紅外光免疫型生化感測器之研製

Development of Infrared Biosensor for Immunoassays Based on Reflection Absorption Spectroscopy

劉俞青(Yu-Ching Liu)

指導教授 ： 楊吉斯

中原大學/理學院/化學研究所/碩士(2002年)

https://doi.org/10.6840/cycu200200098

摘要

任何有機分子在紅外光區均有特殊的吸收峰，藉由這項優點，紅外光能克服傳統生化物質檢測時，需事先標定螢光劑的困擾，達到方便快速檢測的目的。為去除水分子對紅外光的干擾，以及簡化量測裝置，本實驗自行研製反射吸收之測量裝置，以傳統霍式紅外光譜儀測量時樣品直徑可低於3 mm以下。本研究利用銀與酸基之特殊鍵結性，將抗原固定化於反射銀基板上，再利用反射吸收法，測取固化在銀片上紅外光訊號。為有效改善固化後生化分子在水溶液中不穩定的問題，除了加強固化效果之外，實驗中更引入交聯劑，藉由交聯方式來增加生物分子的穩定性。為取得最適化實驗條件，研究過程對光學系統、抗原與交聯劑間的比例、固定化時間，及固定後浸泡於緩衝溶液中穩定性、對抗體分子的選擇性、再現性上均有詳細的探討。由實驗結果得知，以反射吸收法配合紅外線光譜儀之偵測方式能提供足夠之偵測訊號，由測量特定抗體之結果顯示，抗體經由本研究發展之方法固化後仍保有其原先之專一性，由此更證明以紅外光配合抗原抗體之特殊選擇性，可以達到測量生化性樣品之目的，綜觀而言，本研究中所開發之測量法具有取代傳統生化物質測量方法之潛力。

關鍵字

反射吸收法 ； 生化感測器 ； 傅立葉紅外光光譜儀

並列摘要

To eliminate the tedious preparation steps in conventional immunoassays type of methods, infrared sensors based on reflection-absorption mode were developed in this work. Because infrared radiation can interact with any organic molecules, the use of infrared radiation to produce analytical signals can remove the steps in labeling of fluorescencing molecules such as in fluorescence detection method. This new method utilized an immobilizing antibody to absorb its corresponding antigen and in consequence, resulting more intense infrared signal by the absorbed antigen. To study the performance of this proposed method, three pairs of antibody/antigen proteins were used as probe proteins and their infrared absorption bands were carefully examined in order to obtain the best band position for quantitative analyses.
To obtain the optimal conditions for detections, the effect of optical system, the stability of immobilized protein, and the selectivity in detection of antibody/antigen pair were investigated. Results in the construction of optical system indicated that with large incident angle of infrared radiation, the signals of absorbed protein were enhanced tremendously. Meanwhile, the size of the aperture in the optical system should be close to the size of the deposited protein to effectively remove the un-interacted radiations. In the immobilization of protein in reflection substrates, three materials including silver, copper, and stainless steel were examined. Results indicated that silver plate is not only providing the highest reflection capability but also strongly interacting with the protein molecules deposited on it. With 2.5 %(v/v) of glutardialdehyde as cross-linking agent, the deposited protein on silver plate was found to be highly stable in buffer solutions. By blocking with suitable materials, such as polyvinyl alcohol or poly allyl amine, nonspecific interactions between proteins can be largely reduced. Using the optimal conditions, the infrared detection of proteins through the principle of immunology was achieved in this work.

並列關鍵字

FT-IR ； Biosensors ； Reflection Absorption

參考文獻

1. Hulanicki, A.; Glab, S.; Lngman, F. (1989) IUPAC Discussion Paper, Commission, Ⅵ, July.

6. Byfield, M. P.; Abuknesha, R. A. Biosens. Bioelectron. 1994, 9, 373-400.

7. Rieger, P. H. Electrochemistry, Hall, New York. 1994.

9. Uttenthaler, E.; Stephan, D. Biosens. Bioelectron. 1998, 13, 1279-1286.

10. Wessa, T.; Rapp. M.; Ache, H. J. Biosensor and Bioelectronics, 1999, 14, 93-98.