Distinguishing septic from normal donors by detection of sPLA2-IIA from human plasma using a microsieve-based immunoassay
Autor: | Daniel R. Zweitzig, Leon W.M.M. Terstappen, Mark J. Kopnitsky, Kathleen Cichonski, Arjan G.J. Tibbe |
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Přispěvatelé: | Medical Cell Biophysics |
Jazyk: | angličtina |
Rok vydání: | 2017 |
Předmět: |
0301 basic medicine
Male medicine.drug_class Immunology Spla2 iia Enzyme-Linked Immunosorbent Assay Monoclonal antibody law.invention Sepsis 03 medical and health sciences 0302 clinical medicine law medicine Immunology and Allergy Humans Phospholipases A2 Secretory Detection limit Immunoassay Chromatography medicine.diagnostic_test Septic shock Chemistry Antibodies Monoclonal 030208 emergency & critical care medicine Clinical Enzyme Tests medicine.disease Immunity Innate n/a OA procedure 030104 developmental biology Human plasma Recombinant DNA Feasibility Studies |
Zdroj: | Journal of immunological methods, 447, 86-91. Elsevier |
ISSN: | 0022-1759 |
Popis: | Bloodstream infections that progress to septic shock are responsible for hundreds of thousands of deaths each year, and are associated with significant healthcare costs. Recent studies have shown that a member of the secreted phospholipase protein family, termed sPLA2-IIA, may play a role during the innate immune response to bacterial infections, and is elevated in the plasma of septic patients. In this report, the feasibility of a simple microsieve-based sPLA2-IIA detection immunoassay was explored. Microsieves containing 5 μm pores were covalently coupled with a sPLA2-IIA-specific monoclonal antibody at 0.1, 1.0, and 10 μg/mL and then assayed with plasma-based positive and negative controls to determine the optimal coating concentration. Recombinant sPLA2-IIA was then serially diluted to a final concentration of 200, 100, 50, 25, 12.5, and 6.25 ng/mL and tested alongside a non-spiked sample to estimate the detection limit of the prototype assay. Recombinant sPLA2-IIA was also spiked into serum, EDTA-plasma, and Lithium-Heparin plasma, in an effort to evaluate assay performance when analyzing these sample matrices. The preliminary limit of detection studies suggests that the microsieve assay is able to distinguish approximately 6–12 ng/mL of sPLA2-IIA from a non-spiked sample. When compared to an immunoassay diluent, the microsieve assay also yielded acceptable percent recoveries for each of the three sample matrices spiked with clinically significant levels of sPLA2-IIA. The sPLA2-IIA microsieve assay prototype also clearly distinguished five samples from septic patients from five normal donor samples, and the results were in good agreement with a comparator ELISA test system (R2 = 0.9347). |
Databáze: | OpenAIRE |
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