فإن الكتاب أود أن نعترف لويد اونغ لإدخال بناء في الإصدارات الأولى من هذه التقنية، والدكتور جيريمي Agresti لغطاء الضغط نصائح التصميم، والدكتور دونغبينغ تشى لمساعدته في افتعال غطاء الضغط. نحن ممتنون للمختبرات M. Teitell وP. Gunaratne لتوفير مجموعة متنوعة من العينات خلية للاختبار. نحن ممتنون لمؤسسة العلوم الوطنية (التوظيف جائزة DBI-1254185)، ومركز جونسون الشامل للسرطان في جامعة كاليفورنيا، وجامعة كاليفورنيا ومعهد العلوم السريرية بالحركة لدعم هذا العمل.

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الكتاب ليس لديهم تضارب المصالح في الكشف عنها.

<html> نحن هنا توفير إجراء تجريبي شامل لتحليل تشوه الخلايا تمر عبر قنوات ميكروفلويديك مقيدة باستخدام تدفق ضغط يحركها. وهناك سيناريو MATLAB يمكن معالجة البيانات الآلي (المواد التكميلية). يتم الاحتفاظ نسخة محدثة من قانون ( <a href="www.ibp.ucla.edu/research/rowat" target="_blank">www.ibp.ucla.edu/research/rowat</a> )....

التغييرات في شكل الخلية هي الحاسمة في السياقات البيولوجية عديدة. على سبيل المثال، كريات الدم الحمراء وكريات الدم البيضاء تشوه من خلال الشعيرات الدموية التي تكون أصغر من قطر الخاصة بهم 1. في ورم خبيث، يجب أن تشوه الخلايا السرطانية من خلال الثغرات الضيقة الخلالي وكذلك الأوعية الدموية متعرج والشبكات اللمفاوية البذور في مواقع الثانوية 2. للتحقيق في السلوك المادي للخلايا فردية، وأجهزة ميكروفلويديك تمثل منصة مثالية التي يمكن تخصيصها لدراسة مجموعة من السلوكيات الخلية بما في ذلك قدرتهم على الهجرة من خلال الثغرات الضيقة 3 وتشوه بشكل سلبي من خلال التشنج ميكرون النطاق 3- 9. Polydimethylsiloxane (PDMS) أجهزة ميكروفلويديك شفافة بصريا، مما التشوهات الخلية المراد تصور باستخدام المجهر الضوئي وتحليلها باستخدام أدوات معالجة الصور الأساسية. وعلاوة على ذلك، صفائف التشنج يمكن تعريفها بدقة، مما يتيح تحليل خلايا متعددة في وقت واحد معالإنتاجية التي تتجاوز العديد من التقنيات القائمة 10،11. هنا نقدم بروتوكول تجريبي مفصل لبحث التشوه الخلية باستخدام جهاز ميكروفلويديك في &quot;خلية Deformer&quot; PDMS. الجهاز مصمم بحيث خلايا المرور عبر التشنج متتابعة. هذه الهندسة هو شائع في سياقات الفسيولوجية، مثل الشعيرات الرئوية سرير 12. لقياس التشوه خلية، وقت العبور يوفر متري مريحة أن تقاس بسهولة والوقت اللازم للخلية الفردية لعبور خلال 4،6 انقباض واحد. للحفاظ على انخفاض الضغط المستمر عبر قنوات ضيقة أثناء العبور خلية، ونحن نستخدم تدفق ضغط يحركها. ويتضمن بروتوكول لدينا تعليمات مفصلة عن تصميم الجهاز وتصنيع وتشغيل الجهاز عن طريق ضغط يحركها تدفق وإعداد وتصوير من الخلايا، وكذلك معالجة الصور لقياس الوقت للخلايا لتشويه من خلال سلسلة من التشنج. ندرجكلا التصاميم جهاز ورمز معالجة البيانات كملفات رؤية تكميلية. على عينة تمثيلية من البيانات، وتبين لنا خلية العبور مرة من خلال سلسلة من التشنج بوصفها وظيفة من عدد من التشنج passaged. تحليل الجدول الزمني للخلايا لعبور الرغم من التشنج الضيقة للجهاز ميكروفلويديك يمكن أن تكشف عن الاختلافات في التشوه مجموعة متنوعة من أنواع الخلايا 4،5،13. الجهاز أثبت هنا يستكشف فريد عبور الخلية من خلال سلسلة من التشنج ميكرون الحجم؛ هذا التصميم يحاكي مسار متعرج تلك التجربة الخلايا في الدورة الدموية وتمكن أيضا سبر الخصائص الفيزيائية إضافية من الخلايا مثل وقت الاسترخاء.

<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

نحن هنا التفاصيل تصميم وتصنيع واستخدام جهاز ميكروفلويديك لتقييم التشوه من عدد كبير من الخلايا الفردية بطريقة فعالة. عادة، يمكن الحصول على البيانات الخاصة ب ~ 10 2 الخلايا داخل تجربة 1 ساعة. يتيح برنامج تحليل الصور آليا بعد تحليل التجربة كفاءة من بيانات الصورة، مما يتيح تجهيز أن يكتمل في غضون ساعات قليلة. هندسة الجهاز لدينا هي فريدة من نوعها في أن الخلايا يجب أن تشوه من خلال سلسلة من التشنج ميكرون النطاق، وبالتالي تمكين تشوه الأولي وتعتمد على الوقت للاسترخاء من الخلايا الفردية أن يعاير. ويتجلى مدى انطباق هذا الأسلوب لاللوكيميا البشري (HL-60) الخلايا. القيادة الخلايا لتشوه من خلال التشنج ميكرون النطاق باستخدام تدفق ضغط يحركها، نلاحظ أن البرولمفوسيت البشري (HL-60) خلايا تسد حظات انقباض الأول لفترة وسيطة من 9.3 ميللي ثانية قبل الركض بسرعة أكبر من خلال انقباض لاحقالأيونات مع الوقت عبور المتوسط ​​من 4.0 ميللي ثانية في انقباض. على النقيض من ذلك، جميع العابر حمض الريتينويك المعالجة (العدلات من نوع) HL-60 خلايا تسد انقباض الأول فقط 4.3 مللي ثانية قبل الركض من خلال التشنج اللاحقة مع الوقت عبور المتوسط ​​من 3.3 ميللي ثانية. هذه الطريقة يمكن توفير نظرة ثاقبة لطبيعة اللزجة من الخلايا، وتكشف في النهاية أصول الجزيئية من هذا السلوك.

للتحقيق في أنواع مختلفة من التشوه الخلايا، وخلايا سرطان الدم النخاعي الإنسان (HL-60)، خلايا العدلات متباينة، والخلايا اللمفاوية الماوس، والمبيض خطوط الخلايا السرطانية البشرية (OVCAR8، HEYA8) يتم تقييمها باستخدام تقنية على &quot;خلية Deformer&quot; ميكروفلويديك. نتائج ممثلة للمرة ع?...

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

A Microfluidic Technique to Probe Cell Deformability

علينا أن نبرهن على أساس مقايسة على microfluidics لقياس الزمني للخلايا لعبور سلسلة من التشنج ميكرون النطاق.

تقنية ميكروفلويديك للتحقيق التشوه خلية

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.  علينا أن نبرهن على أساس مقايسة على microfluidics لقياس الزمني للخلايا لعبور سلسلة من التشنج ميكرون النطاق. 

Video: A Microfluidic Technique to Probe Cell Deformability

Measurement of Bioelectric Current with a Vibrating Probe

Electric fields, generated by active transport of ions, are present in many biological systems and often serve important functions in tissues and organs. For example, they play an important role in directing cell migration during wound healing. Here we describe the manufacture and use of ultrasensitive vibrating probes for measuring extracellular electric currents. The probe is an insulated, sharpened metal wire with a small platinum-black tip (30-35 &mu;m), which can detect ionic currents in the &mu;A/cm2 range in physiological saline. The probe is vibrated at about 200 Hz by a piezoelectric bender. In the presence of an ionic current, the probe detects a voltage difference between the extremes of its movement. A lock-in amplifier filters out extraneous noise by locking on to the probe's frequency of vibration. Data are recorded onto computer. The probe is calibrated at the start and end of experiments in appropriate saline, using a chamber which applies a current of exactly 1.5 &mu;A/cm2. We describe how to make the probes, set up the system and calibrate. We also demonstrate the technique of cornea measurement, and show some representative results from different specimens (cornea, skin, brain).

The manufacture, calibration and use of non-invasive vibrating probes to measure bioelectric current in various biological systems is described.

قياس الحالية Bioelectric مع التحقيق الذي تجريه بالاهتزاز

Electric fields, generated by active transport of ions, are present in many biological systems and often serve important functions in tissues and organs. ...

Design of a Cyclic Pressure Bioreactor for the Ex Vivo Study of Aortic Heart Valves

The aortic valve, located between the left ventricle and the aorta, allows for unidirectional blood flow, preventing backflow into the ventricle. Aortic valve leaflets are composed of interstitial cells suspended within an extracellular matrix (ECM) and are lined with an endothelial cell monolayer. The valve withstands a harsh, dynamic environment and is constantly exposed to shear, flexion, tension, and compression. Research has shown calcific lesions in diseased valves occur in areas of high mechanical stress as a result of endothelial disruption or interstitial matrix damage1-3. Hence, it is not surprising that epidemiological studies have shown high blood pressure to be a leading risk factor in the onset of aortic valve disease4. 
The only treatment option currently available for valve disease is surgical replacement of the diseased valve with a bioprosthetic or mechanical valve5. Improved understanding of valve biology in response to physical stresses would help elucidate the mechanisms of valve pathogenesis. In turn, this could help in the development of non-invasive therapies such as pharmaceutical intervention or prevention. Several bioreactors have been previously developed to study the mechanobiology of native or engineered heart valves6-9. Pulsatile bioreactors have also been developed to study a range of tissues including cartilage10, bone11 and bladder12. The aim of this work was to develop a cyclic pressure system that could be used to elucidate the biological response of aortic valve leaflets to increased pressure loads. 
The system consisted of an acrylic chamber in which to place samples and produce cyclic pressure, viton diaphragm solenoid valves to control the timing of the pressure cycle, and a computer to control electrical devices. The pressure was monitored using a pressure transducer, and the signal was conditioned using a load cell conditioner. A LabVIEW program regulated the pressure using an analog device to pump compressed air into the system at the appropriate rate. The system mimicked the dynamic transvalvular pressure levels associated with the aortic valve; a saw tooth wave produced a gradual increase in pressure, typical of the transvalvular pressure gradient that is present across the valve during diastole, followed by a sharp pressure drop depicting valve opening in systole. The LabVIEW program allowed users to control the magnitude and frequency of cyclic pressure. The system was able to subject tissue samples to physiological and pathological pressure conditions. This device can be used to increase our understanding of how heart valves respond to changes in the local mechanical environment.

Design of a Cyclic Pressure Bioreactor for the Ex Vivo Study of Aortic Heart Valves

A cyclic pressure bioreactor capable of subjecting heart valve tissue to physiological and pathological pressure conditions has been designed. A LabVIEW program allows users to control pressure magnitude, amplitude and frequency. This device can be used to study the mechanobiology of heart valve tissue or isolated cells.

تصميم مفاعل حيوي لضغط الدوري السابقين فيفو دراسة صمامات القلب الأورطي

The aortic valve, located between the left ventricle and the aorta, allows for unidirectional blood flow, preventing backflow into the ventricle. Aortic ...

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 ...

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.

تحليل خلط التجانس في جهاز ميكروفلويديك التي كتبها الميكروسكيل انعراجي تقنية

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.

ميكروفلويديك Genipin ترسب تقنية للثقافة الموسعة من Micropatterned الأوعية الدموية العضلات الرقيقة

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.

باستخدام لاصق الزخرفة لانشاء 3D ورقة أجهزة ميكروفلويديك

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.

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 ...

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.

الزخرفة مطروح السريع لطبقات الخلايا الحية مع التحقيق ميكروفلويديك

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.

متعدد خطوة الطباعة التصويرية متغير الطول لمحكم بصمام متعدد الأجهزة ميكروفلويديك

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.