Nanosensors-Assisted Quantitative Analysis of Biochemical Processes in Droplets

Abstract

: Here, we present a miniaturized lab-on-a-chip detecting system for an all-electric and label-free analysis of the emulsion droplets incorporating the nanoscopic silicon nanowires-based field-effect transistors (FETs). We specifically focus on the analysis of β-galactosidase e.g.activity, which is an important enzyme of the glycolysis metabolic pathway. Furthermore, the efficiency of the synthesis and action of β-galactosidase can be one of the markers for several diseases, e.g., cancer, hyper/hypoglycemia, cell senescence, or other disruptions in cell functioning. We measure the reaction and reaction kinetics-associated shift of the source-to-drain current I_sd in the system, which is caused by the change of the ionic strength of the microenvironment. With these results, we demonstrate that the ion-sensitive FETs are able to sense the interior of the aqueous reactors; thus, the conjunction of miniature nanosensors and droplet-based microfluidic systems conceptually opens a new route toward a sensitive, optics-less analysis of biochemical processes.

Keywords: droplet-based microfluidics; enzymatic reaction; lab-on-a-chip; nanosensor; point-of-care diagnostics; silicon nanowire-based field-effect transistor; β-galactosidase assay.

Figure 1

Concept of the work. (a) Schematic of the platform comprising a field-effect transistor chip as a sensor and a droplet microfluidics flow-cell as a sample delivery module. (b) The enzymatic reaction of β-galactosidase and ortho-nitrophenyl-β-galactoside (ONPG). (c) Ionic composition before and after the reaction, the arrows represent ion mobility. Field-effect transistor (FET) devices are located directly under the microfluidic channel allowing the detection of droplets one-by-one upon passing. Droplets act as a modulating liquid gate that leads to the tuning of the FET source and drain current.

Figure 2

FET sensors with microfluidic integration. (a) Ready to use devices where a polydimethylsiloxane (PDMS) flow cell is permanently attached to the FET chip with the microfluidic channel aligned directly over the array of FETs: (i) FET chip with 16 devices (ii), SEM magnification of a single FET with Ag/AgCl-modified gate electrode (iii), and Si honeycomb structure (iv). (b) Transfer characteristics of a single FET device (@V_sd = 0.1 V). (c) Design of the microfluidic cell: two inlets for reactants (orange and yellow circles), inlet for oil (green circle), and outlet (gray circle). Reactants mix in the red area and the mixture is incubated while passing the channel (blue area). Droplets are generated (green area, magnified below) and guided to the sensors (yellow area). (d) Images of the channel (top) and droplet generating structure (down).

Figure 3

FET response to variations in sample composition. (a) I_sd signal change with passing droplets. Local maxima correspond to every single droplet, while minima correspond to the oil separating them. (b) Transfer characteristics response to phosphate-buffered saline (PBS) buffer ionic strength variations. The calibration of the flow-rate test is shown in (c).

Figure 4

Data obtained from the assay with the subtracted effect of streaming potential. (a) Reaction with increasing substrate concentration and constant enzyme concentration at (a) 0.1 U and (b) 1 U. The reaction was detected at four time points. (c,d) show the averaged data from each respective set of experiments, calculating the difference between the maxima and minima of the peaks.

Figure 5

Kinetics map of the reaction for different time points, concentrations, and peak values.