基于富集微流控芯片的稻曲病菌孢子光电检测方法

    Photoelectric detection method for false smut fungal spore based on enrichment microfluidic chip

    • 摘要: 针对当前水稻真菌病害发病时间短、传播速度快,缺乏有效早期预警技术的难点,提出一种基于微流控芯片的空气流中水稻真菌病害光电检测方法。该文根据微尺度下孢子富集动力学特征设计了病害孢子高效富集微流控芯片,并结合光电检测系统进行病害孢子的检测。试验根据空气动力学原理以及孢子富集量的大小对微流控芯片通道尺寸进行设置。根据光检测原理和不同浓度孢子在富集区形成的光衰减特性,筛选具有高灵敏度特性的检测光强。在以检测光谱的灵敏度性能为目标并综合考虑线性度的基础上,优选检测波长。以水稻稻曲病菌孢子为检测对象,进行了自动化微流控富集和光电检测试验。试验结果表明:在光源光强1.1×104 cd,波长为650 nm时,利用所述检测方法针对水稻稻曲病菌孢子检测结果(相比与镜检值)误差小于17.5%,根据检测结果建立相关系数为0.992 9的检测模型,具有较好的线性度及可靠性。研究结果为便携式作物病害检测装备的研发提供理论基础。

       

      Abstract: Abstract: The fungal disease of false smut is research focus, which can spread around within short time and cause much loss of rice yield. However, there hasn't been any effective detection and prediction method. For this reason, we designed a set of sample interface, injection channel, enrichment detection area, electric pumping interface in one of the microfluidic chips. The sample interface is connected to the hose for the collection of fungal spores. The injection channel adopts the simple and gentle straight-through design with a fixed width, in order to achieve the barrier-free movement of the fungal spores in the channel, making it easy to clean the chip, and thus improving the recycle rate of the chip. The enrichment detection area is designed as a circular chamber. This design not only slows the drop rate of the spores, which therefore makes the spores arranged in the enrichment detection area efficiently, but it also matches the circular parallel light spot formed by the fiber output lens. In this way, we can reduce the interference and noise introduced by positioning the light source inaccurately and too large spot diameter. Electric suction interface accesses micro-pump pipeline, and compared to the inflatable design, the pumping design reduces the loss in the pump during the collection of spores. In order to make the fungal spores be arranged in the enrichment detection area correctly, the sample channel width was set in the experiment and the diameter of the enrichment detection area was optimized to meet the needs of fungal spore enrichment speed. The experimental results showed that the enriched detection zone had the best enrichment effect when the diameter was 2 000 μm. Then, a photoelectric detection system based on microfluidic chip was established in this paper. When the spore concentration of microfluidic chip was detected by photoelectric detection method, the system condition parameters mainly included light intensity and wavelength. Therefore, different concentrations of spores were tested by the spectrometer, and the results were analyzed to determine the optimal light source parameters of the detection system. The experiment was performed at the Jiangsu University Laboratory in April 2017, and the spores used in the experiment were obtained in the artificial climate room in Jiangsu Vocational College of Agriculture and Forestry. We used the spectrometer, fiber and other structures to build the experimental platform for light intensity and wavelength optimization. According to the principle of light detection and the characteristics of light attenuation formed by different concentrations of spores in the enrichment area, the detection intensity with high sensitivity was screened. Taking the detection of the sensitivity performance of the spectrum as the target and considering the linearity, the detection wavelength was determined; the best light intensity was at 1.1×104 cd and the best wavelength was 650 nm. Finally, with strawberry grey mold spores as the object, we conducted an automated microfluidic chip enrichment and photoelectric detection experiment, and established a curve of spore concentration and absorbance based on the principle of enrichment and photo-detection. The experimental results showed that the detection method we proposed had a linear correlation coefficient of 0.992 9. The method of photoelectric detection of false smut fungal disease based on microfluidic chip realizes the efficient detection of spores concentration, which provides a theoretical foundation for the development of portable false smut disease detection equipment.

       

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