המחברים מבקשים להודות לויד Ung עבור קלט קונסטרוקטיבי בגרסאות המוקדמות של טכניקה זו, ד&quot;ר ג&#39;רמי Agresti לטיפים לעיצוב כובע לחץ, וד&quot;ר Dongping צ&#39;י על עזרתו בבודת כובע הלחץ. אנו מודים למעבדות של מ &#39;Teitell ופ Gunaratne למתן מגוון רחב של דגימות תאים לבדיקה. אנו מודים לקרן הלאומית למדע (קריירת פרס DBI-1,254,185), המרכז לסרטן באוניברסיטת UCLA Jonsson, וUCLA הקליני וTranslational Science Institute לתמיכה בעבודה זו.

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אי לנו ניגודי עניינים לחשוף.

<html> כאן אנו מספקים הליך ניסיוני מקיף לניתוח העיוות של תאים העוברים דרך ערוצי microfluidic מכווצים באמצעות זרימה המונע על ידי לחץ. תסריט MATLAB מאפשר עיבוד נתונים אוטומטי (משלימה חומר); גרסה מעודכנת של הקוד נשמר <a href="www.ibp.ucla.edu/research/rowat" target="_blank">(www.ibp.ucla.edu/research/rowat).</a> באופן רחב יות...

שינויים בצורת תא הם קריטיים בהקשרים ביולוגיים רבים. לדוגמא, אריתרוציטים ולויקוציטים לעוות דרך נימים שהם קטנים יותר מאשר הקוטר שלהם 1. בגרורות, תאים סרטניים חייבים לעוות דרך פערים צרים ביניים, כמו גם כלי דם מפותלים ורשתות הלימפה זרע באתרים משניים 2. כדי לחקור את ההתנהגות הפיסית של תאים בודדים, מכשירי microfluidic להציג פלטפורמה אידיאלית שיכול להיות מותאם אישית כדי ללמוד מגוון רחב של התנהגויות תאים הכוללים את היכולת שלהם להעביר דרך צרים פערים 3 ולעוות באופן פסיבי באמצעות אילוצי מיקרון בקנה מידה 3 9. Polydimethylsiloxane (PDMS) מכשירי microfluidic הם שקופים אופטי, המאפשרים לעיוותי תא להיות דמיינו באמצעות מיקרוסקופ אור ונותחו באמצעות כלים לעיבוד תמונה בסיסיים. יתר על כן, מערכים של אילוצים יכולים להיות מוגדרים במדויק, המאפשרים ניתוח של תאים מרובים בו זמנית עםתפוקה שעולה על טכניקות קיימות רבות 10,11. כאן אנו מציגים פרוטוקול ניסוי מפורט לבדיקת deformability תא באמצעות מכשיר microfluidic PDMS &#39;תא deformer&#39;. המכשיר תוכנן כך שמעבר תאים דרך אילוצים רציפים; גיאומטריה זו היא נפוצה בהקשרים פיסיולוגיים, כגון נימי ריאתי למיטה 12. כדי לאמוד את deformability תא, זמן מעבר מספק מטרי נוחה שנמדד בקלות את הזמן הנדרש לתא יחיד למעבר דרך 4,6 התכווצות אחת. כדי לשמור על ירידה בלחץ קבועה על פני הערוצים המכווצים בזמן מעבר תא, אנו משתמשים בזרימה מונע לחץ. הפרוטוקול שלנו כולל הוראות מפורטות על עיצוב מכשיר וייצור, תפעול מכשיר על ידי-לחץ מונע זרימה, הכנה והדמיה של תאים, כמו גם עיבוד תמונה כדי למדוד את הזמן לתאים כדי לעוות באמצעות סדרה של התכווצויות. אנו כולליםשני עיצובי מכשיר וקוד עיבוד נתונים חזון כקבצים משלימים. כמדגם מייצג של נתונים, אנו מראים את הזמן מעבר תא דרך סדרה של התכווצויות כפונקציה של מספר האילוצים passaged. ניתוח של לוח הזמנים לתאים למעבר למרות אילוצים צרים של מכשיר microfluidic יכול לחשוף הבדלים בdeformability של מגוון רחב של סוגי תאים 4,5,13. המכשיר הפגין כאן באופן ייחודי סוקר מעבר תא דרך סדרה של התכווצויות מיקרון בקנה מידה; עיצוב זה מחקה את הדרך המפותלת שתאים חווים במחזור וגם מאפשרים חיטוט מאפיינים פיזיים נוספים של התאים כגון זמן הרפיה.

<table border="0" cellpadding="0" cellspacing="0" width="573">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Name of Material/ Equipment</td>
			<td style="width:71px;">Company</td>
			<td style="width:119px;">Catalog Number</td>
			<td style="width:167px;">Comments/Description</td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">Pluronic F-127 Block Copolymer Surfactant&#160;</td>
			<td style="width:71px;">Fisher Scientific&#160;</td>
			<td style="width:119px;">8409400</td>
			<td style="width:167px;">Produced by BASF, also available through Sigma</td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">PDMS base and crosslinker</td>
			<td style="width:71px;">Essex Brownell</td>
			<td style="width:119px;">DC-184-1.1</td>
			<td style="width:167px;">Product commonly named Sylgard 184 Elastomer</td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">Oxygen plasma discharge unit</td>
			<td style="width:71px;">Enercon</td>
			<td style="width:119px;">Dyne-A-Mite 3D Treater</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">Biopsy Punch, Harris Uni-Core (0.75 mm)</td>
			<td style="width:71px;">Ted Pella, Inc.</td>
			<td style="width:119px;">15072</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">Fingertight Ferrule, 1/32&#34;</td>
			<td style="width:71px;">Upchurch Scientific</td>
			<td style="width:119px;">UP-F-113</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">Fingertight III Fitting, 10-32</td>
			<td style="width:71px;">Upchurch Scientific</td>
			<td style="width:119px;">UP-F-300X</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">polyetheretherketone (PEEK) tubing, outer diameter = 1/32&#34;or 0.79 mm</td>
			<td style="width:71px;">Valco</td>
			<td style="width:119px;">TPK.515-25M</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">polyethylene (PE-20) tubing, 0.043&#34; or 1.09 mm</td>
			<td style="width:71px;">Becton Dickinson</td>
			<td style="width:119px;">427406</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">Pressure regulator</td>
			<td style="width:71px;">Airgas or Praxair</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">Polyurethane tubing, 5/32&#8221; OD</td>
			<td style="width:71px;">McMaster Carr</td>
			<td style="width:119px;">5648K284</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">Push-to-connect fittings</td>
			<td style="width:71px;">McMaster Carr</td>
			<td style="width:119px;">5111K91</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:216px;">Voltage to Pressure (E/P) Electropneumatic Converter</td>
			<td style="width:71px;">Omega</td>
			<td style="width:119px;">IP413-020</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="52">
			<td height="52" style="height:52px;width:216px;">16-bit,250 kS/S, 80 Analog Inputs Multifunction DAQ</td>
			<td style="width:71px;">National Instruments</td>
			<td style="width:119px;">NI PCI 6225-779295-01</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="52">
			<td height="52" style="height:52px;width:216px;">Analog Connector Block-Screw Terminal</td>
			<td style="width:71px;">National Instruments</td>
			<td style="width:119px;">SCB-68-776844-01</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="52">
			<td height="52" style="height:52px;width:216px;">LabView System Design Software</td>
			<td style="width:71px;">National Instruments</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="52">
			<td height="52" style="height:52px;width:216px;">Matlab Software</td>
			<td style="width:71px;">The MathWorks, Inc.</td>
			<td style="width:119px;">Matlab R2012a</td>
			<td style="width:167px;">Code requires the Image Processing Toolbox</td>
		</tr>
		<tr height="53">
			<td height="53" style="height:53px;width:216px;">Shielded Cable</td>
			<td style="width:71px;">National Instruments</td>
			<td style="width:119px;">SHC68-68</td>
			<td style="width:167px;"></td>
		</tr>
	</tbody>
</table>

a microfluidic technique to probe cell deformability

כאן אנו פירוט עיצוב, ייצור, ושימוש במכשיר microfluidic להעריך deformability של מספר רב של תאים בודדים באופן יעיל. בדרך כלל, נתונים ל~ 10 2 תאים ניתן לרכוש בתוך ניסוי שעה 1. תכנית ניתוח תמונה אוטומטית מאפשרת ניתוח שלאחר ניסוי יעיל של נתוני תמונה, המאפשר עיבוד להיות שלם בתוך כמה שעות. הגיאומטריה המכשיר שלנו היא ייחודית בכך שתאים חייבים לעוות באמצעות סדרה של התכווצויות מיקרון בקנה מידה, ובכך לאפשר את העיוות הראשונית והרפיה של תאים בודדים תלוי זמן להיות assayed. תחולתה של שיטה זו לוקמיה פרומיילוציטית (HL-60) התאים באה לידי ביטוי. נהיגה תאים לעוות דרך אילוצי מיקרון בקנה מידה באמצעות זרימה המונע על ידי לחץ, אנו צופים כי פרומיילוציטית האנושית (HL-60) תאים לרגע לחסום את ההתכווצות הראשונה לזמן החציוני של 9.3 אלפיות שניים לפני passaging במהירות רב יותר דרך להצר הבאיונים עם זמן מעבר החציוני של 4.0 אלפיות שניים להתכווצות. בניגוד לכך, (נויטרופילים סוג) שטופל בחומצה רטינואית כל טרנס HL-60 תאים לחסום את ההתכווצות הראשונה רק 4.3 אלפית שניים לפני passaging דרך האילוצים שלאחר מכן עם זמן מעבר החציוני של 3.3 אלפיות שניים. שיטה זו יכולה לספק תובנות לגבי טבע viscoelastic של תאים, וסופו של דבר לחשוף את המקורות המולקולריים של התנהגות זו.

כדי לחקור את deformability סוגי תאים שונים, תאים אנושיים מיאלואידית לוקמיה (HL-60), תאי נויטרופילים מובחנים, תאי הלימפוציטים עכבר, ושורות תאי סרטן השחלות אנושיות (OVCAR8, HEYA8) שלהם הוערכו באמצעות טכניקת microfluidic &#39;תא deformer&#39;. תוצאות עבור נציג את זמן המעבר של HL-60 תאי HL-60 ונויטרופילים ?...

Watch this Scientific Journal Video about A Microfluidic Technique to Probe Cell Deformability at JoVE.com

A Microfluidic Technique to Probe Cell Deformability

אנו מדגימים מבוססי assay מיקרופלואידיקה כדי למדוד את הזמנים לתאים למעבר דרך רצף של אילוצי מיקרון בקנה מידה.

טכניקת Microfluidic לחקור deformability הסלולרי

Here we detail the design, fabrication, and use of a microfluidic device to evaluate the deformability of a large number of individual cells in an ...

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11.2K Views.  University of California, Los Angeles.  אנו מדגימים מבוססי assay מיקרופלואידיקה כדי למדוד את הזמנים לתאים למעבר דרך רצף של אילוצי מיקרון בקנה מידה. 

Video: A Microfluidic Technique to Probe Cell Deformability

Systematic Analysis of In Vitro Cell Rolling Using a Multi-well Plate Microfluidic System

A major challenge for cell-based therapy is the inability to systemically target a large quantity of viable cells with high efficiency to tissues of interest following intravenous or intraarterial infusion. Consequently, increasing cell homing is currently studied as a strategy to improve cell therapy. Cell rolling on the vascular endothelium is an important step in the process of cell homing and can be probed in-vitro using a parallel plate flow chamber (PPFC). However, this is an extremely tedious, low throughput assay, with poorly controlled flow conditions. Instead, we used a multi-well plate microfluidic system that enables study of cellular rolling properties in a higher throughput under precisely controlled, physiologically relevant shear flow1,2. In this paper, we show how the rolling properties of HL-60 (human promyelocytic leukemia) cells on P- and E-selectin-coated surfaces as well as on cell monolayer-coated surfaces can be readily examined. To better simulate inflammatory conditions, the microfluidic channel surface was coated with endothelial cells (ECs), which were then activated with tumor necrosis factor-&#945; (TNF-&#945;), significantly increasing interactions with HL-60 cells under dynamic conditions. The enhanced throughput and integrated multi-parameter software analysis platform, that permits rapid analysis of parameters such as rolling velocities and rolling path, are important advantages for assessing cell rolling properties in-vitro. Allowing rapid and accurate analysis of engineering approaches designed to impact cell rolling and homing, this platform may help advance exogenous cell-based therapy.

Systematic Analysis of In Vitro Cell Rolling Using a Multi-well Plate Microfluidic System

This study used a multi-well plate microfluidic system, significantly increasing throughput of cell rolling studies under physiologically relevant shear flow. Given the importance of cell rolling in the multi-step cell homing cascade and the importance of cell homing following systemic delivery of exogenous populations of cells in patients, this system offers potential as a screening platform to improve cell-based therapy.

ניתוח שיטתי של<em&gt; במבחנה</em&gt; תא רולינג באמצעות מערכת microfluidic צלחת רב גם

A major challenge for cell-based therapy is the inability to systemically target a large quantity of viable cells with high efficiency to tissues of ...

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.

לוחץ תא כחזק, Microfluidic תאית פלטפורמה להצגה

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

Analyzing Mixing Inhomogeneity in a Microfluidic Device by Microscale Schlieren Technique

In this paper, we introduce the use of microscale schlieren technique to measure mixing inhomogeneity in a microfluidic device. The microscale schlieren system is constructed from a Hoffman modulation contrast microscope, which provides easy access to the rear focal plane of the objective lens, by removing the slit plate and replacing the modulator with a knife-edge. The working principle of microscale schlieren technique relies on detecting light deflection caused by variation of refractive index1-3. The deflected light either escapes or is obstructed by the knife-edge to produce a bright or a dark band, respectively. If the refractive index of the mixture varies linearly with its composition, the local change in light intensity in the image plane is proportional to the concentration gradient normal to the optical axis. The micro-schlieren image gives a two-dimensional projection of the disturbed light produced by three-dimensional inhomogeneity.
To accomplish quantitative analysis, we describe a calibration procedure that mixes two fluids in a T-microchannel. We carry out a numerical simulation to obtain the concentration gradient in the T-microchannel that correlates closely with the corresponding micro-schlieren image. By comparison, a relationship between the grayscale readouts of the micro-schlieren image and the concentration gradients presented in a microfluidic device is established. Using this relationship, we are able to analyze the mixing inhomogeneity from associate micro-schlieren image and demonstrate the capability of microscale schlieren technique with measurements in a microfluidic oscillator4. For optically transparent fluids, microscale schlieren technique is an attractive diagnostic tool to provide instantaneous full-field information that retains the three-dimensional features of the mixing process.

Herein, we describe a procedure that employs microscale schlieren technique to measure mixing inhomogeneity in a microfluidic device. Through calibration, distribution of concentration gradient can be derived from the micro-schlieren image.

ניתוח inhomogeneity ערבוב במכשיר microfluidic על ידי טכניקת microscale Schlieren

In this paper, we introduce the use of microscale schlieren technique to measure mixing inhomogeneity in a microfluidic device. The microscale schlieren ...

Microfluidic Genipin Deposition Technique for Extended Culture of Micropatterned Vascular Muscular Thin Films

The chronic nature of vascular disease progression requires the development of experimental techniques that simulate physiologic and pathologic vascular behaviors on disease-relevant time scales. Previously, microcontact printing has been used to fabricate two-dimensional functional arterial mimics through patterning of extracellular matrix protein as guidance cues for tissue organization. Vascular muscular thin films utilized these mimics to assess functional contractility. However, the microcontact printing fabrication technique used typically incorporates hydrophobic PDMS substrates. As the tissue turns over the underlying extracellular matrix, new proteins must undergo a conformational change or denaturing in order to expose hydrophobic amino acid residues to the hydrophobic PDMS surfaces for attachment, resulting in altered matrix protein bioactivity, delamination, and death of the tissues.
Here, we present a microfluidic deposition technique for patterning of the crosslinker compound genipin. Genipin serves as an intermediary between patterned tissues and PDMS substrates, allowing cells to deposit newly-synthesized extracellular matrix protein onto a more hydrophilic surface and remain attached to the PDMS substrates. We also show that extracellular matrix proteins can be patterned directly onto deposited genipin, allowing dictation of engineered tissue structure. Tissues fabricated with this technique show high fidelity in both structural alignment and contractile function of vascular smooth muscle tissue in a vascular muscular thin film model. This technique can be extended using other cell types and provides the framework for future study of chronic tissue- and organ-level functionality.

We present a method for microfluidic deposition of patterned genipin and fibronectin on PDMS substrates, allowing extended viability of vascular smooth muscle cell-dense tissues. This tissue fabrication method is combined with previous vascular muscular thin film technology to measure vascular contractility over disease-relevant time courses.

טכניקת microfluidic Genipin הפקדת לתרבות מורחבת של כלי דם micropatterned השרירים Thin Films

The chronic nature of vascular disease progression requires the development of experimental techniques that simulate physiologic and pathologic vascular ...

Using Adhesive Patterning to Construct 3D Paper Microfluidic Devices

We demonstrate the use of patterned aerosol adhesives to construct both planar and nonplanar 3D paper microfluidic devices. By spraying an aerosol adhesive through a metal stencil, the overall amount of adhesive used in assembling paper microfluidic devices can be significantly reduced. We show on a simple 4-layer planar paper microfluidic device that the optimal adhesive application technique and device construction style depends heavily on desired performance characteristics. By moderately increasing the overall area of a device, it is possible to dramatically decrease the wicking time and increase device success rates while also reducing the amount of adhesive required to keep the device together. Such adhesive application also causes the adhesive to form semi-permanent bonds instead of permanent bonds between paper layers, enabling single-use devices to be non-destructively disassembled after use. Nonplanar 3D origami devices also benefit from the semi-permanent bonds during folding, as it reduces the likelihood that unrelated faces may accidently stick together. Like planar devices, nonplanar structures see reduced wicking times with patterned adhesive application vs uniformly applied adhesive.

We demonstrate the use of patterned aerosol adhesives to construct 3D paper microfluidic devices. This method of adhesive application forms semi-permanent bonds between layers, enabling single-use devices to be non-destructively disassembled after use and to ease folding complex nonplanar structures.

באמצעות דפוסים דבקים כדי לבנות התקני 3D נייר Microfluidic

We demonstrate the use of patterned aerosol adhesives to construct both planar and nonplanar 3D paper microfluidic devices. By spraying an aerosol ...

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.

פלטפורמת Microfluidic עבור בידוד תפוקה גבוהה מתא בודד ותרבות

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 ...

Rapid Subtractive Patterning of Live Cell Layers with a Microfluidic Probe

The microfluidic probe (MFP) facilitates performing local chemistry on biological substrates by confining nanoliter volumes of liquids. Using one particular implementation of the MFP, the hierarchical hydrodynamic flow confinement (hHFC), multiple liquids are simultaneously brought in contact with a substrate. Local chemical action and liquid shaping using the hHFC, is exploited to create cell patterns by locally lysing and removing cells. By utilizing the scanning ability of the MFP, user-defined patterns of cell monolayers are created. This protocol enables rapid, real-time and spatially controlled cell patterning, which can allow selective cell-cell and cell-matrix interaction studies.

We present a protocol to perform subtractive patterning of live cell monolayers on a surface. This is achieved by local and selective lysis of adherent cells using a microfluidic probe (MFP). The cell lysate retrieved from local regions can be used for downstream analysis, enabling molecular profiling studies.

דפוסי Subtractive מהירים של שכבות תא חיות עם בדיקת Microfluidic

The microfluidic probe (MFP) facilitates performing local chemistry on biological substrates by confining nanoliter volumes of liquids. Using one ...

Multi-step Variable Height Photolithography for Valved Multilayer Microfluidic Devices

Microfluidic systems have enabled powerful new approaches to high-throughput biochemical and biological analysis. However, there remains a barrier to entry for non-specialists who would benefit greatly from the ability to develop their own microfluidic devices to address research questions. Particularly lacking has been the open dissemination of protocols related to photolithography, a key step in the development of a replica mold for the manufacture of polydimethylsiloxane (PDMS) devices. While the fabrication of single height silicon masters has been explored extensively in literature, fabrication steps for more complicated photolithography features necessary for many interesting device functionalities (such as feature rounding to make valve structures, multi-height single-mold patterning, or high aspect ratio definition) are often not explicitly outlined. 
Here, we provide a complete protocol for making multilayer microfluidic devices with valves and complex multi-height geometries, tunable for any application. These fabrication procedures are presented in the context of a microfluidic hydrogel bead synthesizer and demonstrate the production of droplets containing polyethylene glycol (PEG diacrylate) and a photoinitiator that can be polymerized into solid beads. This protocol and accompanying discussion provide a foundation of design principles and fabrication methods that enables development of a wide variety of microfluidic devices. The details included here should allow non-specialists to design and fabricate novel devices, thereby bringing a host of recently developed technologies to their most exciting applications in biological laboratories.

Multilayer microfluidic devices often involve the fabrication of master molds with complex geometries for functionality. This article presents a complete protocol for multi-step photolithography with valves and variable height features tunable to any application. As a demonstration, we fabricate a microfluidic droplet generator capable of producing hydrogel beads.

פוטו-ליתוגרפיה גובה רב שלבי משתנה עבור Valved רב שכבתי מכשירים microfluidic

Microfluidic systems have enabled powerful new approaches to high-throughput biochemical and biological analysis. However, there remains a barrier to ...

Large-area Scanning Probe Nanolithography Facilitated by Automated Alignment and Its Application to Substrate Fabrication for Cell Culture Studies

Scanning probe microscopy has enabled the creation of a variety of methods for the constructive (&#39;additive&#39;) top-down fabrication of nanometer-scale features. Historically, a major drawback of scanning probe lithography has been the intrinsically low throughput of single probe systems. This has been tackled by the use of arrays of multiple probes to enable increased nanolithography throughput. In order to implement such parallelized nanolithography, the accurate alignment of probe arrays with the substrate surface is vital, so that all probes make contact with the surface simultaneously when lithographic patterning begins. This protocol describes the utilization of polymer pen lithography to produce nanometer-scale features over centimeter-sized areas, facilitated by the use of an algorithm for the rapid, accurate, and automated alignment of probe arrays. Here, nanolithography of thiols on gold substrates demonstrates the generation of features with high uniformity. These patterns are then functionalized with fibronectin for use in the context of surface-directed cell morphology studies.

Here we present a protocol for wide-area scanning probe nanolithography enabled by the iterative alignment of probe arrays, as well as the utilization of lithographic patterns for cell-surface interaction studies.

ננוליתוגרפיה בדיקה סריקת שטח גדול בהנחייתם של יישור אוטומטי ויישומו המצע פבריקציה נוספת ללימודי תרבות תא

Scanning probe microscopy has enabled the creation of a variety of methods for the constructive (&#39;additive&#39;) top-down fabrication of ...

A Novel Tenorrhaphy Suture Technique with Tissue Engineered Collagen Graft to Repair Large Tendon Defects

Surgical management of large tendon defects with tendon grafts is challenging, as there are a finite number of sites where donors can be readily identified and used. Currently, this gap is filled with tendon auto-, allo-, xeno-, or artificial grafts, but clinical methods to secure them are not necessarily translatable to animals because of the scale. In order to evaluate new biomaterials or study a tendon graft made up of collagen type 1, we have developed a modified suture technique to help maintain the engineered tendon in alignment with the tendon ends. Mechanical properties of these grafts are inferior to the native tendon. To incorporate engineered tendon into clinically relevant models of loaded repair, a strategy was adopted to offload the tissue engineered tendon graft and allow for the maturation and integration of the engineered tendon in vivo until a mechanically sound neo-tendon was formed. We describe this technique using incorporation of the collagen type 1 tissue engineered tendon construct.

In this paper, we present an in vitro and in situ protocol to repair a tendon gap of up to 1.5 cm by filling it with engineered collagen graft. This was performed by developing a modified suture technique to take the mechanical load until the graft matures into the host tissue.