Rapid measurement of hydrogen sulphide in human blood plasma using a microfluidic method.

Hydrogen sulfide (H2S) is emerging as an important gasotransmitter in both physiological and pathological states. Rapid measurement of H2S 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) H2S 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 R2 ≥ 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 H2S (range 0-100 µM). We have measured the baseline level of endogenous H2S 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 H2S 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 H2S in health and disease.

A microfluidic device with focusing and spacing control for resistance-based sorting of droplets and cells.

This paper reports a novel hydrodynamic technique for sorting of droplets and cells based on size and deformability. The device comprises two modules: a focusing and spacing control module and a sorting module. The focusing and spacing control module enables focusing of objects present in a sample onto one of the side walls of a channel with controlled spacing between them using a sheath fluid. A 3D analytical model is developed to predict the sheath-to-sample flow rate ratio required to facilitate single-file focusing and maintain the required spacing between a pair of adjacent objects. Experiments are performed to demonstrate focusing and spacing control of droplets (size 5-40 µm) and cells (HL60, size 10-25 µm). The model predictions compare well with experimental data in terms of focusing and spacing control within 9%. In the sorting module, the main channel splits into two branch channels (straight and side branches) with the flow into these two channels separated by a "dividing streamline". A sensing channel and a bypass channel control the shifting of the dividing streamline depending on the object size and deformability. While resistance offered by individual droplets of different sizes has been studied in our previous work (P. Sajeesh, M. Doble and A. K. Sen, Biomicrofluidics, 2014, 8, 1-23), here we present resistance of individual cells (HL60) as a function of size. A theoretical model is developed and used for the design of the sorter. Experiments are performed for size-based sorting of droplets (sizes 25 and 40 µm, 10 and 15 µm) and HL60 cells (sizes 11 µm and 19 µm) and deformability-based sorting of droplets (size 10 ± 1.0 µm) and polystyrene microbeads (size 10 ± 0.2 µm). The performance of the device for size- and deformability-based sorting is characterized in terms of sorting efficiency. The proposed device could be potentially used as a diagnostic tool for sorting of larger tumour cells from smaller leukocytes.

Hydrodynamic resistance and mobility of deformable objects in microfluidic channels.

This work reports experimental and theoretical studies of hydrodynamic behaviour of deformable objects such as droplets and cells in a microchannel. Effects of mechanical properties including size and viscosity of these objects on their deformability, mobility, and induced hydrodynamic resistance are investigated. The experimental results revealed that the deformability of droplets, which is quantified in terms of deformability index (D.I.), depends on the droplet-to-channel size ratio [Formula: see text] and droplet-to-medium viscosity ratio [Formula: see text]. Using a large set of experimental data, for the first time, we provide a mathematical formula that correlates induced hydrodynamic resistance of a single droplet [Formula: see text] with the droplet size [Formula: see text] and viscosity [Formula: see text]. A simple theoretical model is developed to obtain closed form expressions for droplet mobility [Formula: see text] and [Formula: see text]. The predictions of the theoretical model successfully confront the experimental results in terms of the droplet mobility [Formula: see text] and induced hydrodynamic resistance [Formula: see text]. Numerical simulations are carried out using volume-of-fluid model to predict droplet generation and deformation of droplets of different size ratio [Formula: see text] and viscosity ratio [Formula: see text], which compare well with that obtained from the experiments. In a novel effort, we performed experiments to measure the bulk induced hydrodynamic resistance [Formula: see text] of different biological cells (yeast, L6, and HEK 293). The results reveal that the bulk induced hydrodynamic resistance [Formula: see text] is related to the cell concentration and apparent viscosity of the cells.