The efficient recognition of circulating tumor cells (CTCs) with an aptamer probe confers numerous benefits; however, the stability and binding affinity of aptamers are significantly hampered in real biological sample matrices. Inspired by the efficient preying mechanism by multiplex tubing feet and endoskeletons of sea urchins, we engineered a superefficient biomimetic single-CTC recognition platform by conjugating dual-multivalent-aptamers (DMAs) Sgc8 and SYL3C onto AuNPs to form a sea urchin-like nanoprobe (sea urchin-DMA-AuNPs). Aptamers Sgc8 and SYL3C selectively bind with the biomarker proteins PTK7 and EpCAM expressed on the surface of CTCs. CTCs were captured with 100% efficiency, followed by sorting on a specially designed multifunctional microfluidic configuration, integrating a single-CTC separation unit and a hydrodynamic filtrating purification unit. After sorting, background-free analysis of biomarker proteins in single CTCs was undertaken with inductively coupled plasma mass spectrometry by measuring the amount of 197Au isotope in sea urchin-DMA-AuNPs. With respect to a single-aptamer nanoprobe/-interface, the dual-aptamer nanoprobe improves the binding efficiency by more than 200% (K d < 0.35 nM). The microchip facilitates the recognition of single CTCs with a sorting separation rate of 93.6% at a flow rate of 60 μL min–1, and it exhibits 73.8 ± 5.0% measurement efficiency for single CTCs. The present strategy ensures the manipulation and detection of a single CTC in 100 μL of whole blood within 1 h.

Single-cell analysis facilitates perception into the most essential processes in life’s mysteries. While it is highly challenging to quantify them at the single-cell level, where precise single-cell sampling is the prerequisite. Herein, a real-time single-cell quantitative platform was established for high-throughput droplet-free single-cell sampling into time-resolved (TRA) ICP-MS and real-time quantification of intracellular target elements. The concentrated cells (2 × 106 cells mL–1) were spontaneously and orderly aligned in a spiral microchannel with 104 periodic dimensional confined micropillars. The quantification is conducted simultaneously by internal standard inducing from another branch channel in the chip. The flow-rate-independent feature of single-cell focusing into an aligned stream within a wide range of fluidic velocities (100–800 μL min–1) facilitates high-throughput, oil-free, single-cell introduction into TRA-ICP-MS. The system was used for real-time exploration of intracellular antagonism of Cu2+ against Cd2+. an obvious antagonistic effect was observed for the MCF-7 cell by culturing for 3, 6, 9, and 12 h with 100 μg L–1 Cd2+ and 100 μg L–1 Cu2+, and a rivalry rate of 12.8% was achieved at 12 h. At identical experimental conditions, however, limited antagonistic effect was encountered for a bEnd3 cell within the same incubation time period, with a rivalry rate of 4.81%. On the contrary, an antagonistic effect was not observed for the HepG2 cell by culturing for 6 h, while an obvious antagonistic effect was found by further culturing to 12 h, with a rivalry rate of 10.43%. For all three cell lines, significant heterogeneity was observed among individual cells.

In this study, we show a more efficient method for isolation and cultivation of dermal papilla cells from hair follicles of human scalp skin. The dermal partments of low hair follicles were pulled out from cutaneous fat and the bulb epithelium was teased out from the fibrous sheath with attached dermal papilla by applying gentle pressure with the tip of an occal forceps. When these fibrous sheaths were entirely digested into isolated cells by collagenase D but the dermal papillae were justly to be digested, collagenase D was discarded and the dermal papillae were isolated completely out from the resuspension solution by repeated low-speed centrifugation and transferred to another dish for free-floating culture. This procedure markedly simplifies the steps of isolated dermal papilla operation and relieves the laborious tension. Furthermore, dermal papillae could be isolated on a large-scale and remained intact. After collagenase digestion, the dermal papillae showed very high adherent rate and quicker growth than that of microdissection, which suggests that the definition factor of dermal papilla cell migration was relaxed and some structure had been activated or exposed. The cells exhibited a multi-layer forming property and spread-out growth style. They showed positive with alcian blue, with toluidine blue O for different gradient pH and PAS, which was similar to the staining results of in situ dermal papilla. It suggests that the culture papilla cells still synthesize and excrete neutral and acid mucopolysaccharides. Our results demonstrate that the papilla cells in culture condition still remain the ability to synthesize the specific extracellular matrix components of in situ dermal papilla, which supports the concept that the dermal papilla cell, a highly specialized fibroblast, especially is involved in hair growth regulation.

A two-dimensional cytometry platform (CytoLM) with high sensitivity and high temporal resolution is developed for single-particle and single-cell sampling and analysis. First, a Dean flow-assisted vortex capillary cell sampling (VCCS) unit confines the sample stream in curved flow and drives to focus and align the particles or cells in a small probe volume. By coupling VCCS to a laser-induced fluorescence (LIF) detector with data acquisition and processing capability, a high-throughput single-particle/cell analysis system (VCCS–LIF) was established. The particle analysis throughput of 119.42/s and a detection recovery of 78.20 ± 1.75% were achieved at a density of 9.16 × 104/mL for fluorescent particles, and the cell analysis throughput is 48.20/s at a density of 1.5 × 105/mL. Second, the CytoLM platform is constructed by hyphenating VCCS–LIF with inductively coupled plasma mass spectrometry (ICP–MS). In the analysis of HepG2 cells by Ag+ incubation and AO staining, 10,760 fluorescence bursts and 3068 MS events were observed in 240 s. Invalid signals due to undispersed cells were controlled at 3.80% for LIF and 1.01% for MS, with a proportion of effective signal of >96.20%. After peak identification and integral processing of the original data, the statistical results including peak area, height, width, and spacing are obtained concurrently and the information on concentration and elemental quantification of single cells is evaluated. CytoLM facilitates high-throughput, multi-dimensional, and multi-parameter characterization of particles and cells, and it may provide vast potential in life science analysis.

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