergillus-specific lateral-flow device tests with that of conventional culture by using bronchoalveolar lavage fluid samples from patients with underlying respiratory diseases. Methods We analyzed 268 bronchoalveolar lavage samples from 221 patients with underlying respiratory diseases (and without hematologic malignancy or previous solid organ transplantation) that were collected for routine microbiological workup between February 2012 and May 2014 at the University Hospital of Graz, Austria. Invasive pulmonary aspergillosis was defined according to EORTC/MSG criteria modified for patients with respiratory diseases. Results 31 patients (14%) had probable or proven, 25 possible and the remaining 165 patients no invasive pulmonary aspergillosis. Probable/proven aspergillosis was associated with a significantly higher (p=0.034) 30-day mortality rate of 32%. Sensitivities, specificities and diagnostic odd ratios differed markedly between galactomannan (cut-off 0.5 optical-density-index: 0.97, 0.81, 124.4; cut-off 1.0: 0.97, 0.93, 422.1; cut-off 3.0: 0.61, 0.99, 109.8), beta-d-glucan (cut-off 80pg/ml: 0.90, 0.42, 6.57; cut-off 200pg/ml: 0.70, 0.61, 3.7), lateral-flow-device tests (0.77, 0.92, 41.8) and mycological culture (0.29, 0.97, 14). Conclusion Probable or proven invasive pulmonary aspergillosis was diagnosed in 14% of our study population and associated with significantly higher 30-day mortality rates. While the performance of beta-d-glucan was limited by low specificity, and that of mycological culture by low sensitivity, the Aspergillus lateral-flow device seems to be a promising alternative to galactomannan testing that remains the diagnostic gold standard for aspergillosis. Clinical trial registration available at, ID NCT02058316.
    American Journal of Respiratory and Critical Care Medicine 09/2014; 190(8). DOI:10.1164/rccm.201407-1275OC
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    ABSTRACT: Rapid and timely diagnosis of infectious diseases is a critical determinant of clinical outcomes and general public health. For the detection of various pathogens, microfluidics-based platforms offer many advantages, including speed, cost, portability, high throughput, and automation. This review provides an overview of the recent advances in microfluidic technologies for point-of-care (POC) diagnostics for infectious diseases. The key aspects of such technologies for the development of a fully integrated POC platform are introduced, including sample preparation, on-chip nucleic acid analysis and immunoassay, and system integration/automation. The current challenges to practical implementation of this technology are discussed together with future perspectives. Copyright © 2014. Published by Elsevier B.V.
    Journal of Chromatography A 12/2014; 1377. DOI:10.1016/j.chroma.2014.12.041
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    ABSTRACT: Invasive fungal infections constitute a serious threat to an ever-growing population of immunocompromised individuals and other individuals at risk. Traditional diagnostic methods, such as histopathology and culture, which are still considered the gold standards, have low sensitivity, which underscores the need for the development of new means of detecting fungal infectious agents. Indeed, novel serologic and molecular techniques have been developed and are currently under clinical evaluation. Tests like the galactomannan antigen test for aspergillosis and the β-glucan test for invasive Candida spp. and molds, as well as other antigen and antibody tests, for Cryptococcus spp., Pneumocystis spp., and dimorphic fungi, have already been established as important diagnostic approaches and are implemented in routine clinical practice. On the other hand, PCR and other molecular approaches, such as matrix-assisted laser desorption ionization (MALDI) and fluorescence in situ hybridization (FISH), have proved promising in clinical trials but still need to undergo standardization before their clinical use can become widespread. The purpose of this review is to highlight the different diagnostic approaches that are currently utilized or under development for invasive fungal infections and to identify their performance characteristics and the challenges associated with their use.
    Clinical Microbiology Reviews 07/2014; 27(3):490-526. DOI:10.1128/CMR.00091-13


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