기금은 NIH MH - 64611와 NARSAD 젊은 탐정 수상에 의해 제공되었다. 저자는 또한 기술 지원 아담 ​​비글리, 마크 Dikopf, 그리고 벤 스미스 인정하고 싶습니다.

SU - 8 몰드 제작 석사 준비 <ol><li> 실리콘 웨이퍼 기판에 SU - 8 주인이 두 단계 표준 부정적인 저항 리소그래피 공정을 사용하여 준비가되어 있습니다. </li><li> 실리콘 웨이퍼의 정렬 마크 이러한 구조의 높이로 면도날을 사용하여 제거하면 (웨이퍼의 바깥 주변을 따라 위치) 실제 장치 구조 이상입니다. </li><li> 실리콘 웨이퍼는 다음 이소 프로필 알코올을 사용하여 청소 및 N 2의 스트림에 건조합니다. 적은 가장 높은 장치 구조보다 두께 테이프로 만들어 지원 기둥은 웨이퍼의 사면에 정렬 마크를 대체합니다. 참고 : 지원 기둥의 높이가 가장 높은 장치 구조보다 더 많은 경우를 통해 구멍이 PDMS 몰드를 형성하지 않습니다. </li><li> 마스터 실리콘 웨이퍼는 실온에서 뜨거운 접시에 배치됩니다. </li></ol> PDMS 솔루션 준비 <ol><li> polydimethylsiloxane (PDMS) 솔루션의 네 그램은 철저히 치료 요원 1 일부로 실리콘 탄성체의 10 부분을 혼합하여 준비가되어 있습니다. 참고 : 두 솔루션은 PDMS 금형에 걸쳐 유사한 물리적 성질을 얻기 위해 균일하게 혼합되어 있는지 확인합니다. </li><li> 혼합 과정에서 PDMS 솔루션에서 생성된 거품은 진공 건조기를 사용하여 제거됩니다. </li></ol> PDMS 코팅 및 경화 <ol><li> 거품없는 PDMS 솔루션은 서서히 거품이 분배 과정에서 생성되지 않도록하는 SU - 8 마스터에 적절합니다. </li><li> 쓰기에서 투명 필름의 한쪽 끝을 그 다음 철판에 위치하고 천천히 고르게 SU - 8 마스터에 PDMS를 확산하기 위해 PDMS 솔루션에 배치됩니다. 이 과정에서 생성된 모든 거품이 프로브를 사용하여 제거해야합니다. [참고 : 자세한 거품의 생성을 방지하기 위해 투명성을 들지 마십시오.] </li><li> borofloat의 슬래브는 균일한 압력을 적용하기 위해 투명 시트의 상단에 위치합니다. 젠틀 압력은 투명성과 원형의 SU - 8 표면 사이에 끼워 넣으면 최소한의 또는 전혀 PDMS가 그러한 높은 원형 구조의 상단 표면 (채널 길이의 중간에 위치하고 채널의 넓이를 따라 중심으로)에 적용 구조. </li><li> 세 이상 borofloat의 슬라브는 전에 원형 개방 구조의 상단 표면에와​​ 치료 과정에서 지속적인 압력을 적용하기 위해 실리콘 웨이퍼 borofloat 슬래브 샌드위치 상단에 표시됩니다. </li><li> 뜨거운 접시의 온도가 다음 75 증가 ° C와 PDMS는 1 시간 동안이 온도에서 치료됩니다. 핫 플레이트 50 ° C.의 온도를 데려 </li><li> 슬라브이 제거되고 투명도가 부드럽게 마스터 및 그것의 위에 코팅된 얇은 PDMS 시트 남겨두고 제거됩니다. </li></ol> microfluidic 장치의 건설 마스터에서 PDMS 시트 제거 <ol><li> 재관류 챔버의 외부 경계는 SU - 8 마스터에있는 포트와 재관류 챔버에있는 포트를 정렬 후 면도날을 사용하여 PDMS 시트에 나가 새겨져있다. </li><li> PDMS 시트는 부드럽게 PDMS의 찢어 방지하기 위해 채널의 길이를 따라 당기, 마스터에서 제거됩니다. PDMS 시트가 위쪽을 향하고 microfluidic 네트워크 표면을 가진 투명 시트에 배치됩니다. </li><li> 입구와 출구 포트는 다음 코크 송곳을 사용하여 만들어집니다. </li></ol> 유리 coverslip에 PDMS 시트의 결합 <ol><li> PDMS 시트 (표면이 microfluidic 네트워크를 포함) 및 유리 coverslip의 결합 표면은 부드럽게 투명성의 시트에 배치, 3M의 스카치 테이프를 사용하여 세척하고, 마지막으로 O 2 플라즈마 챔버에 배치됩니다. </li><li> 접착 표면 10 S.위한 165 왓츠 플라즈마로 처리됩니다 유리 coverslip의 플라즈마 표면 처리는 즉시 PDMS 시트의 플라즈마 표면 처리 접착됩니다. 참고 : 결합 표면 사이에 갇혀있는 공기 방울을 제거하기 위해 유리 표면에 부드러운 압력을 적용합니다. </li><li> 5 분 PDMS와 유리 사이 좋은 결합을 얻을 수 있습니다. 5 분 후에 부드럽게 PDMS 시트가 결합 프로세스에 삽입되었던에 대한 투명성을 제거합니다. </li><li> microfluidic 장치는 채널이 친수성 ​​만들기 위해 산소 플라즈마 시스템에 배치됩니다. 플라즈마 처리는 1 분 165 왓츠에서 이루어집니다. </li></ol> microfluidic 장치 및 재관류 챔버의 통합 재관류 챔버의 준비 <ol><li> 재고품의 재관류 챔버가 주문되었고, 입구와 콘센트 포트 microfluidic 장치와 챔버가 보세 때, 챔버에있는 포트와 장치가 정렬되도록 뚫고 있었다. 재관류 챔버의 바닥 표면은 P로 코팅되어DMS 챔버와 microfluidic 장치 사이에 긴밀한 인감을 얻을 수 있습니다. </li><li> 쓰기에 투명 시트는 실온에서 뜨거운 접시에 배치됩니다. </li><li> 한 g PDMS 솔루션은 앞에서 설명한 것과 유사한 과정에 준비가되어 있습니다. 거품없는 PDMS 솔루션은 다음 천천히 거품의 투명 시트 피하 세대에 적절합니다. </li><li> 재관류 챔버는 다음 PDMS 솔루션에 배치됩니다. </li><li> borofloat의 슬래브는 다음 균일한 압력을 적용하고 챔버의 바닥 표면에 PDMS의 얇은 코팅을 얻기 위해 챔버의 상단에 위치합니다. </li><li> 뜨거운 접시의 온도가 다음 75 증가 ° C와 PDMS는 1 시간 동안이 온도에서 치료됩니다. </li></ol> microfluidic 장치 및 재관류 챔버의 결합 <ol><li> 슬래브가 제거되고 투명성이 부드럽게은 아래쪽 표면에 재관류 챔버와 얇은 PDMS 시트 코팅 남겨두고 제거됩니다. 원치 않는 PDMS는 면도날 및 액세스 포트에서 PDMS 제거에 대한 날카로운 팁 프로브를 사용하여 제거됩니다. </li><li> microfluidic 장치와 챔버의 PDMS 바닥 표면의 PDMS의 표면은 부드럽게, 3M 테이프를 사용하여 청소 투명성의 시트에 배치하고, 마지막으로 산소 플라즈마 챔버에 배치됩니다. </li><li> 표면 10 S.위한 165 왓츠 플라즈마로 처리됩니다 챔버의 PDMS 코팅 표면은 즉시 microfluidic 장치의 PDMS의 표면에 접착됩니다. 참고 : 결합 표면 사이에 갇혀있는 공기 방울을 제거하기 위해 유리 표면에 부드러운 압력을 적용합니다. </li><li> 즉시 사용하기 전에 장치가 채널이 친수성 ​​만들기 위해 산소 플라즈마 시스템에 배치됩니다. 플라즈마 처리는 1 분 165 왓츠에서 이루어집니다. </li></ol> microfluidic 장치를 사용 neurochemical microenvironment 두뇌 조각 노출 <ol><li> (microfluidic 장치)의 친수성 ​​microchannels - (재관류 챔버)을 조합 표준 ACSF (인공 대뇌 척수) 솔루션으로 가득합니다. </li><li> 입구 포트에 ACSF 솔루션의 작은 방울을 분배하고 솔루션은 최대 채널로 악인 될 수 있습니다. 콘센트 포트에서 주사기를 사용하여 남아있는 거품을 제거합니다. 입구에서 microfluidic 장치의 출구 포트에 유체의 수동 펌프 수 있도록 콘센트 포트에서 ACSF 솔루션의 큰 방울을 분배. 참고 : 관리가 수동 펌프 방식을 사용하여 유체의 흐름을 허용하는 채널에서 모든 거품을 제거로 이동해야합니다. </li><li> (microfluidic 장치) 수정 - (재관류 챔버) 일반 플랫폼에서 콤보를하고 현미경 어댑터에 일반 플랫폼을 수정. </li><li> 표준 입구와 콘센트 (흡입) 표준 ACSF 솔루션으로 뇌 슬라이스의 지속적인 재관류에 대한 재관류 챔버에 튜브를 연결합니다. ACSF 솔루션은 지속적으로 95% O2 - 5% CO 2 aspirated입니다. 참고 : 재관류 챔버의 욕조에서 ACSF의 일정한 수준을 유지하기 위해 흡입 튜브의 위치를 조정합니다. </li><li> 재관류 챔버가 ACSF 솔루션으로 가득되면, 스포이트를 사용하여 재관류 챔버의 뇌 조각을 놓으십시오. 프로브를 사용 microfluidic 장치에 원형의 구멍 위에 뇌 조각의 위치. 두뇌 슬라이스가 원형 구멍을 통해 원하는 위치에되면, 뇌의 슬라이스를 무력화하기 위해 슬라이스 앵커를 사용합니다. </li><li> microfluidic 장치가 지금은 유체의 펌핑 패시브를 사용하여 다양한 신경 전달 물질로 두뇌 조각 (재관류 챔버와 microfluidic 네트워크의 상단에 위치) 노출하는 데 사용할 수 있습니다. </li></ol>

<ol>
	<li>Blake, A. J., Pearce, T. M., Rao, N. S., Johnson, S. M., Williams, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multilayer+PDMS+microfluidic+chamber+for+controlling+brain+slice+microenvironment.">Multilayer PDMS microfluidic chamber for controlling brain slice microenvironment.</a> Lab on a Chip. 7, 842-849 (2007).</li><li>Walker, G. M., Beebe, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+passive+pumping+method+for+microfluidic+devices.">A passive pumping method for microfluidic devices.</a> Lab on a Chip. 2, 131-134 (2002).</li></ol>

저자는 생물학의 다른 분야에 대한 시연 microfluidic 기술을 포함한 협력 열려 있습니다.

기존 macroscale 또는 microscale 두뇌 슬라이스 재관류 챔버스들은 신경 전달 물질로 뇌 조각을 폭로 제공하는 공간 해상도 측면에서 한계가 있습니다. 여기 시연 microfluidic 장치 기술은 간단한 bioMEMS 기법을 사용하여이 제한을 극복. 그것은 microfluidic 장치의 제조 및 기존 전기 생리학의 설정과 통합의 용이성에 단순이 증명 장치 기술의 광범위한 응용 프로그램을 만들 것이라는 것을 예상합니다. 불가능되지 않은 흥?...

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		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>RC-26GPL</td>
<td>Tool</td>
<td>Warner Instruments</td>
<td>W2-64-0236</td>
<td>Low Profile Large Bath RC-26GLP Recording Chamber</td>
</tr>
<tr>
<td>SHD-26GH/10</td>
<td>Tool</td>
<td>Warner Instruments</td>
<td>W2-64-0253</td>
<td>Stainless steel slice hold-down for RC-26G, 1.0 mm thread spacing </td>
</tr>
<tr>
<td>PDMS (polydimethylsiloxane)</td>
<td>Reagent</td>
<td>Dow Corning</td>
<td>Sylgard 184</td>
<td>Silicone Elastomer Kit</td>
</tr>
<tr>
<td>Plasma Preen-II 862</td>
<td>Tool</td>
<td>Plasmatic Systems, Inc.</td>
<td>&nbsp;</td>
<td>Microwave plasma system</td>
</tr>
<tr>
<td>Model P-1</td>
<td>Tool</td>
<td>Warner Instruments</td>
<td>W2-64-0277</td>
<td>Series 20 Plain Platform, Model P-1</td>
</tr>
<tr>
<td>SA-NIK</td>
<td>Tool</td>
<td>Warner Instruments</td>
<td>W2-64-0291</td>
<td>Adapter for Nikon Diaphot/TE200/TE2000, SA-NIK</td>
</tr>
<tr>
<td>Oxygenated, heated ACSF (Artificial cerebro-spinal fluid)</td>
<td>Reagent</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>Exact composition will vary with application</td>
</tr>		</tbody>
		</table>
		</div>

brain slice stimulation using a microfluidic network and standard perfusion chamber

우리는 쉽게 기존의 전기 생리학의 설정과 통합 수있는 두 수준 microfluidic 장치의 제조를 증명하고있다. 두 수준 microfluidic 장치는 두 단계 표준 리소그래피 프로세스 1 저항 부정을 사용하는 조작이다. 첫 번째 수준은 각 끝에 유입 및 배출구 포트 microchannels가 포함되어 있습니다. 두 번째 수준은 채널 길이의 중간에 위치하고 채널 폭과 함께 중심으로 microscale 원형 구멍을 포함하고 있습니다. 수동 펌프 방법은 입구 포트에서 배출구 포트 2 유체를 펌프하는 데 사용됩니다. microfluidic 장치는 재고품 재관류의 챔버와 통합 및 전기 생리학 설정과 완벽한 통합이 가능합니다. 유체의 출구 포트를 향해 microchannels를 통해 유입 포트 흐름에 도입하고 또한 재관류의 목욕탕에 microchannels 상단에있는 원형 구멍을 통해 탈출. 따라서 재관류 챔버 목욕과 microfluidic 장치 위에 위치 뇌 슬라이스의 바닥 표면은 서로 다른 신경 전달 물질과 함께 노출 수 있습니다. microfluidic 장치의 microscale 두께와 소재의 투명한 자연 [유리 coverslip와 PDMS (polydimethylsiloxane)] microfluidic 장치가 뇌 슬라이스의 현미경을 허용하기 위해 사용됩니다. microfluidic 장치는 뇌의 슬라이스 m​​icroenvironments 소개 화학 자극의 변조 (공간과 시간적 모두) 수 있습니다.

Watch this Scientific Journal Video about Brain Slice Stimulation Using a Microfluidic Network and Standard Perfusion Chamber at JoVE.com

Brain Slice Stimulation Using a Microfluidic Network and Standard Perfusion Chamber

우리는 서로 다른 신경 전달 물질로 잘 제어 방식으로 뇌 슬라이스의 microscale 표면 노출 표준 전기 생리학의 설정과 통합되는 간단한 microfluidic 장치의 제조를 보여줍니다.

Microfluidic 네트워크 및 표준 재관류 회의소를 사용하여 뇌 슬라이스 자극

We have demonstrated the fabrication of a two-level microfluidic device that can be easily integrated with existing electrophysiology setups. The ...

brain-slice-stimulation-using-microfluidic-network-standard-perfusion

Research

JoVE Journal

Biology

10.8K 조회수.  University of Illinois, Chicago. 우리는 서로 다른 신경 전달 물질로 잘 제어 방식으로 뇌 슬라이스의 microscale 표면 노출 표준 전기 생리학의 설정과 통합되는 간단한 microfluidic 장치의 제조를 보여줍니다.

비디오: Brain Slice Stimulation Using a Microfluidic Network and Standard Perfusion Chamber

Fabrication of a Microfluidic Device for the Compartmentalization of Neuron Soma and Axons

In this video, we demonstrate the technique of soft lithography with polydimethyl siloxane (PDMS) which we use to fabricate a microfluidic device for culturing neurons. Previously, a silicon wafer was patterned with the design for the neuron microfluidic device using SU-8 and photolithography to create a master mold, or what we simply refer to as a "master". Next, we pour the silicon polymer PDMS on top of the master which is then cured by heating the PDMS to 80&deg;C for 1 hour. The PDMS forms a negative mold of the device. The PDMS is then carefully cut and lifted away from the master. Holes are punched where the reservoirs will be and the excess PDMS trimmed away from the device. Nitrogen is used to blow away any excess debris from the device. At this point the devices are now ready for use and can either bonded to corning No. 1 cover glass with a plasma sterilizer/cleaner or can be reversibly bound to the cover glass by simply placing the device on top of the cover glass. The reversible bonding of the device to glass is covered in a separate video and requires first that the device be sterilized either with 70% ethanol or by autoclaving. Plasma treating sterilizes the devices so no further treatment is necessary. It is, however, important, when plasma-treating the devices, to add liquid to the devices within 10 minutes of the plasma treatment while the surfaces are still hydrophilic. Waiting longer than 10 minutes to add liquid to the device makes it difficult for the liquid to enter the device. The neuron devices are typically plasma-bound to cover glass and 0.5 mg/ml poly-L-lysine (PLL) in pH 8.5 borate buffer is immediately added to the device. After a minimum of 3 hours incubating with PLL, the devices are washed with dH2O water a minimum of 3 times with at least 15 minutes between each wash. Next, the water is removed and fresh media is added to the device. At this point the device is ready for use. It is important to remember at this point to never remove all the media from the device. Always leave media in the main channel.

In this video we demonstrate the technique of soft lithography with polydimethyl siloxane (PDMS) which we use to farbricate a microfluidic device for culturing neurons.

신경 소마와 Axons의 Compartmentalization위한 Microfluidic 장치의 제작

In this video, we demonstrate the technique of soft lithography with polydimethyl siloxane (PDMS) which we use to fabricate a microfluidic device for ...

연구

생물학

Preparing E18 Cortical Rat Neurons for Compartmentalization in a Microfluidic Device

In this video, we demonstrate the preparation of E18 cortical rat neurons. E18 cortical rat neurons are obtained from E18 fetal rat cortex previously dissected and prepared. The E18 cortex is, upon dissection, immediately dissociated into individual neurons. It is possible to store E18 cortex in Hibernate E buffer containing B27 at 4&deg;C for up to a week before the dissociation is performed. However, there will be a drop in cell viability. Typically we obtain our E18 Cortex fresh. It is transported to the lab in ice cold Calcium free Magnesium free dissection buffer (CMFM). Upon arrival, trypsin is added to the cortex to a final concentration of 0.125%. The cortex is then incubated at 37&deg;C for 8 minutes. DMEM containing 10% FBS is added to the cortex to stop the reaction. The cortex is then centrifuged at 2500 rpm for 2 minutes. The supernatant is removed and 2 ml of Neural Basal Media (NBM) containing 2% B27 (vol/vol) and 0.25% Glutamax (vol/vol) is added to the cortex which is then re-suspended by pipetting up and down. Next, the cortex is triturated with previously fire polished glass pipettes, each with a successive smaller opening. After triturating, the cortex is once again centrifuged at 2500 rpm for 2 minutes. The supernatant is then removed and the cortex pellet re-suspended with 2 ml of NBM containing B27 and Glutamax. The cell suspension is then passed through a 40 um nylon cell strainer. Next the cells are counted. The neurons are now ready for loading into the neuron microfluidic device.

In this video we demonstrate the preparation of E18 Cortical Rat Neurons.

Microfluidic 장치에서 Compartmentalization에 대한 E18 두피 랫 뉴런 준비

In this video, we demonstrate the preparation of E18 cortical rat neurons. E18 cortical rat neurons are obtained from E18 fetal rat cortex previously ...

CD4+ T-Lymphocyte Capture Using a Disposable Microfluidic Chip for HIV

CD4 + HIV에 대한 일회용 Microfluidic 칩을 사용하여 T - 림프구 캡처

Cell Capture Using a Microfluidic Device

Microfluidic 장치를 사용하여 셀 캡처

Assessing Neural Stem Cell Motility Using an Agarose Gel-based Microfluidic Device

While microfluidic technology is reaching a new level of maturity for macromolecular assays, cell-based assays are still at an infant stage1. This is largely due to the difficulty with which one can create a cell-compatible and steady microenvironment using conventional microfabrication techniques and materials. We address this problem via the introduction of a novel microfabrication material, agarose gel, as the base material for the microfluidic device. Agarose gel is highly malleable, and permeable to gas and nutrients necessary for cell survival, and thus an ideal material for cell-based assays. We have shown previously that agarose gel based devices have been successful in studying bacterial and neutrophil cell migration2. In this report, three parallel microfluidic channels are patterned in an agarose gel membrane of about 1mm thickness. Constant flows with media/buffer are maintained in the two side channels using a peristaltic pump. Cells are maintained in the center channel for observation. Since the nutrients and chemicals in the side channels are constantly diffusing from the side to center channel, the chemical environment of the center channel is easily controlled via the flow along the side channels. Using this device, we demonstrate that the movement of neural stem cells can be monitored optically with ease under various chemical conditions, and the experimental results show that the over expression of epidermal growth factor receptors (EGFR) enhances the motility of neural stem cells. Motility of neural stem cells is an important biomarker for assessing cells aggressiveness, thus tumorigenic factor3. Deciphering the mechanism underlying NSC motility will yield insight into both disorders of neural development and into brain cancer stem cell invasion.

We demonstrate that the over expression of epidermal growth factor receptors (EGFR) enhances the motility of neural stem cells(NSCs) using a novel agarose gel based microfluidic device. This technology can be readily adaptable to other mammalian cell systems where cell sources are scarce, such as human neural stem cells, and the turn around time is critical.

아가로 오스 겔 기반 Microfluidic 장치를 사용 신경 줄기 세포의 운동성 평가

While microfluidic technology is reaching a new level of maturity for macromolecular assays, cell-based assays are still at an infant stage1. This is ...

Trypsinizing and Subculturing Mammalian Cells

As cells reach confluency, they must be subcultured or passaged. Failure to subculture confluent cells results in reduced mitotic index and eventually in cell death. The first step in subculturing is to detach cells from the surface of the primary culture vessel by trypsinization or mechanical means. The resultant cell suspension is then subdivided, or reseeded, into fresh cultures. Secondary cultures are checked for growth and fed periodically, and may be subsequently subcultured to produce tertiary cultures. The time between passaging of cells varies with the cell line and depends on the growth rate.

As cells reach confluency, they must be subcultured or passaged. This video will demonstrate a procedure for subculturing both adherent and suspension cells.

포유 동물 세포를 Trypsinizing 및 Subculturing

As cells reach confluency, they must be subcultured or passaged. Failure to subculture confluent cells results in reduced mitotic index and eventually in ...

The Microfluidic Probe: Operation and Use for Localized Surface Processing

Microfluidic devices allow assays to be performed using minute amounts of sample and have recently been used to control the microenvironment of cells. Microfluidics is commonly associated with closed microchannels which limit their use to samples that can be introduced, and cultured in the case of cells, within a confined volume. On the other hand, micropipetting system have been used to locally perfuse cells and surfaces, notably using push-pull setups where one pipette acts as source and the other one as sink, but the confinement of the flow is difficult in three dimensions. Furthermore, pipettes are fragile and difficult to position and hence are used in static configuration only. 
The microfluidic probe (MFP) circumvents the constraints imposed by the construction of closed microfluidic channels and instead of enclosing the sample into the microfluidic system, the microfluidic flow can be directly delivered onto the sample, and scanned across the sample, using the MFP. . The injection and aspiration openings are located within a few tens of micrometers of one another so that a microjet injected into the gap is confined by the hydrodynamic forces of the surrounding liquid and entirely aspirated back into the other opening. The microjet can be flushed across the substrate surface and provides a precise tool for localized deposition/delivery of reagents which can be used over large areas by scanning the probe across the surface. 
In this video we present the microfluidic probe1 (MFP). We explain in detail how to assemble the MFP, mount it atop an inverted microscope, and align it relative to the substrate surface, and finally show how to use it to process a substrate surface immersed in a buffer.

In this video we present the microfluidic probe1 (MFP). We explain in detail how to assemble the MFP, mount it atop an inverted microscope, and align it relative to the substrate surface, and finally show how to use it to process a substrate surface immersed in a buffer solution.

Microfluidic 프로브 : 운영 및 현지화된 표면 처리에 사용

Microfluidic devices allow assays to be performed using minute amounts of sample and have recently been used to control the microenvironment of cells. ...

Platelet Adhesion and Aggregation Under Flow using Microfluidic Flow Cells

Platelet aggregation occurs in response to vascular injury where the extracellular matrix below the endothelium has been exposed. The platelet adhesion cascade takes place in the presence of shear flow, a factor not accounted for in conventional (static) well-plate assays. This article reports on a platelet-aggregation assay utilizing a microfluidic well-plate format to emulate physiological shear flow conditions. Extracellular proteins, collagen I or von Willebrand factor are deposited within the microfluidic channel using active perfusion with a pneumatic pump. The matrix proteins are then washed with buffer and blocked to prepare the microfluidic channel for platelet interactions. Whole blood labeled with fluorescent dye is perfused through the channel at various flow rates in order to achieve platelet activation and aggregation. Inhibitors of platelet aggregation can be added prior to the flow cell experiment to generate IC50 dose response data.

The platelet adhesion cascade takes place in the presence of shear flow, a factor not accounted for in conventional (static) well-plate assays. This article reports on a platelet-aggregation assay utilizing a microfluidic well-plate format to emulate physiological shear flow conditions.

Microfluidic 흐름 전지를 사용하여 흐름에서 혈소판 접착 및 집계

Platelet aggregation occurs in response to vascular injury where the extracellular matrix below the endothelium has been exposed. The platelet adhesion ...

Mapping the After-effects of Theta Burst Stimulation on the Human Auditory Cortex with Functional Imaging

Auditory cortex pertains to the processing of sound, which is at the basis of speech or music-related processing1. However, despite considerable recent progress, the functional properties and lateralization of the human auditory cortex are far from being fully understood. Transcranial Magnetic Stimulation (TMS) is a non-invasive technique that can transiently or lastingly modulate cortical excitability via the application of localized magnetic field pulses, and represents a unique method of exploring plasticity and connectivity. It has only recently begun to be applied to understand auditory cortical function 2. 
An important issue in using TMS is that the physiological consequences of the stimulation are difficult to establish. Although many TMS studies make the implicit assumption that the area targeted by the coil is the area affected, this need not be the case, particularly for complex cognitive functions which depend on interactions across many brain regions 3. One solution to this problem is to combine TMS with functional Magnetic resonance imaging (fMRI). The idea here is that fMRI will provide an index of changes in brain activity associated with TMS. Thus, fMRI would give an independent means of assessing which areas are affected by TMS and how they are modulated 4. In addition, fMRI allows the assessment of functional connectivity, which represents a measure of the temporal coupling between distant regions. It can thus be useful not only to measure the net activity modulation induced by TMS in given locations, but also the degree to which the network properties are affected by TMS, via any observed changes in functional connectivity.
Different approaches exist to combine TMS and functional imaging according to the temporal order of the methods. Functional MRI can be applied before, during, after, or both before and after TMS. Recently, some studies interleaved TMS and fMRI in order to provide online mapping of the functional changes induced by TMS 5-7. However, this online combination has many technical problems, including the static artifacts resulting from the presence of the TMS coil in the scanner room, or the effects of TMS pulses on the process of MR image formation. But more importantly, the loud acoustic noise induced by TMS (increased compared with standard use because of the resonance of the scanner bore) and the increased TMS coil vibrations (caused by the strong mechanical forces due to the static magnetic field of the MR scanner) constitute a crucial problem when studying auditory processing. 
This is one reason why fMRI was carried out before and after TMS in the present study. Similar approaches have been used to target the motor cortex 8,9, premotor cortex 10, primary somatosensory cortex 11,12 and language-related areas 13, but so far no combined TMS-fMRI study has investigated the auditory cortex. The purpose of this article is to provide details concerning the protocol and considerations necessary to successfully combine these two neuroscientific tools to investigate auditory processing. 
Previously we showed that repetitive TMS (rTMS) at high and low frequencies (resp. 10 Hz and 1 Hz) applied over the auditory cortex modulated response time (RT) in a melody discrimination task 2. We also showed that RT modulation was correlated with functional connectivity in the auditory network assessed using fMRI: the higher the functional connectivity between left and right auditory cortices during task performance, the higher the facilitatory effect (i.e. decreased RT) observed with rTMS. However those findings were mainly correlational, as fMRI was performed before rTMS. Here, fMRI was carried out before and immediately after TMS to provide direct measures of the functional organization of the auditory cortex, and more specifically of the plastic reorganization of the auditory neural network occurring after the neural intervention provided by TMS. 
Combined fMRI and TMS applied over the auditory cortex should enable a better understanding of brain mechanisms of auditory processing, providing physiological information about functional effects of TMS. This knowledge could be useful for many cognitive neuroscience applications, as well as for optimizing therapeutic applications of TMS, particularly in auditory-related disorders.

Auditory processing is the basis of speech and music-related processing. Transcranial Magnetic Stimulation (TMS) has been used successfully to study cognitive, sensory and motor systems but has rarely been applied to audition. Here we investigated TMS combined with functional Magnetic Resonance Imaging to understand the functional organization of auditory cortex.

쎄의 후유증을 매핑하는 기능 영상과 함께 인간 청각 컴퓨터에 접속 자극을 버스트

Auditory cortex pertains to the processing of sound, which is at the basis of speech or music-related processing1. However, despite considerable recent ...

Mechanical Stimulation of Chondrocyte-agarose Hydrogels

Articular cartilage suffers from a limited repair capacity when damaged by mechanical insult or degraded by disease, such as osteoarthritis. To remedy this deficiency, several medical interventions have been developed. One such method is to resurface the damaged area with tissue-engineered cartilage; however, the engineered tissue typically lacks the biochemical properties and durability of native cartilage, questioning its long-term survivability. This limits the application of cartilage tissue engineering to the repair of small focal defects, relying on the surrounding tissue to protect the implanted material. To improve the properties of the developed tissue, mechanical stimulation is a popular method utilized to enhance the synthesis of cartilaginous extracellular matrix as well as the resultant mechanical properties of the engineered tissue. Mechanical stimulation applies forces to the tissue constructs analogous to those experienced in vivo. This is based on the premise that the mechanical environment, in part, regulates the development and maintenance of native tissue1,2. The most commonly applied form of mechanical stimulation in cartilage tissue engineering is dynamic compression at physiologic strains of approximately 5-20% at a frequency of 1 Hz1,3. Several studies have investigated the effects of dynamic compression and have shown it to have a positive effect on chondrocyte metabolism and biosynthesis, ultimately affecting the functional properties of the developed tissue4-8. In this paper, we illustrate the method to mechanically stimulate chondrocyte-agarose hydrogel constructs under dynamic compression and analyze changes in biosynthesis through biochemical and radioisotope assays. This method can also be readily modified to assess any potentially induced changes in cellular response as a result of mechanical stimuli.

The biosynthesis of cartilaginous extracellular matrix by chondrocytes can be affected by application of mechanical stimuli. This method describes the technique of applying dynamic compressive strains to chondrocytes encapsulated in 3D constructs and the evaluation of induced changes in chondrocyte metabolism.

Chondrocyte - 아가로 오스 Hydrogels의 기계적 자극

Articular cartilage suffers from a limited repair capacity when damaged by mechanical insult or degraded by disease, such as osteoarthritis. To remedy ...