Department of Mechanical Engineering, Indian Institute of Technology Madras, Chennai, 600036, India.
Department of Chemistry, Indian Institute of Technology Madras, Chennai, 600036, India.
Sci Rep. 2019 Mar 1;9(1):3258. doi: 10.1038/s41598-019-39389-7.
Hydrogen sulfide (HS) is emerging as an important gasotransmitter in both physiological and pathological states. Rapid measurement of HS remains a challenge. We report a microfluidic method for rapid measurement of sulphide in blood plasma using Dansyl-Azide, a fluorescence (FL) based probe. We have measured known quantities of externally added (exogenous) HS to both buffer and human blood plasma. Surprisingly, a decrease in FL intensity with increase in exogenous sulphide concentration in plasma was observed which is attributed to the interaction between the proteins and sulphide present in plasma underpinning our observation. The effects of mixing and incubation time, pH, and dilution of plasma on the FL intensity is studied which revealed that the FL assay required a mixing time of 2 min, incubation time of 5 min, a pH of 7.1 and performing the test within 10 min of sampling; these together constitute the optimal parameters at room temperature. A linear correlation (with R ≥ 0.95) and an excellent match was obtained when a comparison was done between the proposed microfluidic and conventional spectrofluorometric methods for known concentrations of HS (range 0-100 µM). We have measured the baseline level of endogenous HS in healthy volunteers which was found to lie in the range of 70 μM - 125 μM. The proposed microfluidic device with DNS-Az probe enables rapid and accurate estimation of a key gasotransmitter HS in plasma in conditions closely mimicking real time clinical setting. The availability of this device as at the point of care, will help in understanding the role of HS in health and disease.
硫化氢（HS）在生理和病理状态下都是一种重要的气体递质。快速测量 HS 仍然是一个挑战。我们报告了一种使用荧光（FL）探针 Dansyl-Azide 快速测量血浆中硫化物的微流控方法。我们已经测量了已知数量的外源性 HS 添加到缓冲液和人血浆中。令人惊讶的是，在血浆中外源硫化物浓度增加时，FL 强度降低，这归因于存在于血浆中的蛋白质和硫化物之间的相互作用，这支持了我们的观察。研究了混合和孵育时间、pH 值和血浆稀释对 FL 强度的影响，结果表明 FL 测定需要 2 分钟的混合时间、5 分钟的孵育时间、pH 值为 7.1 并在采样后 10 分钟内进行测试；这些共同构成了室温下的最佳参数。当将提出的微流控和传统荧光光谱法用于已知 HS 浓度（范围 0-100μM）进行比较时，获得了线性相关性（R≥0.95）和极好的匹配。我们已经测量了健康志愿者内源性 HS 的基线水平，发现其范围在 70μM-125μM 之间。带有 DNS-Az 探针的提出的微流控装置能够在非常接近实时临床环境的条件下快速准确地估计血浆中的关键气体递质 HS。这种设备在即时护理点的可用性将有助于了解 HS 在健康和疾病中的作用。
