Rigid and Conductive Dual Nanoprobe for Single Cell Analysis

2014 ◽  
Vol 69 (8) ◽  
Author(s):  
Abdul Hafiz Mat Sulaiman ◽  
Mohd Ridzuan Ahmad
Keyword(s):  
Single Cell ◽  
Single Cell Analysis ◽  
Single Cells ◽  
Electrical Measurement ◽  
Microfluidic System ◽  
General Information ◽  
Aluminium Copper ◽  
Element Approach ◽  
Cell Surgery ◽  
Electrical Characterizations

Electrical property characterization of a single cell can be used to infer about its physiological condition, e.g. cell viability.  Due to that, a dual nanoprobe-microfluidic system for electrical properties measurement of single cells has been proposed. This paper is concerned about the mechanical and electrical characterizations of the dual nanoprobe. Electrical and mechanical characterizations were conducted to measure the resistance and the strength of the dual nanoprobe for five different metals i.e. Aluminium, Copper, Silver, Tungsten, and Zinc using finite element approach. From the findings, Tungsten’s nanoprobe has the highest strength while the resistance values for the five materials are not significantly different. Therefore, Tungsten is selected as the most recommended metal for the dual nanoprobe. We also performed single cell electrical measurement to test the functionality of the sensor. This work provides general information of the nanoprobe which can be used as a framework in other applications involving Nano devices i.e. cell surgery and drug delivery.

Statistical single-cell analysis of cell cycle-dependent quantum dot cytotoxicity and cellular uptake using a microfluidic system

RSC Advances ◽  
2014 ◽  
Vol 4 (47) ◽  
pp. 24929-24934 ◽  
Author(s):  
Jing Wu ◽  
Haifang Li ◽  
Qiushui Chen ◽  
Xuexia Lin ◽  
Wu Liu ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Quantum Dot ◽  
Single Cell ◽  
Cellular Uptake ◽  
Single Cell Analysis ◽  
Single Cells ◽  
Microfluidic System ◽  
Cell Analysis ◽  
System P ◽  
Cycle Dependent

The response of single cells in different cell cycle phases to QD cytotoxicity studied on a microfluidic device.

A SIMULATION STUDY OF SINGLE CELL INSIDE AN INTEGRATED DUAL NANONEEDLE-MICROFLUDIC SYSTEM

2016 ◽  
Vol 78 (7-5) ◽  
Author(s):  
Muhammd Asraf Mansor ◽  
Mohd Ridzuan Ahmad
Keyword(s):  
Single Cell ◽  
Simulation Study ◽  
Single Cell Analysis ◽  
Biological Activities ◽  
Single Cells ◽  
Microfluidic Channel ◽  
Electrical Measurement ◽  
Solution Flow Rate ◽  
Solution Flow ◽  
Novel Method

Electrical properties of living cells have been proven to play significant roles in understanding of various biological activities including disease progression both at the cellular and molecular levels. Analyzing the cell’s electrical states especially in single cell analysis (SCA) lead to differentiate between normal cell and cancer cell. This paper presents a simulation study of micro-channel and nanoneedle structure, fluid manipulation and current flow through HeLa cell inside a microfluidic channel. To perform electrical measurement, gold dual nanoneedle has been utilized. The simulation result revealed, the cell penetration occurs at microchannel dimension and solution flow rate is 22 µm x 70 µm x 25 µm (width x length x height) and 0.396 pL/min, respectively. The purposed device has capability to characterize the electrical property of single cells can be used as a novel method for cell viability detection in instantaneous manner.

scholarly journals Microfluidic platform accelerates tissue processing into single cells for molecular analysis and primary culture models

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Jeremy A. Lombardo ◽  
Marzieh Aliaghaei ◽  
Quy H. Nguyen ◽  
Kai Kessenbrock ◽  
Jered B. Haun
Keyword(s):  
Single Cell ◽  
Single Cell Analysis ◽  
Single Cells ◽  
Cell Analysis ◽  
Processing Technologies ◽  
Microfluidic Platform ◽  
Tissue Processing ◽  
Single Cell Rna Sequencing ◽  
Culture Models ◽  
Tissue Digestion

AbstractTissues are complex mixtures of different cell subtypes, and this diversity is increasingly characterized using high-throughput single cell analysis methods. However, these efforts are hindered, as tissues must first be dissociated into single cell suspensions using methods that are often inefficient, labor-intensive, highly variable, and potentially biased towards certain cell subtypes. Here, we present a microfluidic platform consisting of three tissue processing technologies that combine tissue digestion, disaggregation, and filtration. The platform is evaluated using a diverse array of tissues. For kidney and mammary tumor, microfluidic processing produces 2.5-fold more single cells. Single cell RNA sequencing further reveals that endothelial cells, fibroblasts, and basal epithelium are enriched without affecting stress response. For liver and heart, processing time is dramatically reduced. We also demonstrate that recovery of cells from the system at periodic intervals during processing increases hepatocyte and cardiomyocyte numbers, as well as increases reproducibility from batch-to-batch for all tissues.

scholarly journals Multiplexed profiling of single-cell extracellular vesicles secretion

2019 ◽  
Vol 116 (13) ◽  
pp. 5979-5984 ◽  
Author(s):  
Yahui Ji ◽  
Dongyuan Qi ◽  
Linmei Li ◽  
Haoran Su ◽  
Xiaojie Li ◽  
...  
Keyword(s):  
Single Cell ◽  
Cell Lines ◽  
Extracellular Vesicles ◽  
Single Cell Analysis ◽  
Comprehensive Evaluation ◽  
Single Cells ◽  
Deep Understanding ◽  
Principal Component ◽  
Cell Heterogeneity ◽  
Indispensable Tool

Extracellular vesicles (EVs) are important intercellular mediators regulating health and diseases. Conventional methods for EV surface marker profiling, which was based on population measurements, masked the cell-to-cell heterogeneity in the quantity and phenotypes of EV secretion. Herein, by using spatially patterned antibody barcodes, we realized multiplexed profiling of single-cell EV secretion from more than 1,000 single cells simultaneously. Applying this platform to profile human oral squamous cell carcinoma (OSCC) cell lines led to a deep understanding of previously undifferentiated single-cell heterogeneity underlying EV secretion. Notably, we observed that the decrement of certain EV phenotypes (e.g.,CD63+EV) was associated with the invasive feature of both OSCC cell lines and primary OSCC cells. We also realized multiplexed detection of EV secretion and cytokines secretion simultaneously from the same single cells to investigate the multidimensional spectrum of cellular communications, from which we resolved tiered functional subgroups with distinct secretion profiles by visualized clustering and principal component analysis. In particular, we found that different cell subgroups dominated EV secretion and cytokine secretion. The technology introduced here enables a comprehensive evaluation of EV secretion heterogeneity at single-cell level, which may become an indispensable tool to complement current single-cell analysis and EV research.

scholarly journals Direct observation of frequency modulated transcription in single cells using light activation

eLife ◽  
2013 ◽  
Vol 2 ◽  
Author(s):  
Daniel R Larson ◽  
Christoph Fritzsch ◽  
Liang Sun ◽  
Xiuhau Meng ◽  
David S Lawrence ◽  
...  
Keyword(s):  
Single Cell ◽  
Single Molecule ◽  
Single Cell Analysis ◽  
Cell Model ◽  
Single Cells ◽  
Single Gene ◽  
Gene Activation ◽  
Light Activation ◽  
Dynamic Synthesis ◽  
Single Cell Model

Single-cell analysis has revealed that transcription is dynamic and stochastic, but tools are lacking that can determine the mechanism operating at a single gene. Here we utilize single-molecule observations of RNA in fixed and living cells to develop a single-cell model of steroid-receptor mediated gene activation. We determine that steroids drive mRNA synthesis by frequency modulation of transcription. This digital behavior in single cells gives rise to the well-known analog dose response across the population. To test this model, we developed a light-activation technology to turn on a single steroid-responsive gene and follow dynamic synthesis of RNA from the activated locus.

scholarly journals More than efficacy revealed by single-cell analysis of antiviral therapeutics

2019 ◽  
Author(s):  
Wu Liu ◽  
Mehmet U. Caglar ◽  
Zhangming Mao ◽  
Andrew Woodman ◽  
Jamie J. Arnold ◽  
...  
Keyword(s):  
Single Cell ◽  
Drug Targets ◽  
Single Cell Analysis ◽  
Single Cells ◽  
Principal Component ◽  
Viral Population ◽  
Cell Analysis ◽  
Toxic Dose ◽  
Time Required

SUMMARYDevelopment of antiviral therapeutics emphasizes minimization of the effective dose and maximization of the toxic dose, first in cell culture and later in animal models. Long-term success of an antiviral therapeutic is determined not only by its efficacy but also by the duration of time required for drug-resistance to evolve. We have developed a microfluidic device comprised of ~6000 wells, with each well containing a microstructure to capture single cells. We have used this device to characterize enterovirus inhibitors with distinct mechanisms of action. In contrast to population methods, single-cell analysis reveals that each class of inhibitor interferes with the viral infection cycle in a manner that can be distinguished by principal component analysis. Single-cell analysis of antiviral candidates reveals not only efficacy but also properties of the members of the viral population most sensitive to the drug, the stage of the lifecycle most affected by the drug, and perhaps even if the drug targets an interaction of the virus with its host.

scholarly journals μCB-seq: Microfluidic cell barcoding and sequencing for high-resolution imaging and sequencing of single cells

2020 ◽  
Author(s):  
Tyler N. Chen ◽  
Anushka Gupta ◽  
Mansi Zalavadia ◽  
Aaron M. Streets
Keyword(s):  
High Resolution ◽  
Single Cell ◽  
Single Cell Analysis ◽  
Protein Localization ◽  
Single Cells ◽  
Optical Measurements ◽  
Detection Sensitivity ◽  
High Resolution Imaging ◽  
Cell Analysis ◽  
Resolution Imaging

AbstractSingle-cell RNA sequencing (scRNA-seq) enables the investigation of complex biological processes in multicellular organisms with high resolution. However, many phenotypic features that are critical to understanding the functional role of cells in a heterogeneous tissue or organ are not directly encoded in the genome and therefore cannot be profiled with scRNA-seq. Quantitative optical microscopy has long been a powerful approach for characterizing diverse cellular phenotypes including cell morphology, protein localization, and chemical composition. Combining scRNA-seq with optical imaging has the potential to provide comprehensive single-cell analysis, allowing for functional integration of gene expression profiling and cell-state characterization. However, it is difficult to track single cells through both measurements; therefore, coupling current scRNA-seq protocols with optical measurements remains a challenge. Here, we report Microfluidic Cell Barcoding and Sequencing (μCB-seq), a microfluidic platform that combines high-resolution imaging and sequencing of single cells. μCB-seq is enabled by a novel fabrication method that preloads primers with known barcode sequences inside addressable reaction chambers of a microfluidic device. In addition to enabling multi-modal single-cell analysis, μCB-seq improves gene detection sensitivity, providing a scalable and accurate method for information-rich characterization of single cells.

scholarly journals MBRS-59. SINGLE-CELL WHOLE-GENOME SEQUENCING DISSECTS INTRA-TUMOURAL GENOMIC HETEROGENEITY AND CLONAL EVOLUTION IN CHILDHOOD MEDULLOBLASTOMA

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii408-iii408
Author(s):  
Marina Danilenko ◽  
Masood Zaka ◽  
Claire Keeling ◽  
Stephen Crosier ◽  
Rafiqul Hussain ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Single Cell ◽  
Genome Sequencing ◽  
Copy Number ◽  
Single Cell Analysis ◽  
Mutational Analysis ◽  
Single Cells ◽  
Clonal Evolution ◽  
Whole Genome ◽  
Clinically Significant

Abstract Medulloblastomas harbor clinically-significant intra-tumoral heterogeneity for key biomarkers (e.g. MYC/MYCN, β-catenin). Recent studies have characterized transcriptional heterogeneity at the single-cell level, however the underlying genomic copy number and mutational architecture remains to be resolved. We therefore sought to establish the intra-tumoural genomic heterogeneity of medulloblastoma at single-cell resolution. Copy number patterns were dissected by whole-genome sequencing in 1024 single cells isolated from multiple distinct tumour regions within 16 snap-frozen medulloblastomas, representing the major molecular subgroups (WNT, SHH, Group3, Group4) and genotypes (i.e. MYC amplification, TP53 mutation). Common copy number driver and subclonal events were identified, providing clear evidence of copy number evolution in medulloblastoma development. Moreover, subclonal whole-arm and focal copy number alterations covering important genomic loci (e.g. on chr10 of SHH patients) were detected in single tumour cells, yet undetectable at the bulk-tumor level. Spatial copy number heterogeneity was also common, with differences between clonal and subclonal events detected in distinct regions of individual tumours. Mutational analysis of the cells allowed dissection of spatial and clonal heterogeneity patterns for key medulloblastoma mutations (e.g. CTNNB1, TP53, SMARCA4, PTCH1) within our cohort. Integrated copy number and mutational analysis is underway to establish their inter-relationships and relative contributions to clonal evolution during tumourigenesis. In summary, single-cell analysis has enabled the resolution of common mutational and copy number drivers, alongside sub-clonal events and distinct patterns of clonal and spatial evolution, in medulloblastoma development. We anticipate these findings will provide a critical foundation for future improved biomarker selection, and the development of targeted therapies.

scholarly journals Methods and sensors for functional genomic studies of cell-cycle transitions in single cells

2020 ◽  
Vol 52 (10) ◽  
pp. 468-477
Author(s):  
Alexander C. Zambon ◽  
Tom Hsu ◽  
Seunghee Erin Kim ◽  
Miranda Klinck ◽  
Jennifer Stowe ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Single Cell ◽  
Single Cell Analysis ◽  
Imaging System ◽  
Single Cells ◽  
Cell Types ◽  
Cell Analysis ◽  
Mcf7 Cells ◽  
Genomic Studies ◽  
Cell Subpopulations

Much of our understanding of the regulatory mechanisms governing the cell cycle in mammals has relied heavily on methods that measure the aggregate state of a population of cells. While instrumental in shaping our current understanding of cell proliferation, these approaches mask the genetic signatures of rare subpopulations such as quiescent (G0) and very slowly dividing (SD) cells. Results described in this study and those of others using single-cell analysis reveal that even in clonally derived immortalized cancer cells, ∼1–5% of cells can exhibit G0 and SD phenotypes. Therefore to enable the study of these rare cell phenotypes we established an integrated molecular, computational, and imaging approach to track, isolate, and genetically perturb single cells as they proliferate. A genetically encoded cell-cycle reporter (K67p-FUCCI) was used to track single cells as they traversed the cell cycle. A set of R-scripts were written to quantify K67p-FUCCI over time. To enable the further study G0 and SD phenotypes, we retrofitted a live cell imaging system with a micromanipulator to enable single-cell targeting for functional validation studies. Single-cell analysis revealed HT1080 and MCF7 cells had a doubling time of ∼24 and ∼48 h, respectively, with high duration variability in G1 and G2 phases. Direct single-cell microinjection of mRNA encoding (GFP) achieves detectable GFP fluorescence within ∼5 h in both cell types. These findings coupled with the possibility of targeting several hundreds of single cells improves throughput and sensitivity over conventional methods to study rare cell subpopulations.

scholarly journals Synchrotron macro ATR-FTIR microspectroscopy for high-resolution chemical mapping of single cells

The Analyst ◽  
2019 ◽  
Vol 144 (10) ◽  
pp. 3226-3238 ◽  
Author(s):  
Jitraporn Vongsvivut ◽  
David Pérez-Guaita ◽  
Bayden R. Wood ◽  
Philip Heraud ◽  
Karina Khambatta ◽  
...  
Keyword(s):  
High Resolution ◽  
Single Cell ◽  
Spatial Resolution ◽  
Environmental Science ◽  
Single Cell Analysis ◽  
Single Cells ◽  
Cell Analysis ◽  
Chemical Mapping ◽  
Ftir Microspectroscopy ◽  
Resolution Single

Coupling synchrotron IR beam to an ATR element enhances spatial resolution suited for high-resolution single cell analysis in biology, medicine and environmental science.

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