Hydrogen sulfide (H 2 S) is emerging as an important gasotransmitter in both physiological and pathological states. Rapid measurement of H 2 S 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) H 2 S 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 2 ≥ 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 H 2 S (range 0–100 µM). We have measured the baseline level of endogenous H 2 S 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 H 2 S 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 H 2 S in health and disease.

The levels of hydrogen peroxide ($${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 ) in human blood is of great relevance as it has emerged as an important signalling molecule in a variety of disease states. Fast and reliable measurement of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 levels in the blood, however, continues to remain a challenge. Herein we report an automated method employing a microfluidic device for direct and rapid measurement of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 in human blood based on laser-induced fluorescence measurement. Our study delineates the critical factors that affect measurement accuracy—we found blood cells and soluble proteins significantly alter the native $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 levels in the time interval between sample withdrawal and detection. We show that separation of blood cells and subsequent dilution of the plasma with a buffer at a ratio of 1:6 inhibits the above effect, leading to reliable measurements. We demonstrate rapid measurement of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 in plasma in the concentration range of 0–49 µM, offering a limit of detection of 0.05 µM, a sensitivity of 0.60 µM−1, and detection time of 15 min; the device is amenable to the real-time measurement of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 in the patient’s blood. Using the linear correlation obtained with known quantities of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 , the endogenous $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 concentration in the blood of healthy individuals is found to be in the range of 0.8–6 µM. The availability of this device at the point of care will have relevance in understanding the role of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 in health and disease.

The levels of hydrogen peroxide (H2O2) in human blood is of great relevance as it has emerged as an important signalling molecule in a variety of disease states. Fast and reliable measurement of H2O2 levels in the blood, however, continues to remain a challenge. Herein we report an automated method employing a microfluidic device for direct and rapid measurement of H2O2 in human blood based on laser-induced fluorescence measurement. Our study delineates the critical factors that affect measurement accuracy – we found blood cells and soluble proteins significantly alter the native H2O2 levels in the time interval between sample withdrawal and detection. We show that separation of blood cells and subsequent dilution of the plasma with a buffer at a ratio of 1:6 inhibits the above effect, leading to reliable measurements. We demonstrate rapid measurement of in plasma in the concentration range of 0 – 49 µM, offering a limit of detection of 0.05 µM, a sensitivity of 0.60 µM-1, and detection time of 15 min; the device is amenable to the real-time measurement of H2O2 in the patient’s blood. Using the linear correlation obtained with known quantities of H2O2, the endogenous H2O2 concentration in the blood of healthy individuals is found to be in the range 2 – 6 µM. The availability of this device at the point of care will have relevance in understanding the role of H2O2 in health and disease.

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Life evolved in an euxinic world with subsequent oxic 'invasion' leading to two parallel but interconnected biospheres, hydrogen sulphide (H2S) and hydrogen peroxide (H2O2) exemplify these worlds respectively. Their concentration gradients have informational value in meromictic lakes. Similarly, it is posited, there exists a whole body chemocline in humans in which the two molecules form an inversely coupled oxic/sulphidic oscillator (OSO). The OSO is hormetic and characterised by a range of amplitudes and frequencies in health. Deviations from its baseline profile heralds the onset of SIRS before the appearance of clinical signs. Loss of oscillator status and transition to a steady state causes widespread intercellular and inter-organ communication failure presaging multi-organ dysfunction. The salient clinico-pathophysiological features of SIRS of any aetiology are emergent phenomena related to the OSO profile. Extent of recovery of organ function will mirror the recovery of the OSO profile thereby providing a tool to predict outcomes in SIRS.

Vasoplegia observed post-cardiopulmonary bypass (CPB), is associated with substantial morbidity, multiple organ failure and mortality. Circulating counts of hematopoietic stem cells (HSCs) and endothelial progenitor cells (EPC) are potential markers of neovascularization and vascular repair. However, changes in the circulating levels of these progenitors in perioperative CPB and their association with post-CPB vasoplegia remains to be determined. HSC and EPC counts were enumerated at different timepoints during CPB in 19 individuals who underwent elective cardiac surgery via flow cytometry. These 19 individuals were categorized into two groups based on severity of post-operative vasoplegia as: clinically insignificant vasoplegic Group 1 (G1) and clinically significant vasoplegic Group 2 (G2). The differential changes in progenitor cell counts during different stages of surgery were compared across these two groups.Machine-learning classifiers (logistic regression and gradient boosting) were employed to determine if differential changes in progenitor counts could differentiate and group subjects based on the severity of vasoplegia. Enumeration of progenitor cells revealed an early and significant increase in the circulating counts of CD34 + and CD34 + CD133 + hematopoietic stem cells (HSC) in G1 subjects which was attenuated in G2 individuals.Additionally, EPCs (CD34 + VEGFR2 + ) were lower in G2 individuals compared to G1.Gradient boosting outperformed logistic regression in assessing the vasoplegia grouping based on fold change in circulating CD 34 + levels. Our findings indicate that a lack of early response of CD34 + cells and CD34 + CD133 + HSCs might serve as an early marker for development of clinically significant vasoplegia after CPB.

Vasoplegia observed post cardiopulmonary bypass (CPB) is associated with substantial morbidity, multiple organ failure and mortality. Circulating counts of hematopoietic stem cells (HSCs) and endothelial progenitor cells (EPC) are potential markers of neo-vascularization and vascular repair. However, the significance of changes in the circulating levels of these progenitors in perioperative CPB, and their association with post-CPB vasoplegia, are currently unexplored. We enumerated HSC and EPC counts, via flow cytometry, at different time-points during CPB in 19 individuals who underwent elective cardiac surgery. These 19 individuals were categorized into two groups based on severity of post-operative vasoplegia, a clinically insignificant vasoplegic Group 1 (G1) and a clinically significant vasoplegic Group 2 (G2). Differential changes in progenitor cell counts during different stages of surgery were compared across these two groups. Machine-learning classifiers (logistic regression and gradient boosting) were employed to determine if differential changes in progenitor counts could aid the classification of individuals into these groups. Enumerating progenitor cells revealed an early and significant increase in the circulating counts of CD34+ and CD34+CD133+ hematopoietic stem cells (HSC) in G1 individuals, while these counts were attenuated in G2 individuals. Additionally, EPCs (CD34+VEGFR2+) were lower in G2 individuals compared to G1. Gradient boosting outperformed logistic regression in assessing the vasoplegia grouping based on the fold change in circulating CD 34+ levels. Our findings indicate that a lack of early response of CD34+ cells and CD34+CD133+ HSCs might serve as an early marker for development of clinically significant vasoplegia after CPB.