Microscope slide | EPFL Graph Search

A microscope slide is a thin flat piece of glass, typically 75 by 26 mm (3 by 1 inches) and about 1 mm thick, used to hold objects for examination under a microscope. Typically the object is mounted (secured) on the slide, and then both are inserted together in the microscope for viewing. This arrangement allows several slide-mounted objects to be quickly inserted and removed from the microscope, labeled, transported, and stored in appropriate slide cases or folders etc. Microscope slides are often used together with a cover slip or cover glass, a smaller and thinner sheet of glass that is placed over the specimen. Slides are held in place on the microscope's stage by slide clips, slide clamps or a cross-table which is used to achieve precise, remote movement of the slide upon the microscope's stage (such as in an automated/computer operated system, or where touching the slide with fingers is inappropriate either due to the risk of contamination or lack of precision).

Real-time monitoring of single-cell secretion with a high-throughput nanoplasmonic microarray

Hatice Altug, Saeid Ansaryan, Xiaokang Li, Yen-Cheng Liu, Eduardo Romero Arvelo

Proteins secreted by cells play significant roles in mediating many physiological, developmental, and pathological processes due to their functions in intra/intercellular communication and signaling. Conventional end-point methods are insufficient for understanding the temporal response in cell secretion process, which is often highly dynamic. Furthermore, cellular heterogeneity makes it essential to analyze secretory proteins from single cells. To uncover individual cellular activities and the underlying kinetics, new technologies are needed for real-time analysis of the secretomes of many cells at single-cell resolution. This study reports a high-throughput biosensing microarray platform, which is capable of label-free and real-time secretome monitoring from a large number of living single cells using a biochip integrating ultrasensitive nanoplasmonic substrate and microwell compartments having volumes of ∼0.4 nL. Precise synchronization of image acquisition and microscope stage movement of the developed optical platform enables spectroscopic analysis with high temporal and spectral resolution. In addition, our system allows simultaneous optical imaging of cells to track morphology changes for a comprehensive understanding of cellular behavior. We demonstrated the platform performance by detecting interleukin-2 secretion from hundreds of single lymphoma cells in real-time over many hours. Significantly, the analysis of the secretion kinetics allows us to study cellular response to the stimulations in a statistical way. The new platform is a promising tool for the characterization of single-cell functionalities given its versatility, throughput and label-free configuration.

2022

Improvement of breast cancer diagnosis through microfluidics

Huu Tuan Nguyen

In breast cancer, 15-20% of cases are reported with overexpression of human epidermal growth factor receptor 2 (HER2) that causes rapid cancer progression and poor prognosis. Fortunately, HER2-targeted therapy using specific antibodies such as Trastuzumab is effective for treating these cases. In situ hybridization (ISH) is a standard technique used for HER2 assessment. This technique locates the HER2 gene by using complementary DNA probes. Two main ISH methods are fluorescence in situ hybridization (FISH) and chromogenic in situ hybridization (CISH). However, both FISH and CISH are expensive and laborious. Moreover, they can only be assessed in a small area of the tissue and thus are prone to errors in case of HER2 intra-tumoral heterogeneity (ITH). This thesis aims to improve HER2 assessment in breast cancer through various techniques related to microfluidics. The main component of our technology is a microfluidic tissue processor. This micro-fabricated chip is clamped to a microscope slide carrying a human tissue slice, creating a chamber that accommodates the tissue and delivering reagents to stain it. Five applications of this microfluidic technology to breast cancer diagnostics are presented in the three chapters of this thesis. In chapter 1, we present a microfluidic FISH protocol for HER2 gene assessment using a standard FISH probe and demonstrate that, when applying an oscillatory flow within the chip, hybridization efficiency is increased thanks to molecular replenishment. This back-and-forth motion of the diluted FISH probe inside a thin chamber above the tissue slide is the principle of microfluidic-assistance for ISH. We thereby succeed in drastically reducing the experimental time from 2 days to 1, and the amount of the expensive probe used per test by a factor of 10. We highlight the performance and reliability of microfluidic-assistance FISH by comparing the FISH scores obtained by this method to standard FISH technique scores using several clinical tissue samples. The principle of microfluidic assistance in FISH is also applicable to other types of ISH probes, including fast FISH based on Ethylene Carbonate and CISH. In chapter 2, we describe a new microfluidic method allowing the quantification of HER2 expression levels from formalin-fixed breast cancer tissues. After partial extraction of proteins from the tissue slide, the extract is routed to an antibody microarray for HER2 titration by fluorescence. HER2-negative and positive samples can be distinguished using this simple test, and the obtained results agree with the FISH scores. In chapter 3, we establish a method allowing high content, cell-by-cell analysis of both protein overexpression and gene amplification using successive microfluidic immunofluorescence (IF) and FISH staining combined with image processing. We demonstrate that by using high-content automatic analysis, the HER2 status of the sample can be precisely assessed using both a quantitative IF technique based on HER2 and cytokeratin protein quantification and automatic scoring of HER2 loci and centromere of chromosome 17 signals in a FISH image. Furthermore, this method characterizes HER2 ITH quantitatively. In particular, heterogeneous clusters and individual cells are visualized in a reconstructed map of the tissue. We conclude that high-content IF/FISH analysis is a powerful tool that can assist clinical diagnostics in the future.

EPFL2018

Extra short incubation microfluidic assisted – fluorescence in situ hybridization (ESIMA-FISH)

Estelle Annick Cuttaz, Laurène Nathalie Dupont, Martinus Gijs, Huu Tuan Nguyen, Raphaël Etienne Jean Trouillon

Fluorescence in situ hybridization (FISH) is a powerful technique for evaluating the HER2 gene status in breast cancer specimen (1). However, most of FISH assays currently used in clinical laboratories are expensive and require long experimental time, up to two days with an overnight incubation (2). Indeed, despite the development of faster FISH probes (HER2 IQFISH pharmDxTM from DAKO, Denmark) cutting the assay time to one day (3), the cost of these new probes is still high (more than 200$/test) and impedes the dissemination of this technique. In this study, we present the extra short incubation microfluidic assisted- fluorescence in situ hybridization (ESIMA-FISH) technique that uses microfluidics to improve FISH for HER2 assessment in breast cancer samples. ESIMA-FISH requires a very short incubation time (35 minutes) and uses 4-fold less probe solution per test. The system is based on a microfluidic chip, developed in our laboratory (4), that is clamped against a microscope slide containing a breast cancer tissue specimen (figure 1). A fluorescent DNA probe solution, specific to the target DNA, is then applied to the tissue section within a thin chamber using the microfluidic system. The probe solution used is obtained by diluting 4 times the standard HER2 IQFISH pharmDxTM probe solution (DAKO, Denmark). Oscillating flows can then be implemented using syringe pumps to improve the delivery of the probe to the tissue. Thanks to this hydrodynamic enhancement of mass transport, the probe-target hybridization efficiency is increased, resulting in overall reductions in the cost and duration of the assay. To validate the ESIMA-FISH technique, several tissue specimens were blindly tested with ESIMA-FISH and standard IQFISH. The results from these two techniques were comparable (figure 2, 3), supporting the possibility of a future clinical use of ESIMA-FISH.

2016