研發自動化檢測人體血清中總抗氧化能力之測定方法及其臨床應用

Development of a high throughput automated measurement of serum total antioxidative capacity and its clinical application

陳季芬(Chi-Fen Chen)

指導教授 ： 蔡麗玉

高雄醫學大學/健康科學院/生物醫學檢驗學研究所/碩士(2007年)

摘要

無資料

關鍵字

總抗氧化能力 ； 自動化 ； 臨床應用

並列摘要

There is overwhelming evidence to indicate that reactive oxygen species (ROS) have been implicated in the etiology of a host of degenerative diseases including cardiovascular disease, cancer and a variety of inflammatory disorders. If ROS are involved in all of these clinical conditions, then antioxidants should be effective in preventing their occurrence. For these reasons, there is a considerable interest in the use of certain methods to assessment the total antioxidative capacity (TAC) aiming to replenish the needed antioxidants to correct the deficit of prooxidant / antioxidant imbalance. Thus far, a variety of methodologies have been devised for the measurement of TAC in the sera. However, these methods are complicated and required multiple procedural steps. Along the same vein, these procedures are incapable of large-scale screening of TAC because they are not being able to automate. For these reasons, it is desirable to develop a rapid and facile method for assaying TAC and is capable of automating for a large scale screening purpose. In this project, I have firstly devised a novel microassay method for rapid screening of TAC in human sera. The method was also being automated using Olympus AU 600 Chemistry Autoanalyzer. The principle of the method is based upon the concept that various antioxidants in serum, being recognizes as reluctant, are capable of reducing a ferric tripyridytriazine (Fe3+-TPTZ), to its Fe2+-TPTZ complex which exhibits a maximum absorption as 593 nm. The automated procedure has the following analytic parameters; Pipettor settings are : sample volume = 2 μL and reagent volume = 300 μL; analyzer settings are : reaction mode = endpoint; wavelength = 600 nm, temperature = 37℃, absorbance readings number = 27, interval = 18 sec. The results of serum TAC were reported as mmole/L Trolox-eguivalent wait (mmole/L TEU). The assay performed under our optimum condition has excellent precision data, with within-run and day-to-day reproducibilities (expressed as CVs) as three control levels were all below 3.0%. The recovery studies revealed an average of 101.5 ± 4.0%.The stability studies revealed that specimens freshly drawn serum or storage at -20℃ could stable for 21days. The reference range has ales been established which was in the range of 0.53 to 0.97 mmole/L TEU (n = 850). The proposed TAC assay has a throughput of 300 tests/hr. Also, the clinical utility was exemplified by measuring the TAC values for a group of liver disease (n = 82), pneumonia (n = 41)and malignancy without icteric (n = 154). The TAC values for all these diseased groups were significantly lower than the normal reference range (P<0.001). Collectively, our proposed high throughput microassay for TAC has been shown to be simple, rapid and clinically useful in identifying an individual who may be suffering from antioxidant depletion further studies may be necessary to evaluate the interference exerted by specimens that may contain extremely high bilirubin, uric acid and creatinine concentrations. These studies are currently under investigation.

並列關鍵字

Total antioxidative capacity ； automated ； clinical application

參考文獻

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