title: A rapid and simple chemiluminescence method for screening levels of inosine and hypoxanthine in non-traumatic chest pain patients.
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.
authors.affiliation: Department of Internal Medicine, Divisions of Nephrology, Clinical Pharmacology and Hypertension, Virginia Commonwealth University, Richmond, VA, USA. defarthi@vcu.edu
authors.forename: Don E
authors.initials: DE
authors.lastname: Farthing
authors.forename: Domenic
authors.initials: D
authors.lastname: Sica
authors.forename: Michael
authors.initials: M
authors.lastname: Hindle
authors.forename: Les
authors.initials: L
authors.lastname: Edinboro
authors.forename: Lei
authors.initials: L
authors.lastname: Xi
authors.forename: Todd W B
authors.initials: TW
authors.lastname: Gehr
authors.forename: Lynne
authors.initials: L
authors.lastname: Gehr
authors.forename: Christine A
authors.initials: CA
authors.lastname: Farthing
authors.forename: Terri L
authors.initials: TL
authors.lastname: Larus
authors.forename: Itaf
authors.initials: I
authors.lastname: Fakhry
authors.forename: H Thomas
authors.initials: HT
authors.lastname: Karnes
mesh_terms: D002637:Chest Pain; D006801:Humans; D019271:Hypoxanthine / Q000097:blood*; D007288:Inosine / Q000097:blood*; D008163:Luminescent Measurements / Q000379:methods*; D015394:Molecular Structure; D017202:Myocardial Ischemia / Q000175:diagnosis; D012015:Reference Standards; D013997:Time Factors
chemical_list: D019271:Hypoxanthine; D007288:Inosine