Nanokit for single-cell electrochemical analyses

Abstract

The development of more intricate devices for the analysis of small molecules and protein activity in single cells would advance our knowledge of cellular heterogeneity and signaling cascades. Therefore, in this study, a nanokit was produced by filling a nanometersized capillary with a ring electrode at the tip with components from traditional kits, which could be egressed outside the capillary by electrochemical pumping. At the tip, femtoliter amounts of the kit components were reacted with the analyte to generate hydrogen peroxide for the electrochemical measurement by the ring electrode. Taking advantage of the nanotip and small volume injection, the nanokit was easily inserted into a single cell to determine the intracellular glucose levels and sphingomyelinase (SMase) activity, which had rarely been achieved. High cellular heterogeneities of these two molecules were observed, showing the significance of the nanokit. Compared with the current methods that use a complicated structural design or surface functionalization for the recognition of the analytes, the nanokit has adapted features of the well-established kits and integrated the kit components and detector in one nanometer-sized capillary, which provides a specific device to characterize the reactivity and concentrations of cellular compounds in single cells.

Electrochemical Monitoring of ROS/RNS Homeostasis Within Individual Phagolysosomes Inside Single Macrophages

“…Thesame procedure was used to compare the homeostatic performances of different phenotypes of RAW2 64.7 murine macrophages,t hat is,i nt he resting stage (M0 type) or after their 24 hincubation (M1 type) with LPS/IFN-g. [4,5,15,16] Both phenotypes exhibited extremely different morphologies and diverse mean intracellular ROS/RNS concentrations as revealed, respectively,b yb right-field microscopy and DCFH-DAf luorescent staining (see insets in Figure 3A). [17] Figures 3B,C (Supporting Information, Table S2) show that the mean Q 0 value for M1 cells (18 fC) was nearly double that (9.1 fC) for M0 cells.…”

Electrochemical Monitoring of ROS/RNS Homeostasis Within Individual Phagolysosomes Inside Single Macrophages

“…The same procedure was used to compare the homeostatic performances of different phenotypes of RAW 264.7 murine macrophages, that is, in the resting stage (M0 type) or after their 24 h incubation (M1 type) with LPS/IFN‐γ . Both phenotypes exhibited extremely different morphologies and diverse mean intracellular ROS/RNS concentrations as revealed, respectively, by bright‐field microscopy and DCFH‐DA fluorescent staining (see insets in Figure A)…”

Intracellular Electrochemical Sensing

“…As another example, a nanometer‐sized capillary with a ring electrode was used to detect intracellular glucose (Figure B) . There, a GOx solution filled the tip capillary, and the capillary was inserted into cells.…”

“…There, a GOx solution filled the tip capillary, and the capillary was inserted into cells. It was then pumped into the cells, and the reaction by‐product was detected using the ring electrode .…”