Development of an enzymatic chromatography strip with nicotinamide adenine dinucleotide-tetrazolium coupling reactions for quantitative l-lactate analysis.

Autor: Kan SC; Department of Chemical Engineering, National Chung Hsing University, Taichung 402, Taiwan., Chang WF; Department of Chemical Engineering, National Chung Hsing University, Taichung 402, Taiwan., Lan MC; Department of Chemical Engineering, National Chung Hsing University, Taichung 402, Taiwan., Lin CC; Department of Chemical Engineering, National Chung Hsing University, Taichung 402, Taiwan., Lai WS; Department of Chemical Engineering, National Chung Hsing University, Taichung 402, Taiwan., Shieh CJ; Biotechnology Center, National Chung Hsing University, Taichung 402, Taiwan., Hsiung KP; Research and Development Department, Taiwan Unison Biotech, Hsinchu 300, Taiwan., Liu YC; Department of Chemical Engineering, National Chung Hsing University, Taichung 402, Taiwan. Electronic address: ycliu@dragon.nchu.edu.tw.
Jazyk: angličtina
Zdroj: Analytical biochemistry [Anal Biochem] 2015 Feb 15; Vol. 471, pp. 61-6. Date of Electronic Publication: 2014 Nov 29.
DOI: 10.1016/j.ab.2014.11.015
Abstrakt: In this study, a dry assay of l-lactate via the enzymatic chromatographic test (ECT) was developed. An l-lactate dehydrogenase plus a nicotinamide adenine dinucleotide (NADH) regeneration reaction were applied simultaneously. Various tetrazolium salts were screened to reveal visible color intensities capable of determining the lactate concentrations in the sample. The optimal analysis conditions were as follows. The diaphorase (0.5 μl, 2(-6)U/μl) was immobilized in the test line of the ECT strip. Nitrotetrazolium blue chloride (5 μl, 12 mM), l-lactate dehydrogenase (1 μl, 0.25U/μl), and NAD(+) (2μl, 1.5×10(-5)M) were added into the mobile phase (100 μl) composed of 0.1% (w/w) Tween 20 in 10mM phosphate buffer (pH 9.0), and the process was left to run for 10 min. This detection had a linear range of 0.039 to 5mM with a detection limit of 0.047 mM. This quantitative analysis process for l-lactate was easy to operate with good stability and was proper for the point-of-care testing applications.
(Copyright © 2014 Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
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