Showing papers by "Tuhin Subhra Santra published in 2016"

Microfluidic Devices in Advanced Caenorhabditis elegans Research

Abstract: The study of model organisms is very important in view of their potential for application to human therapeutic uses. One such model organism is the nematode worm, Caenorhabditis elegans. As a nematode, C. elegans have ~65% similarity with human disease genes and, therefore, studies on C. elegans can be translated to human, as well as, C. elegans can be used in the study of different types of parasitic worms that infect other living organisms. In the past decade, many efforts have been undertaken to establish interdisciplinary research collaborations between biologists, physicists and engineers in order to develop microfluidic devices to study the biology of C. elegans. Microfluidic devices with the power to manipulate and detect bio-samples, regents or biomolecules in micro-scale environments can well fulfill the requirement to handle worms under proper laboratory conditions, thereby significantly increasing research productivity and knowledge. The recent development of different kinds of microfluidic devices with ultra-high throughput platforms has enabled researchers to carry out worm population studies. Microfluidic devices primarily comprises of chambers, channels and valves, wherein worms can be cultured, immobilized, imaged, etc. Microfluidic devices have been adapted to study various worm behaviors, including that deepen our understanding of neuromuscular connectivity and functions. This review will provide a clear account of the vital involvement of microfluidic devices in worm biology.

Dielectric passivation layer as a substratum on localized single-cell electroporation

Abstract: Single-cell electroporation is a powerful technique to understand cellular behavior with heterogeneity, which might be impossible based on bulk measurements of millions of cells together. In this study, a dielectric passivation layer was deposited on top of an indium-tin oxide micro-electrode-based transparent chip surface using a plasma enhanced chemical vapour deposition technique. We theoretically and experimentally investigated the key effects of the dielectric passivation layer on localized single-cell electroporation for different cancer cells, which were randomly distributed with a high density throughout the chip surface as a monolayer. The passivation layer not only prevented the conventional or bulk electroporation with bubble and ion generation, but also provide an intense electric field in-between electrode gap for localized single-cell electroporation with high cell viability. Thus, devices with dielectric passivation layers are potentially applicable for single-cell studies.

Electroporation for Single-Cell Analysis

Abstract: Single-cell analysis is a powerful technique to understand cell to cell or cell to environment behaviors. It can provide detailed information of cell proliferation, differentiation, and different responses to external stimuli and intracellular reactions. For single cell analysis, electroporation or electropermeabilization is an efficient and fast method, where high external electric field is applied on cell membrane to form transient membrane pores to deliver ions or molecules in or out of the cell. Conventional electroporation or bulk electroporation (BEP) is performed in a batch mode with millions of cells in suspension, which can only provide an average value. Since the last decade, microfabricated devices have been developed to perform single cell electroporation, where electric field is only intense on a single cell, resulting in high transfection efficiency with high cell viability compared to BEP. Single cell analysis using electroporation technique can be performed with microfabricated electrode arrays, carbon fiber microelectrodes, micropipettes or electrolyte-filled capillaries-based devices. Recently developed nanofabricated electrodes can realize localized single cell electroporation for delivery of different molecules with high transfection efficiency and high cell viability. Thus, single cell electroporation technique opens up the new window for the manipulation of genomics, proteomics, trascriptomics, metabolomics, or fluxomics at single cell level. This chapter emphasizes the recent advancement of electroporation technique for cellular delivery and the analysis method which might be potentially applicable for biological research and therapeutic applications.

Microinjection for Single-Cell Analysis

Abstract: The mere existence of life, from unicellular organisms to well-organized multicellular organisms, pathological conditions and death, has always fascinated human beings and demanded understanding of biological systems over several centuries. Further, an efficient treatment strategy for genetic disorders requires understanding of pathological conditions at the single-cell level. Numerous experimental methodologies have been developed over several decades to facilitate our understanding of cellular functions, by modulating the molecular pathways. Needle microinjection is one of them and it is widely used to modulate cellular functions by introducing foreign cargo into the cell. Microinjection is a method that can directly deliver a precise amount of foreign cargo either into the cytoplasm or the nucleus of a single cell using micropipettes. It is considered a gold standard method of direct cargo delivery. After introduction of this technique in the early 1900s, numerous modifications were made to improve its efficiency and it was applied to a wide variety of fields from scientific research to clinical therapy. This chapter is intended to provide a basic knowledge of the microinjection technique, its advantages and disadvantages, its development, the basic instrumentation required along with a basic protocol, and its uses collected extensively from numerous literatures up-to-date. Further, several modifications that have been carried out to improve the basic instrumentation setup, in order to increase the efficiency and rate of microinjection by the addition of semi-automated and automated computerized systems, are also discussed. Finally, this chapter provides a gateway to explore advanced understanding of microinjection for single-cell analysis.

Photothermal nanoblades for delivery of large-sized cargo into mammalian cells at high throughput

Abstract: Technologies for transferring large-sized cargo into mammalian cells are needed to advance key applications in cell engineering. However, reliable methodologies for introducing large-sized cargo into mammalian cells are nearly completely lacking. This talk will present two new technologies, photothermal nanoblade and BLAST, that overcome the size limitation of cargo delivery into mammalian cells.