Development of an enzymatic chromatography strip with nicotinamide adenine dinucleotide–tetrazolium coupling reactions for quantitative l-lactate analysis
| Autor: | Kuang-Pin Hsiung, Min-Chi Lan, Wei-Feng Chang, Chia-Chi Lin, Wei-Shiang Lai, Shu-Chen Kan, Yung-Chuan Liu, Chwen-Jen Shieh |
|---|---|
| Rok vydání: | 2015 |
| Předmět: |
Biophysics
Tetrazolium Salts Nicotinamide adenine dinucleotide Biochemistry chemistry.chemical_compound Limit of Detection Lactate dehydrogenase Diaphorase Animals Humans Lactic Acid Molecular Biology Reagent Strips Detection limit Chromatography L-Lactate Dehydrogenase Cell Biology Hydrogen-Ion Concentration Enzymes Immobilized NAD Clostridium kluyveri Lactic acid Kinetics chemistry Rabbits NAD+ kinase Formazan Quantitative analysis (chemistry) |
| Zdroj: | Analytical Biochemistry. 471:61-66 |
| ISSN: | 0003-2697 |
| DOI: | 10.1016/j.ab.2014.11.015 |
| Popis: | 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, 0.25 U/μl), and NAD + (2 μl, 1.5 × 10 −5 M) were added into the mobile phase (100 μl) composed of 0.1% (w/w) Tween 20 in 10 mM 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 5 mM 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. |
| Databáze: | OpenAIRE |
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