Passivated Covalently Functionalized Quartz Crystal Microbalance for Reusable Detection of Small Proteins

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This thesis outlines the potential of quartz crystal microbalance (QCM)for small protein detection. Such small protein is a signaling protein named cytokine which is related to various diseases. Currently used biosensors are an extension of the aptamer-based sensor described in Yue Ling's thesis [1].
Covalently functionalized QCM crystals were tested with Tumour Necrosis Factor Alpha (TNF-α) on three aspects, for instance, sensitivity, reusability, and selectivity. The detection method was injecting solution contained target cytokine into a chamber with QCM crystal and comparing the frequency difference before and after the introduction. For the selectivity test, another cytokine, Interleukin 6(IL-6) was presented, and frequency shift should not be observed. To reuse the biosensors, they were rinsed with 7M urea and phosphate buffered saline, respectively.
In the sensitivity test, the biosensors had successfully detected 0.1 ug/ml of TNF-α. The modified urea wash method also demonstrated its reusable for further experiments. Whereas the biosensors have shown response to the nontarget cytokine, IL-6. To alleviate response to nontarget cytokine, the functionalization process was modified by adding a blocking agent, ethanolamine. This chemical substance will forma passivation layer on the surfaces of the biosensors resulting in the reduction of nonreacted surface groups (GOPS-CDI) binding. The following experiments with these passivated QCM crystals have shown improvement in selectivity, and the sensitivity of the crystals was not impacted. Rather, the key factor of maintaining the sensitivity is related to the aptamer concentration on the crystal surface.
Further studies on the selectiveness of biosensors will be needed to examine the effectiveness of ethanolamine with a relatively consistent amount of aptamers on the QCM crystal.
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