وأيد هذا العمل من قبل منحة ماري كوري لإعادة الإدماج.

<ol>
	<li>Maerkl, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Integration+column:+Microfluidic+high-throughput+screening.">Integration column: Microfluidic high-throughput screening.</a> Integrative biology quantitative biosciences from nano to macro. 1, 19-29 (2009).</li><li>Hong, J. W., Quake, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Integrated+nanoliter+systems.">Integrated nanoliter systems.</a> Nature. 21, 1179-1183 (2003).</li><li>Unger, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Monolithic+Microfabricated+Valves+and+Pumps+by+Multilayer+Soft+Lithography.">A Monolithic Microfabricated Valves and Pumps by Multilayer Soft Lithography.</a> Science. 288, 113-116 (2000).</li><li>Gerber, D., Maerkl, S. J., Quake, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+in+vitro+microfluidic+approach+to+generating+protein-interaction+networks.">An in vitro microfluidic approach to generating protein-interaction networks.</a> Nature. 6, 71-74 (2009).</li><li>Einav, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Discovery+of+a+hepatitis+C+target+and+its+pharmacological+inhibitors+by+microfluidic+affinity+analysis.">Discovery of a hepatitis C target and its pharmacological inhibitors by microfluidic affinity analysis.</a> Nature. 26, 1019-1027 (2008).</li><li>Fordyce, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=De+novo+identification+and+biophysical+characterization+of+transcription-factor+binding+sites+with+microfluidic+affinity+analysis.">De novo identification and biophysical characterization of transcription-factor binding sites with microfluidic affinity analysis.</a> Nature Biotechnology. 28, 962-967 (2010).</li><li>Zhu, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Global+analysis+of+protein+activities+using+proteome+chips.">Global analysis of protein activities using proteome chips.</a> Science (New York, N.Y.). 293, 2101-2105 (2001).</li><li>Ramachandran, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Self-assembling+protein+microarrays.">Self-assembling protein microarrays.</a> Science (New York, N.Y.). 305, 86-90 (2004).</li><li>Zhong, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+microfluidic+processor+for+gene+expression+profiling+of+single+human+embryonic+stem+cells.">A microfluidic processor for gene expression profiling of single human embryonic stem cells.</a> Lab on a chip. 8, 68-74 (2008).</li><li>Kusnezow, W., Hoheisel, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Solid+supports+for+microarray+immunoassays.">Solid supports for microarray immunoassays.</a> Journal of molecular recognition JMR. 16, 165-176 (2003).</li><li>Lundin, M., Monne, M., Widell, A., Von Heijne, G., Persson, M. A. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Topology+of+the+membrane-associated+hepatitis+C+virus+protein+NS4B.">Topology of the membrane-associated hepatitis C virus protein NS4B.</a> Journal of virology. 77, 5428 (2003).</li></ol>

الإعلان عن أي تضارب في المصالح.

في هذه الورقة نقدم طريقة لتوليد البروتين صفائف في الإنتاجية العالية باستخدام منصة ميكروفلويديك. ويستند الجيل مجموعة على الطباعة من القوالب ميكروأري DNA والبروتين في التعبير عن DNA داخل جهاز ميكروفلويديك. برنامجنا ميكروفلويد?...

<table border="1" style=";text-align:right;direction:rtl"><tbody><tr><td> كاشف / معدات </td><td> شركة </td><td> كتالوج رقم </td></tr><tr><td> PDMS-184 SYLGARD </td><td> داو كورنينج USA </td><td> ESSEX-DC </td></tr><tr><td> Chlorotrimethylsilane (TMCS </td><td> سيغما الدريخ </td><td> C72854 </td></tr><tr><td> ركائز الزجاج المطلي الايبوكسي </td><td> CEL المنتسبين USA </td><td> VEPO-25C </td></tr><tr><td> البولي إثيلين glycole (PEG) </td><td> سيغما الدريخ </td><td> 81260 </td></tr><tr><td> D-طرهالوز تبلور </td><td> سيغما الدريخ </td><td> T9531 </td></tr><tr><td> المعقدة البيروكسيديز-BSA </td><td> خرق </td><td> PIR-29130 </td></tr><tr><td> Neutravidin </td><td> خرق </td><td> 31050 </td></tr><tr><td> خماسي صاحب-البيوتين </td><td> QIAGEN </TD&gt; <td> 34440 </td></tr><tr><td> HEPES </td><td> <a href="http://www.bioind.com/htmls/article.aspx?c0=741&amp;bsp=13384" target="_blank">صناعات البيولوجية</a> </td><td> 03-025-1B </td></tr><tr><td> TNT-T7 </td><td> Promega </td><td> L5540 </td></tr><tr><td> C-MYC Cy3 الأجسام المضادة </td><td> سيغما الدريخ </td><td></td></tr><tr><td> تحكم مربع </td><td> ستانفورد على microfluidics مسبك </td><td></td></tr><tr><td> قالب </td><td> ستانفورد على microfluidics مسبك </td><td></td></tr><tr><td> دبوس </td><td> نيو انغلاند الصغيرة أنابيب شركة </td><td></td></tr><tr><td> Tygon microbore أنابيب </td><td> Tygon </td><td> S-54-HL </td></tr><tr><td> Microarrayer </td><td> الروبوتات الحيوية </td><td> MicroGrid 610 </td></tr><tr><td> سيليكون دبابيس </td><td> بالتوازي SYNأطروحة </td><td> SMT-S75 </td></tr></tbody></table>

high-throughput protein expression generator using a microfluidic platform

الحقول يتزايد بسرعة، مثل علم الأحياء النظم، تتطلب وضع وتنفيذ تكنولوجيات جديدة، وتمكين قياسات عالية الإنتاجية وارتفاع الإخلاص، نظم كبيرة. على microfluidics وعود لتحقيق العديد من هذه المتطلبات، مثل إجراء التجارب الفرز الفائق الإنتاجية على الرقاقة، والتي تشمل فحوصات الكيمياء الحيوية، والفيزياء الحيوية، ومقرها الخلية-1. منذ الأيام الأولى من أجهزة على microfluidics، وقد تطورت بشكل كبير هذا المجال، مما يؤدي إلى تطوير التكامل على نطاق واسع ميكروفلويديك 2،3. هذه التكنولوجيا تسمح لإدماج الآلاف من الصمامات الميكرو ميكانيكية على جهاز واحد مع بصمة البريد الحجم (الشكل 1). قمنا بتطوير منصة عالية الإنتاجية لتوليد ميكروفلويديك في المختبر من صفائف التعبير البروتين (الشكل 2) اسمه PING (مولد الشبكة بروتين التفاعل). يمكن لهذه المصفوفات تخدم كنموذج لتجارب العديد منمثل البروتين البروتين 4، البروتين RNA 5 أو 6 DNA البروتين التفاعلات. الجهاز يتكون من آلاف من غرف رد فعل، والتي مبرمجة بشكل فردي باستخدام microarrayer. التوفيق بين هذه المطبوعة إلى ميكروأرس برامج الأجهزة على microfluidics كل غرفة مع بقعة واحدة القضاء على التلوث المحتملة أو عبر التفاعل-علاوة على ذلك، باستخدام تقنيات توليد ميكروأرس ميكروأري القياسية اكتشاف هو أيضا وحدات للغاية، مما يسمح للarraying من البروتينات 7، DNA 8، جزيئات صغيرة، والمعلقات حتى الغروية. الأثر المحتمل للعلوم البيولوجية على microfluidics على أمر مهم. وقد قدم عدد من فحوصات على microfluidics القائمة بالفعل رؤى جديدة في هيكل ووظيفة النظم البيولوجية، وميدان على microfluidics سوف تظل تؤثر البيولوجيا.

Watch this Scientific Journal Video about High-throughput Protein Expression Generator Using a Microfluidic Platform at JoVE.com

High-throughput Protein Expression Generator Using a Microfluidic Platform

نقدم نهج ميكروفلويديك للتعبير عن المصفوفات البروتين. الجهاز يتكون من آلاف من الدوائر التي تسيطر عليها رد فعل الصغيرة الميكانيكية الصمامات. تزاوج الجهاز ميكروفلويديك إلى مكتبة الجينات ميكروأري المطبوعة. وكتب بعد ذلك هذه الجينات وترجمتها على الرقاقة، مما أدى إلى مجموعة البروتين جاهزة للاستخدام التجريبي.

عالية الإنتاجية مولد بروتين تعبير باستخدام منصة ميكروفلويديك

Rapidly increasing fields, such as systems biology, require the development and implementation of new technologies, enabling high-throughput and ...

high-throughput-protein-expression-generator-using-microfluidic

Research

JoVE Journal

Bioengineering

11.6K Views.  Bar-Ilan University. نقدم نهج ميكروفلويديك للتعبير عن المصفوفات البروتين. الجهاز يتكون من آلاف من الدوائر التي تسيطر عليها رد فعل الصغيرة الميكانيكية الصمامات. تزاوج الجهاز ميكروفلويديك إلى مكتبة الجينات ميكروأري المطبوعة. وكتب بعد ذلك هذه الجينات وترجمتها على الرقاقة، مما أدى إل...

Video: High-throughput Protein Expression Generator Using a Microfluidic Platform

CometChip: A High-throughput 96-Well Platform for Measuring DNA Damage in Microarrayed Human Cells

DNA damaging agents can promote aging, disease and cancer and they are ubiquitous in the environment and produced within human cells as normal cellular metabolites. Ironically, at high doses DNA damaging agents are also used to treat cancer. The ability to quantify DNA damage responses is thus critical in the public health, pharmaceutical and clinical domains. Here, we describe a novel platform that exploits microfabrication techniques to pattern cells in a fixed microarray. The &#8216;CometChip&#8217; is based upon the well-established single cell gel electrophoresis assay (a.k.a. the comet assay), which estimates the level of DNA damage by evaluating the extent of DNA migration through a matrix in an electrical field. The type of damage measured by this assay includes abasic sites, crosslinks, and strand breaks. Instead of being randomly dispersed in agarose in the traditional assay, cells are captured into an agarose microwell array by gravity. The platform also expands from the size of a standard microscope slide to a 96-well format, enabling parallel processing. Here we describe the protocols of using the chip to evaluate DNA damage caused by known genotoxic agents and the cellular repair response followed after exposure. Through the integration of biological and engineering principles, this method potentiates robust and sensitive measurements of DNA damage in human cells and provides the necessary throughput for genotoxicity testing, drug development, epidemiological studies and clinical assays.

We describe here a platform that allows comet assay detection of DNA damage with unprecedented throughput. The device patterns mammalian cells into a microarray and enables parallel processing of 96 samples. The approach facilitates analysis of base level DNA damage, exposure-induced DNA damage and DNA repair kinetics.

CometChip: إنتاجية عالية منصة 96-حسنا لقياس الحمض النووي من التلف في خلايا الإنسان Microarrayed

DNA damaging agents can promote aging, disease and cancer and they are ubiquitous in the environment and produced within human cells as normal cellular ...

High Throughput Microfluidic Rapid and Low Cost Prototyping Packaging Methods

In this work, 3 different packaging and assembly techniques are presented. They can be classified into two categories: one-time use and reusable packaging techniques.
The one-time use packaging technique employs UV-based and temperature curing epoxies to connect microtubes to access holes, wire-bonding for integrated circuit connections, and silver epoxy for electrical connections. This method is based on a robust assembly technique that can support relatively high pressure close to 1 psi and does not need any support to strengthen the microfluidic architecture.
Reusable packaging techniques consist of PDMS-based microtube interconnectors and anisotropic adhesive films for electrical connections. These devices are more sensitive and fragile. Consequently, Plexiglas support is added to the microfluidic structure to improve the electrical contact when anisotropic adhesive films are used, and also to strengthen the microfluidic architecture. In addition, a micromanipulator is needed to maintain tubes while using a thin PDMS layer to connect them to the access holes. Different PDMS layer thicknesses, ranging from 0.45-3 mm, are tested to compare the best adherence versus injection rates. Applied injection rates are varied from 50-300 &#956;l/hr for 0.45-3 mm PDMS layers, respectively. These techniques are mainly applicable for low-pressure applications. However, they can be extended for high-pressure ones through plasma-oxygen process to permanently seal the PDMS to glass substrates. The main advantage of this technique, besides the fact that it is reusable, consists of keeping the device observable when the microchannel length is very short (in the range of 3 mm or lower).

In this article we describe different techniques for microfluidic rapid prototyping platforms. The proposed techniques are based on ultraviolet (UV) sensitive and temperature curing epoxies, polydimethylsiloxane (PDMS) based tubing, wire-bonding, and anisotropic adhesive films. The assembling procedures presented are developed for both one-time use devices as well as reusable microfluidic systems.

عالية الإنتاجية ميكروفلويديك السريع وانخفاض تكلفة النماذج طرق التعبئة والتغليف

In this work, 3 different packaging and assembly techniques are presented. They can be classified into two categories: one-time use and reusable packaging ...

Cell Squeezing as a Robust, Microfluidic Intracellular Delivery Platform

Rapid mechanical deformation of cells has emerged as a promising, vector-free method for intracellular delivery of macromolecules and nanomaterials. This technology has shown potential in addressing previously challenging applications; including, delivery to primary immune cells, cell reprogramming, carbon nanotube, and quantum dot delivery. This vector-free microfluidic platform relies on mechanical disruption of the cell membrane to facilitate cytosolic delivery of the target material. Herein, we describe the detailed method of use for these microfluidic devices including, device assembly, cell preparation, and system operation. This delivery approach requires a brief optimization of device type and operating conditions for previously unreported applications. The provided instructions are generalizable to most cell types and delivery materials as this system does not require specialized buffers or chemical modification/conjugation steps. This work also provides recommendations on how to improve device performance and trouble-shoot potential issues related to clogging, low delivery efficiencies, and cell viability.

Rapid mechanical deformation of cells has emerged as a promising, vector-free method for intracellular delivery of macromolecules and nanomaterials. This protocol provides detailed steps on how to use the system for a broad range of applications.

الضغط خلية باعتبارها قوية، ميكروفلويديك بين الخلايا توصيل منصة

Rapid mechanical deformation of cells has emerged as a promising, vector-free method for intracellular delivery of macromolecules and nanomaterials. This ...

Use of a High-throughput In Vitro Microfluidic System to Develop Oral Multi-species Biofilms

There are few high-throughput in vitro systems which facilitate the development of multi-species biofilms that contain numerous species commonly detected within in vivo oral biofilms. Furthermore, a system that uses natural human saliva as the nutrient source, instead of artificial media, is particularly desirable in order to support the expression of cellular and biofilm-specific properties that mimic the in vivo communities. We describe a method for the development of multi-species oral biofilms that are comparable, with respect to species composition, to supragingival dental plaque, under conditions similar to the human oral cavity. Specifically, this methods article will describe how a commercially available microfluidic system can be adapted to facilitate the development of multi-species oral biofilms derived from and grown within pooled saliva. Furthermore, a description of how the system can be used in conjunction with a confocal laser scanning microscope to generate 3-D biofilm reconstructions for architectural and viability analyses will be presented. Given the broad diversity of microorganisms that grow within biofilms in the microfluidic system (including Streptococcus, Neisseria, Veillonella, Gemella, and Porphyromonas), a protocol will also be presented describing how to harvest the biofilm cells for further subculture&#160;or DNA extraction and analysis. The limits of both the microfluidic biofilm system and the current state-of-the-art data analyses will be addressed. Ultimately, it is envisioned that this article will provide a baseline technique that will improve the study of oral biofilms and aid in the development of additional technologies that can be integrated with the microfluidic platform.

Use of a High-throughput In Vitro Microfluidic System to Develop Oral Multi-species Biofilms

The goal of this methods paper is to describe the use of a microfluidic system for the development of multi-species biofilms that contain species typically identified in human supragingival dental plaque.&#160;Methods to describe biofilm architecture, biofilm viability, and an approach to harvest biofilm for culture-dependent or culture-independent analyses are highlighted.

استخدام إنتاجية عالية<em&gt; في المختبر</em&gt; نظام ميكروفلويديك لتطوير الأغشية الحيوية متعدد الأنواع عن طريق الفم

There are few high-throughput in vitro systems which facilitate the development of multi-species biofilms that contain numerous species commonly detected ...

A Microfluidic Platform for High-throughput Single-cell Isolation and Culture

Studying the heterogeneity of single cells is crucial for many biological questions, but is technically difficult. Thus, there is a need for a simple, yet high-throughput, method to perform single-cell culture experiments. Here, we report a microfluidic chip-based strategy for high-efficiency single-cell isolation (~77%) and demonstrate its capability of performing long-term single-cell culture (up to 7 d) and cellular heterogeneity analysis using clonogenic assay. These applications were demonstrated with KT98 mouse neural stem cells, and A549 and MDA-MB-435 human cancer cells. High single-cell isolation efficiency and long-term culture capability are achieved by using different sizes of microwells on the top and bottom of the microfluidic channel. The small microwell array is designed for precisely isolating single-cells, and the large microwell array is used for single-cell clonal culture in the microfluidic chip. This microfluidic platform constitutes an attractive approach for single-cell culture applications, due to its flexibility of adjustable cell culture spaces for different culture strategies, without decreasing isolation efficiency.

Here, we present a protocol for isolating and culturing single cells with a microfluidic platform, which utilizes a new microwell design concept to allow for high-efficiency single cell isolation and long-term clonal culture.

ومنصة ميكروفلويديك لعزل وحيدة الخلية عالية الإنتاجية والثقافة

Studying the heterogeneity of single cells is crucial for many biological questions, but is technically difficult. Thus, there is a need for a simple, yet ...

A High-throughput Cell Microarray Platform for Correlative Analysis of Cell Differentiation and Traction Forces

Microfabricated cellular microarrays, which consist of contact-printed combinations of biomolecules on an elastic hydrogel surface, provide a tightly controlled, high-throughput engineered system for measuring the impact of arrayed biochemical signals on cell differentiation. Recent efforts using cell microarrays have demonstrated their utility for combinatorial studies in which many microenvironmental factors are presented in parallel. However, these efforts have focused primarily on investigating the effects of biochemical cues on cell responses. Here, we present a cell microarray platform with tunable material properties for evaluating both cell differentiation by immunofluorescence and biomechanical cell&#8211;substrate interactions by traction force microscopy. To do so, we have developed two different formats utilizing polyacrylamide hydrogels of varying Young&#39;s modulus fabricated on either microscope slides or glass-bottom Petri dishes. We provide best practices and troubleshooting for the fabrication of microarrays on these hydrogel substrates, the subsequent cell culture on microarrays, and the acquisition of data. This platform is well-suited for use in investigations of biological processes for which both biochemical (e.g., extracellular matrix composition) and biophysical (e.g., substrate stiffness) cues may play significant, intersecting roles.

Cell differentiation is regulated by a host of microenvironmental factors, including both matrix composition and substrate material properties. We describe here a technique utilizing cell microarrays in conjunction with traction force microscopy to evaluate both cell differentiation and biomechanical cell&#8211;substrate interactions as a function of microenvironmental context.

وإنتاجية عالية منصة ميكروأري الخليوي لتحليل مترابط من تمايز الخلايا وقوات الجر

Microfabricated cellular microarrays, which consist of contact-printed combinations of biomolecules on an elastic hydrogel surface, provide a tightly ...

A Microfluidic Platform for Longitudinal Imaging in Caenorhabditis elegans

In the last decade, microfluidic techniques have been applied to study small animals, including the nematode Caenorhabditis elegans, and have proved useful as a convenient live imaging platform providing capabilities for precise control of experimental conditions in real time. In this article, we demonstrate live imaging of individual worms employing WormSpa, a previously-published custom microfluidic device. In the device, multiple worms are individually confined to separate chambers, allowing multiplexed longitudinal surveillance of various biological processes. To illustrate the capability, we performed proof-of-principle experiments in which worms were infected in the device with pathogenic bacteria, and the dynamics of expression of immune response genes and egg laying were monitored continuously in individual animals. The simple design and operation of this device make it suitable for users with no previous experience with microfluidic-based experiments. We propose that this approach will be useful for many researchers interested in longitudinal observations of biological processes under well-defined conditions.

A Microfluidic Platform for Longitudinal Imaging in Caenorhabditis elegans

In this article, we demonstrate live imaging of individual worms employing a custom microfluidic device. In the device, multiple worms are individually confined to separate chambers, allowing multiplexed longitudinal surveillance of various biological processes.

منصة موائع جزيئية "التصوير الطولي" في ايليجانس كاينورهابديتيس

In the last decade, microfluidic techniques have been applied to study small animals, including the nematode Caenorhabditis elegans, and have proved ...

A Multilayer Microfluidic Platform for the Conduction of Prolonged Cell-Free Gene Expression

The limitations of cell-based synthetic biology are becoming increasingly apparent as researchers aim to develop larger and more complex synthetic genetic regulatory circuits. The analysis of synthetic genetic regulatory networks in vivo is time consuming and suffers from a lack of environmental control, with exogenous synthetic components interacting with host processes resulting in undesired behavior. To overcome these issues, cell-free characterization of novel circuitry is becoming more prevalent. In vitro transcription and translation (IVTT) mixtures allow the regulation of the experimental environment and can be optimized for each unique system. The protocols presented here detail the fabrication of a multilayer microfluidic device that can be utilized to sustain IVTT reactions for prolonged durations. In contrast to batch reactions, where resources are depleted over time and (by-) products accumulate, the use of microfluidic devices allows the replenishment of resources as well as the removal of reaction products. In this manner, the cellular environment is emulated by maintaining an out-of-equilibrium environment in which the dynamic behavior of gene circuits can be investigated over extended periods of time. To fully exploit the multilayer microfluidic device, hardware and software have been integrated to automate the IVTT reactions. By combining IVTT reactions with the microfluidic platform presented here, it becomes possible to comprehensively analyze complex network behaviors, furthering our understanding of the mechanisms that regulate cellular processes.

The fabrication process of a PDMS-based, multilayer, microfluidic device that allows in vitro transcription and translation (IVTT) reactions to be performed over prolonged periods is described. Furthermore, a comprehensive overview of the hardware and software required to automate and maintain these reactions for prolonged durations is provided.

منصة ميكروفلويدريك متعددة الطبقات لتوصيل التعبير الجيني الخالي من الخلايا لفترات طويلة

The limitations of cell-based synthetic biology are becoming increasingly apparent as researchers aim to develop larger and more complex synthetic genetic ...

A Microfluidic Platform for Stimulating Chondrocytes with Dynamic Compression

Mechanical stimuli are known to modulate biological functions of cells and tissues. Recent studies have suggested that compressive stress alters growth plate cartilage architecture and results in growth modulation of long bones of children. To determine the role of compressive stress in bone growth, we created a microfluidic device actuated by pneumatic pressure, to dynamically (or statically) compress growth plate chondrocytes embedded in alginate hydrogel cylinders. In this article, we describe detailed methods for fabricating and characterizing this device. The advantages of our protocol are: 1) Five different magnitudes of compressive stress can be generated on five technical replicates in a single platform, 2) It is easy to visualize cell morphology via a conventional light microscope, 3) Cells can be rapidly isolated from the device after compression to facilitate downstream assays, and 4) The platform can be applied to study mechanobiology of any cell type that can grow in hydrogels.

This article provides detailed methods for fabricating and characterizing a pneumatically actuating microfluidic device for chondrocyte compression.

منصة ميكروفلويديك لتحفيز الخلايا الغضروفية مع الضغط الديناميكي

Mechanical stimuli are known to modulate biological functions of cells and tissues. Recent studies have suggested that compressive stress alters growth ...

High-Throughput Identification of Resistance to Pseudomonas syringae pv. Tomato in Tomato using Seedling Flood Assay

Tomato is an agronomically important crop that can be infected by Pseudomonas syringae, a Gram-negative bacterium, resulting in bacterial speck disease. The tomato-P. syringae pv. tomato pathosystem is widely used to dissect the genetic basis of plant innate responses and disease resistance. While disease was successfully managed for many decades through the introduction of the Pto/Prf gene cluster from Solanum pimpinellifolium into cultivated tomato, race 1 strains of P. syringae have evolved to overcome resistance conferred by the Pto/Prf gene cluster and occur worldwide.
Wild tomato species are important reservoirs of natural diversity in pathogen recognition, because they evolved in diverse environments with different pathogen pressures. In typical screens for disease resistance in wild tomato, adult plants are used, which can limit the number of plants that can be screened due to their extended growth time and greater growth space requirements. We developed a method to screen 10-day-old tomato seedlings for resistance, which minimizes plant growth time and growth chamber space, allows a rapid turnover of plants, and allows large sample sizes to be tested. Seedling outcomes of survival or death can be treated as discrete phenotypes or on a resistance scale defined by amount of new growth in surviving seedlings after flooding. This method has been optimized to screen 10-day-old tomato seedlings for resistance to two P. syringae strains and can easily be adapted to other P. syringae strains.

High-Throughput Identification of Resistance to Pseudomonas syringae pv. Tomato in Tomato using Seedling Flood Assay

The seedling flood assay facilitates rapid screening of wild tomato accessions for the resistance to the Pseudomonas syringae bacterium. This assay, used in conjunction with the seedling bacterial growth assay, can assist in further characterizing the underlying resistance to the bacterium, and can be used to screen mapping populations to determine the genetic basis of resistance.

تعريف عالي الإنتاجية للمقاومة لـ Pseudomonas syringae pv. الطماطم في الطماطم باستخدام الشتلات الفيضانات أساي

Tomato is an agronomically important crop that can be infected by Pseudomonas syringae, a Gram-negative bacterium, resulting in bacterial speck disease. ...