Cortisol in the human hair is a clinical biomarker of long-term social-and-work-related mental stress, which is of high morbidity rate in the current modern society. This study developed a sensitive hair cortisol assay, featuring surface- enhanced Raman spectroscopy, immunoreaction, and a double-layer paper microdevice. The first layer of the paper microdevice was used to remove the hair residue in the hair extract by filtration (sample pretreatment). The second layer was used for competitive immunoreaction and detection. Standard cortisol antigen immobilized in the second layer and the free cortisol in hair extract competed to bind the spiked Raman-active cortisol monoclonal antibody solution. The hair cortisol can be quantitated by the intensity of Raman signal of monoclonal antibody bound on the paper. We found the Raman signal decreased as the cortisol concentration increases in hair samples. The relative Raman intensity measured was linearly proportional to the logarithmic value of the cortisol concentration in hair samples we measured. The detection of limit (LOD) was 1 pg/mL with the portable Raman spectrometer. The RSD of measurement was 8.38% (n=6). In addition, we used LC-MS to measure two real samples as a comparison with our method as above. The results are 0.771 and 0.153 ng/mL by LC-MS method and 0.63 and 0.247 ng/mL by the proposed method. It can be observed that the results are in same order, demonstrating the validity of the proposed method. In addition, 48 samples can be measured in a single chip. These results showed that this method is sensitive, specific, and suitable for large-scale screening of hair cortisol samples.

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基于便携式拉曼光谱仪和双层纸微器件的头发皮质醇超灵敏检测

人头发中的皮质醇是长期与社会和工作相关的精神压力的临床生物标志物，在当今现代社会发病率很高。本研究开发了一种灵敏的头发皮质醇检测方法，具有表面增强拉曼光谱、免疫反应和双层纸微型装置。纸微装置的第一层用于通过过滤（样品预处理）去除毛发提取物中的毛发残留物。第二层用于竞争性免疫反应和检测。固定在第二层中的标准皮质醇抗原和头发提取物中的游离皮质醇竞争结合加标的拉曼活性皮质醇单克隆抗体溶液。毛发皮质醇可以通过结合在纸上的单克隆抗体的拉曼信号强度来定量。我们发现拉曼信号随着头发样本中皮质醇浓度的增加而降低。测量的相对拉曼强度与我们测量的头发样品中皮质醇浓度的对数值成线性比例。便携式拉曼光谱仪的检测限 (LOD) 为 1 pg/mL。测量的 RSD 为 8.38% (n=6)。此外，我们使用 LC-MS 测量两个真实样品，作为与我们上述方法的比较。LC-MS 方法的结果为 0.771 和 0.153 ng/mL，建议方法的结果为 0.63 和 0.247 ng/mL。可以看出，结果的顺序相同，证明了所提出方法的有效性。此外，单芯片可测量48个样品。这些结果表明该方法灵敏、特异，适用于毛发皮质醇样品的大规模筛选。