A rapid and simple chemiluminescence method for screening levels of inosine and hypoxanthine in non‐traumatic chest pain patients |
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| Authors: | Don E Farthing Domenic Sica Michael Hindle Les Edinboro Lei Xi Todd WB Gehr Lynne Gehr Christine A Farthing Terri L Larus Itaf Fakhry H Thomas Karnes |
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| Institution: | 1. Department of Internal Medicine, Divisions of Nephrology, Clinical Pharmacology and Hypertension, Virginia Commonwealth University, Richmond, VA, USA;2. Department of Pharmaceutics, Virginia Commonwealth University, Richmond, VA, USA;3. Department of Forensic Science, State of Virginia, Richmond, VA, USA;4. Department of Internal Medicine, Division of Cardiology, Virginia Commonwealth University, Richmond, VA, USA;5. Department of Anesthesiology, Virginia Commonwealth University, Richmond, VA, USADepartment of Anesthesiology, Virginia Commonwealth University, Richmond, VA, USA |
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| Abstract: | A rapid and simple chemiluminescence method was developed for detection of inosine and hypoxanthine in human plasma. The method utilized a microplate luminometer with direct injectors to automatically dispense reagents during sample analysis. Enzymatic conversions of inosine to hypoxanthine, followed by hypoxanthine to xanthine to uric acid, generated superoxide anion radicals as a useful metabolic by‐product. The free radicals react with Pholasin®, a sensitive photoprotein used for chemiluminescence detection, to produce measurable blue‐green light. The use of Pholasin® and a chemiluminescence signal enhancer, Adjuvant‐K?, eliminated the need for plasma clean‐up steps prior to analysis. The method used 20 μL of heparinized plasma, with complete analysis of total hypoxanthine levels (inosine is metabolized to hypoxanthine using purine nucleoside phosphorylase) in approximately 3.7 min. The rapid chemiluminescence method demonstrated the capability of differentiating total hypoxanthine levels between healthy individuals, and patients presenting with non‐traumatic chest pain and potential acute cardiac ischemia. The results support the potential use of chemiluminescence methodology as a diagnostic tool to rapidly screen for elevated levels of inosine and hypoxanthine in human plasma, potential biomarkers of acute cardiac ischemia.Copyright ©2009 John Wiley & Sons, Ltd. |
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| Keywords: | inosine hypoxanthine biomarker chemiluminescence Pholasin® acute cardiac ischemia |
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