이 작품은 양적 생명 과학의 캘리포니아 공과 대학 (QB3), 로저스 가족 재단, 혁신적인 생물 의학 연구, BASF에서 보조금에 대한 UCSF / 샌들러 재단 프로그램에서 브리징 격차 상, 그리고 NSF를 통해로부터 연구 상 수상에 의해 지원되었다 학부 조기 경력 개발 (경력) 프로그램 (DBI-1253293).

1. SU8 마스터 제작 <ol><li> AutoCAD 소프트웨어를 사용하여 두 개의 층을 제조하기위한 미세 유체 구조를 디자인하고 10 μm의 해상도로 회로 기판 필름 공급 업체에서 인쇄 된 디자인을 가지고있다. 장치 설계의 세부 사항은 첨부 된 기준 (11)에 부여하고, 채널 형상은도 1에 도시되어있다. 각 층은 층 (12)의 제조에서의 기능을 배치 할 수 있도록 정렬 마크를 포함한다. </li><li> 스핀 코터에 미리 청소 3 인치 직경의 실리콘 웨이퍼를 놓고 척에 부착하는 진공를 켜. 50 ㎛의 막 두께를 제공하고, 2,000 rpm에서 30 초, 500 rpm에서 20 초간 스핀 웨이퍼의 중심에 SU8-3035 1 ㎖를 적용. </li><li> 웨이퍼를 제거하고, 30 분 동안 135 ° C 열판에 굽는다. 웨이퍼는 다음 단계로 이동하기 전에 실온으로 냉각되도록. </li><li> 1 차 레이어 마스크로 코팅 된 웨이퍼 (그림 2A 노출 </stronG&gt;) 90 초 동안 LED nm의 평행 190 mW의 365 미만. 노출 후, 다음 단계를 진행하기 전에, 1 분 동안 실온에 다음 시원한를 135 ° C 형 열판에 웨이퍼를 놓습니다. </li><li> 스핀 코터에 웨이퍼를 놓고 척에 부착하는 진공의 전원을 켭니다. 135 ㎛의 두께의 부가 층을 제공 결과, 1,375 rpm에서 30 초, 500 rpm에서 20 초간 스핀 웨이퍼의 중심에 SU8-2050 1 ㎖를 적용. </li><li> 다음 단계로 이동하기 전에 30 분 동안 135 ° C의 열판에 RT에 다음 멋진 웨이퍼와 빵을 제거합니다. </li><li> 1.3 패턴 형상에 2 차 레이어 마스크 (그림 2B)를 맞추고 3 분 동안 LED nm의 평행 190 mW의 365에 코팅 된 웨이퍼를 노출. 노출 후, 다음 단계를 진행하기 전에, 1 분 동안 135 ° C의 열판에 RT에 다음 시원한 놓습니다. </li><li> 30 분 동안 프로필렌 글리콜 모노 메틸 에테르 아세테이트 교반 조에서 침지하여 마스크를 개발한다. 웨이퍼를 세척1 분 동안 135 ° C의 열판에 이소프로판올 및 빵에. PDMS 성형 100mm 페트리 접시에서 개발 마스터를 놓습니다. </li></ol> 2. PDMS 소자 제조 <ol><li> 플라스틱 컵에 경화제 5 g과 실리콘계의 50g을 조합하여 1 PDMS : 10을 준비한다. 볶음 스틱 장착 된 회전 도구를 사용하여 내용을 섞는다. 모든 기포가 제거되고 30 분 동안, 또는 건조기 내부까지 혼합물을 탈기. </li><li> 마스터 이상 3mm의 두께를주고 더 탈 가스에 대한 데시 케이 터에 다시 배치 할 PDMS를 붓고. 일단 모든 기포를 제거하고, 2 시간 동안 60 ℃에서 구워 장치. </li><li> 패턴면을 위로하여 깨끗한 표면에 메스과 장소를 사용하여 금형에서 장치를 잘라. 마스터 2 (그림 3a)에서 석사 (1)를 분리하기 위해 면도날로 절반에 PDMS 몰드를 잘라. 마스터 (1)에 의해 임프린트 된 50㎛의 유체 취급을 포함하는 형상 부재에, 0.7와 유체 유입구 및 출구를 펀치5mm 생검 펀치. </li><li> 플라즈마는 300 W 플라즈마 청소기에 60 초 동안 1 mbar에서의 O 2 플라즈마에 장치를 취급합니다. 일시적 PDMS-PDMS 본딩을 지연 및 윤활제 역할을 DI 물 방울 PDMS의 unpunched 조각의 표면을 적시고. 마스터 2면에 스테레오 현미경, 장소 마스터 1을 통해보기 및 기계식 잠금 장치가 달성 될 때까지 상대적으로 표면을 밀어 동안 때 오목 프레임과 그림 3A 메이트에서 돌출 프레임. </li><li> 60 ° C의 오븐에서 장치를 놓고 물과 완전한 결합을 증발시켜 60 ℃에서 이틀 동안 조립 된 장치 (도 3b)을 굽는다. </li></ol> 시약 3. 준비 <ol><li> 내부 단계의 증류수로 1 ML의 주사기를 채우십시오. </li><li> 1 중량으로 HFE 7500 불소계 오일 1 ML의 주사기를 채우십시오. 중간 단계에 대한 % 생체 적합성 계면 활성제 계면 활성제 (13). </li><li> 10 중량으로 10 ML의 주사기를 채우십시오. % 폴리에틸렌1 중량 % 함유하는 수용액에서는 E 글리콜 (PEG). % 트윈 20과 1 중량. 연속 상 % 나트륨 도데 실 설페이트. </li></ol> 4. 시스템 준비 <ol><li> &lt;100 마이크로 초 셔터 속도를 낼 수있는 디지털 카메라와 결합 거꾸로 현미경의 무대에서 마이크로 유체 칩을 놓습니다. </li><li> 주사기 펌프에 모든 주사기를 탑재하고 27 G 바늘을 연결합니다. 바늘 ~ PE-2 튜브의 30cm 길이를 연결하고 해당 장치에 구멍을 천공으로 느슨한 끝을 삽입합니다. </li><li> 장치의 출력 포트에 PE-2의 10cm 길이를 삽입하고 폐기물 수집 용기에서 다른 쪽 끝을 배치합니다. </li><li> 프라임 튜브 세그먼트에서 유체의 속도까지 높은 속도로 시린지 펌프 (000 μL / 분)를 실행하여 장치는 장치의 흡입구에 도달한다. </li></ol> 5. 에멀젼 생성 <ol><li> 50 μm의 × 50 μm의 구멍과를 포함하는 지역에 현미경 초점하류 출구 채널. </li><li> 연속 상 250 μL / 내부 단계의 시간, 100 μL / 중간 단계의 시간, 700 μL / hr의 유량으로 이중 에멀젼 발생기에 유체를 제공하고 평균 10 분을 기다려야 주사기 펌프를 설정합니다. </li><li> 각각 250 μL / 시간에서 100 μL / 시간 내측 및 중간 단계의 유량을 유지한다. 1,050 μL / 시간 외측 위상의 유량을 설정한다. 이중 에멀젼 생성이 유동 조건의 설정에 따라 안정을 위해 3 ~ 5 분을 기다립니다. </li><li> 수동 이미지 분석을 통해 오프라인으로 처리를위한 30 Hz에서 영상 5 초를 취득. </li><li> 표 1에 주어진 유량과 5.3 및 5.4를 반복한다. 내측 및 중간 단계의 유량은 일정하게 유지되고 반송파 위상 유량 주사기 펌프의 설정을 조절함으로써 변화된다. </li></ol>

<ol>
	<li>Van Der Graaf, S., Schro&#235;n, C. G. P. H., Boom, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preparation+of+double+emulsions+by+membrane+emulsification+-+A+review.">Preparation of double emulsions by membrane emulsification - A review.</a> J. Membrane Sci. 251 (1-2), 7-15 (2005).</li><li>Laugel, C., Baillet, A. P., Youenang Piemi, M., Marty, J., Ferrier, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oil-water-oil+multiple+emulsions+for+prolonged+delivery+of+hydrocortisone+after+topical+application:+comparison+with+simple+emulsions.">Oil-water-oil multiple emulsions for prolonged delivery of hydrocortisone after topical application: comparison with simple emulsions.</a> Int. J. Pharm. 160 (1), 109-117 (1998).</li><li>Kim, S. H., Kim, J. W., Cho, J. C., Weitz, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Double-emulsion+drops+with+ultra-thin+shells+for+capsule+templates.">Double-emulsion drops with ultra-thin shells for capsule templates.</a> Lab Chip. 11 (18), 3162-3166 (2011).</li><li>Lim, S. W., Abate, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ultrahigh-throughput+sorting+of+microfluidic+drops+with+flow+cytometry.">Ultrahigh-throughput sorting of microfluidic drops with flow cytometry.</a> Lab Chip. 13 (23), 4563-4572 (2013).</li><li>Bernath, K., Hai, M., Mastrobattista, E., Griffiths, A. D., Magdassi, S., Tawfik, D. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+compartmentalization+by+double+emulsions:+sorting+and+gene+enrichment+by+fluorescence+activated+cell+sorting.">In vitro compartmentalization by double emulsions: sorting and gene enrichment by fluorescence activated cell sorting.</a> Anal. Biochem. 325 (1), 151-157 (2004).</li><li>Seemann, R., Brinkmann, M., Pfohl, T., Herminghaus, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Droplet+based+microfluidics.">Droplet based microfluidics.</a> Rep. Prog. Phys. 75 (1), 016601 (2012).</li><li>Bauer, W. A. C., Fischlechner, M., Abell, C., Huck, W. T. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hydrophilic+PDMS+microchannels+for+high-throughput+formation+of+oil-in-water+microdroplets+and+water-in-oil-in-water+double+emulsions.">Hydrophilic PDMS microchannels for high-throughput formation of oil-in-water microdroplets and water-in-oil-in-water double emulsions.</a> Lab Chip. 10 (14), 1814-1819 (2010).</li><li>Utada, A. S., Lorenceau, E., Link, D. R., Kaplan, P. D., Stone, H. A., Weitz, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Monodisperse+double+emulsions+generated+from+a+microcapillary+device.">Monodisperse double emulsions generated from a microcapillary device.</a> Science. 308 (5721), 537-541 (2005).</li><li>Chang, F. C., Su, Y. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Controlled+double+emulsification+utilizing+3D+PDMS+microchannels.">Controlled double emulsification utilizing 3D PDMS microchannels.</a> J. Micromech. Microeng. 18 (6), 065018 (2008).</li><li>Romanowsky, M. B., Abate, A. R., Rotem, A., Holtze, C., Weitz, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+throughput+production+of+single+core+double+emulsions+in+a+parallelized+microfluidic+device.">High throughput production of single core double emulsions in a parallelized microfluidic device.</a> Lab Chip. 12 (4), 802-807 (2012).</li><li>Tran, T. M., Cater, S., Abate, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Coaxial+flow+focusing+in+poly(dimethylsiloxane)+microfluidic+devices.">Coaxial flow focusing in poly(dimethylsiloxane) microfluidic devices.</a> Biomicrofluidics. 8 (1), 016502 (2014).</li><li>. Lithography Available from: <a target="_blank" href="https://www.memsnet.org/mems/processes/lithography.html">https://www.memsnet.org/mems/processes/lithography.html</a> (2015)</li><li>O'Donovan, B., Eastburn, D. J., Abate, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrode-free+picoinjection+of+microfluidic+drops.">Electrode-free picoinjection of microfluidic drops.</a> Lab Chip. 12 (20), 4029-4032 (2012).</li><li>Chang, F. C., Lin, H. H., Su, Y. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Controlled+W/O/W+double+emulsification+in+3-D+PDMS+micro-channels.">Controlled W/O/W double emulsification in 3-D PDMS micro-channels.</a> , 792-795 (2008).</li><li>Romanowsky, M. B., Abate, A. R., Rotem, A., Holtze, C., Weitz, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+throughput+production+of+single+core+double+emulsions+in+a+parallelized+microfluidic+device.">High throughput production of single core double emulsions in a parallelized microfluidic device.</a> Lab Chip. 12 (4), 802 (2012).</li></ol>

The authors have nothing to disclose.

여기에 설명 된 이중 에멀젼 발생 형상은 유리 모세관 장치 (8)의 물리를 모방하도록 설계된다. 이들에 정렬 원통형 유리 모세관 균일 이중 에멀젼 방울에 전단되는 삼상 동축 제트을 만드는 데 사용됩니다. 우리 3D PDMS 소자의 기능은 전체 높이가 320 μm의 아르 반송파 위상 채널과 50 μm의 높이로 형성 작은 ​​제작 기능의 중앙 정렬에 의존한다. 마스크가 정확히 정렬되지 않는 경우에 50 ?...

<html> 이중 에멀젼은 인해 산업, 의약품에서 사용 가능성, 및 생물학적 응용 1 중간, 비혼 화성 유체 층에 의해 담체 상으로부터 분리 액적로 구성하고, 특히 중요하다. 어떤 경우에는, 이중 에멀젼의 코어에 고 부가가치 화합물을 캡슐화 할 수있는 능력은 보호 물질과 제어 방식으로 방출 될 수있다. 예를 들어, 약물이 외부 캐리어 유체 (2)에 적합하지 않은 조건에서 용해도 캡슐화 될 수있다. 또한, 중간 유층 약물, 화장품 및 영양 3의 캡슐화 및 전송을위한 캡슐 템플릿으로 사용될 수있다. 그들은 서브 나노 리터 실험의 대규모 수가 다음, 실시 감지 (FACS) 악기 4,5 정렬 형광 활성화 된 셀을 사용하여 정렬 할 수 있기 때문에 생물학, 이중 에멀젼은 또한 높은 처리량 검사에 유용하다. ENT는 &quot;&gt; 원하는 성능 특성을 갖는 이중 에멀젼의 디자인은 이중 에멀젼 크기, 조성 및 균일도의 정밀한 제어를 필요로한다. 이러한 막 유화 같은 벌크 유화 공정은, 업계에서 사용되지만, 생성 된 에멀젼은 높은 다 분산되어 보이는 기능적 특성 (1)의 다양한. 액적 미세 유체 분야는 자연적으로 신중하게 제어 구성 6 단 분산 에멀젼의 생성을 적합하다. 미세 유체 이중 에멀젼 생성이 초점을 두 가지 주요 전략을 순차적 드롭 결정과 유리 모세관 흐름 달성되었다. 두 번 에멀젼 수 공정을 두 단계의 방울을 사용 평면 PDMS 장치에서 생성 될 수있다. 먼저, 수성 유 에멀젼이 유 중수 드롭하게 소수성 채널 벽을 가진 디바이스의 영역이있다. 다음을 사용하여 생성되며, 에멀젼 일 수있다 흐르는 물에 기름에 적합한 친수성 ​​벽 드롭 만드는 영역에 재 주입드롭 만들기 (4). 그러나, PMD를 친수성 표면 처리는 추가적인 제조 공정을 필요로하며 종종 임시 7이다. 이중 에멀젼을 형성하기 위해 대부분의 제어 및 반복 가능한 방법이 동축 흐름을 중심으로하고, 기술은 세 단계를 포함하는 동심 제트 단 분산 액적 (8)을 생성하기 위해 작은 오리피스를 통해 전단된다 유리 모세관 마이크로 유체를 사용 개척했다. 이 기술은 각상의 유속의 함수 인 이중 에멀젼의 정확한 크기 및 구성으로, 채널 치수보다 훨씬 작은 방울의 생산을 허용한다. 액적 크기 및 채널과 상기 보호 외피 흐름 사이의 큰 차이는, 채널 벽과 접촉 불필요한 표면 처리 렌더링로부터 방울을 방지한다. 그러나, 유리 장치는주의 조립 및 씰링과 함께, 테이퍼 모세관 팁 사용자 정의 제조를 필요로한다. 이전 연구자들은 3D 소프트 리소그래피를 사용했다그래피로 물리학을 중심으로 흐름을 사용하여 이중 에멀젼을 생성하지만, 이러한 장치는 직경&gt; 150 μm의 9, 10, 일반적으로 외과로 분류 개체보다 큰 크기의 약 주문 에멀젼을 생산했다. 매력적인 대안은 PDMS 소프트 리소그래피 제조의 용이성에 초점을 맞추고 유리 모세관 동축 흐름의 강력한 기능과 작은 방울 생성을 포함한다. 본 논문에서는 50 μm의 에멀젼 ≤ 생산 초점 동축 흐름을 사용하여 완전히 3D 소프트 리소그래피 (11)를 사용하여 구성되어 이중 에멀젼 발생기를 설명합니다. 우리는 디바이스 뽑아 유리 모세관 노즐 에멀젼 형성 공정을 근사하는 작은 전단 채널 (도 1)를 포함하는 장치를 제조 폴더 형 접근법을 사용한다. 더 중요하게, 이러한 장치는 특별한 표면 처리를 필요로하지 않고, 모든 중합체 구조는 간단하고 반복적 인 제조 SC드립니다중복 된 장치들의 많은 수 alable. 여기서는 더블 에멀젼 발생기의 설계, 제조 및 테스트를 개요. 더블 에멀젼 세대는 14 μm의의 물방울 직경에 이르기까지 강력하고 반복적으로 표시됩니다. 제조의 용이성과 기능의 결합은이 장치 새로운 더블 에멀젼 응용 프로그램의 개발을위한 매력적인 옵션이 있습니다.

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double emulsion generation using a polydimethylsiloxane (pdms) co-axial flow focus device

Double emulsions are useful in a number of biological and industrial applications in which it is important to have an aqueous carrier fluid. This paper presents a polydimethylsiloxane (PDMS) microfluidic device capable of generating water/oil/water double emulsions using a coaxial flow focusing geometry that can be fabricated entirely using soft lithography. Similar to emulsion devices using glass capillaries, double emulsions can be formed in channels with uniform wettability and with dimensions much smaller than the channel sizes. Three dimensional flow focusing geometry is achieved by casting a pair of PDMS slabs using two layer soft lithography, then mating the slabs together in a clamshell configuration. Complementary locking features molded into the PDMS slabs enable the accurate registration of features on each of the slab surfaces. Device testing demonstrates formation of double emulsions from 14 &#181;m to 50 &#181;m in diameter while using large channels that are robust against fouling and clogging.

이중 에멀젼 발생기 3D PDMS 제조를 (도 1a)를 사용하여 만든 공동 축류 집속 장치로 구성된다. 지오메트리는 삼상 동축 제트의 형성 물 / 오일 / 물 이중 에멀젼 (도 1b,도 1c)의 형성을 허용 광장 × 50 μm의 오리피스 50㎛의로 전단 될 수있다. 내부 수상과 유상 중간 10 ㎛, 채널 치수에 함께 접합하게된다 × 50 μm의 (도 1D,</...

Watch this Scientific Journal Video about Double Emulsion Generation Using a Polydimethylsiloxane PDMS Co-axial Flow Focus Device at JoVE.com

Double Emulsion Generation Using a Polydimethylsiloxane PDMS Co-axial Flow Focus Device

Microfluidic double emulsions generation typically involves devices with patterned wettability or custom-fabricated glass components. Here we describe the fabrication and testing of an all polydimethylsiloxane (PDMS) double emulsion generator that does not require surface treatment or complicated fabrication processes, and is capable of producing double emulsions down to 14 &#181;m.

폴리 디메틸 실록산 (PDMS) 공동 축류 초점 장치를 사용하여 이중 에멀젼 생성

Double emulsions are useful in a number of biological and industrial applications in which it is important to have an aqueous carrier fluid. This paper ...

double-emulsion-generation-using-polydimethylsiloxane-pdms-co-axial

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JoVE Journal

Bioengineering

Double Emulsion Generation Using a Polydimethylsiloxane (PDMS) Co-axial Flow Focus Device

15.9K 조회수.  University of California, San Francisco. Microfluidic double emulsions generation typically involves devices with patterned wettability or custom-fabricated glass components. Here we describe the fabrication and testing of an all polydimethylsiloxane (PDMS) double emulsion generator that does not require surface treatment or complicated fabrication processes, and is capable of producing double emulsions down to 14 µm.

비디오: Double Emulsion Generation Using a Polydimethylsiloxane PDMS Co-axial Flow Focus Device

A Novel Three-dimensional Flow Chamber Device to Study Chemokine-directed Extravasation of Cells Circulating under Physiological Flow Conditions

Extravasation of circulating cells from the bloodstream plays a central role in many physiological and pathophysiological processes, including stem cell homing and tumor metastasis. The three-dimensional flow chamber device (hereafter the 3D device) is a novel in vitro technology that recreates physiological shear stress and allows each step of the cell extravasation cascade to be quantified. The 3D device consists of an upper compartment in which the cells of interest circulate under shear stress, and a lower compartment of static wells that contain the chemoattractants of interest. The two compartments are separated by porous inserts coated with a monolayer of endothelial cells (EC). An optional second insert with microenvironmental cells of interest can be placed immediately beneath the EC layer. A gas exchange unit allows the optimal CO2 tension to be maintained and provides an access point to add or withdraw cells or compounds during the experiment. The test cells circulate in the upper compartment at the desired shear stress (flow rate) controlled by a peristaltic pump. At the end of the experiment, the circulating and migrated cells are collected for further analyses. The 3D device can be used to examine cell rolling on and adhesion to EC under shear stress, transmigration in response to chemokine gradients, resistance to shear stress, cluster formation, and cell survival. In addition, the optional second insert allows the effects of crosstalk between EC and microenvironmental cells to be examined. The translational applications of the 3D device include testing of drug candidates that target cell migration and predicting the in vivo behavior of cells after intravenous injection. Thus, the novel 3D device is a versatile and inexpensive tool to study the molecular mechanisms that mediate cellular extravasation.

The three-dimensional flow chamber device is a novel in vitro technology for the quantitative and step-wise evaluation of the extravasation cascade of cells circulating under conditions of physiological shear stress. The device therefore fills a critical need for basic, preclinical, and clinical studies of cell migration.

생리 흐름 조건에서 순환 세포의 케모카인 - 감독 넘쳐을 연구하는 소설 입체 흐름 챔버 장치

Extravasation of circulating cells from the bloodstream plays a central role in many physiological and pathophysiological processes, including stem cell ...

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PLGA Nanoparticles Formed by Single- or Double-emulsion with Vitamin E-TPGS

Poly(lactic-co-glycolic acid) (PLGA) is a biocompatible member of the aliphatic polyester family of biodegradable polymers. PLGA has long been a popular choice for drug delivery applications, particularly since it is already FDA-approved for use in humans in the form of resorbable sutures. Hydrophobic and hydrophilic drugs are encapsulated in PLGA particles via single- or double-emulsion. Briefly, the drug is dissolved with polymer or emulsified with polymer in an organic phase that is then emulsified with the aqueous phase. After the solvent has evaporated, particles are washed and collected via centrifugation for lyophilization and long term storage. PLGA degrades slowly via hydrolysis in aqueous environments, and encapsulated agents are released over a period of weeks to months. Although PLGA is a material that possesses many advantages for drug delivery, reproducible formation of nanoparticles can be challenging; considerable variability is introduced by the use of different equipment, reagents batch, and precise method of emulsification. Here, we describe in great detail the formation and characterization of microparticles and nanoparticles formed by single- or double-emulsion using the emulsifying agent vitamin E-TPGS. Particle morphology and size are determined with scanning electron microscopy (SEM). We provide representative SEM images for nanoparticles produced with varying emulsifier concentration, as well as examples of imaging artifacts and failed emulsifications. This protocol can be readily adapted to use alternative emulsifiers (e.g. poly(vinyl alcohol), PVA) or solvents (e.g.&#160;dichloromethane, DCM).

We describe the production and characterization of nanoparticles and microparticles composed of poly(lactic-co-glycolic acid) using vitamin E-TPGS as an emulsifier. By varying formulation parameters such as the concentration of emulsifier, it is possible to produce nanoparticles with mean diameters ranging from 220 nm to&#160;1.98 &#181;m.

Poly(lactic-co-glycolic acid) (PLGA) is a biocompatible member of the aliphatic polyester family of biodegradable polymers. PLGA has long been a popular ...

Stretching Micropatterned Cells on a PDMS Membrane

Mechanical forces exerted on cells and/or tissues play a major role in numerous processes. We have developed a device to stretch cells plated on a PolyDiMethylSiloxane (PDMS) membrane, compatible with imaging. This technique is reproducible and versatile. The PDMS membrane can be micropatterned in order to confine cells or tissues to a specific geometry. The first step is to print micropatterns onto the PDMS membrane with a deep UV technique. The PDMS membrane is then mounted on a mechanical stretcher. A chamber is bound on top of the membrane with biocompatible grease to allow gliding during the stretch. The cells are seeded and allowed to spread for several hours on the micropatterns. The sample can be stretched and unstretched multiple times with the use of a micrometric screw. It takes less than a minute to apply the stretch to its full extent (around 30%). The technique presented here does not include a motorized device, which is necessary for applying repeated stretch cycles quickly and/or computer controlled stretching, but this can be implemented. Stretching of cells or tissue can be of interest for questions related to cell forces, cell response to mechanical stress or tissue morphogenesis. This video presentation will show how to avoid typical problems that might arise when doing this type of seemingly simple experiment.

This manuscript presents a technique to apply or release forces on adherent cells or tissues using unidirectional stretching.

PDMS 막에 마이크로 패턴 세포를 스트레칭

Mechanical forces exerted on cells and/or tissues  play a major role in numerous processes. We have developed a device to stretch  cells plated on a ...

Lipid Bilayer Vesicle Generation Using Microfluidic Jetting

Bottom-up synthetic biology presents a novel approach for investigating and reconstituting biochemical systems and, potentially, minimal organisms. This emerging field engages engineers, chemists, biologists, and physicists to design and assemble basic biological components into complex, functioning systems from the bottom up. Such bottom-up systems could lead to the development of artificial cells for fundamental biological inquiries and innovative therapies1,2. Giant unilamellar vesicles (GUVs) can serve as a model platform for synthetic biology due to their cell-like membrane structure and size. Microfluidic jetting, or microjetting, is a technique that allows for the generation of GUVs with controlled size, membrane composition, transmembrane protein incorporation, and encapsulation3. The basic principle of this method is the use of multiple, high-frequency fluid pulses generated by a piezo-actuated inkjet device to deform a suspended lipid bilayer into a GUV. The process is akin to blowing soap bubbles from a soap film. By varying the composition of the jetted solution, the composition of the encompassing solution, and/or the components included in the bilayer, researchers can apply this technique to create customized vesicles. This paper describes the procedure to generate simple vesicles from a droplet interface bilayer by microjetting.

Microfluidic jetting against a droplet interface lipid bilayer provides a reliable way to generate vesicles with control over membrane asymmetry, incorporation of transmembrane proteins, and encapsulation of material. This technique can be applied to study a variety of biological systems where compartmentalized biomolecules are desired.

미세 유체 제트를 사용하여 지질 이중층 소포 세대

Bottom-up synthetic biology presents a novel approach for investigating and reconstituting biochemical systems and, potentially, minimal organisms. This ...

Automated Lipid Bilayer Membrane Formation Using a Polydimethylsiloxane Thin Film

An artificial lipid bilayer, or black lipid membrane (BLM), is a powerful tool for studying ion channels and protein interactions, as well as for biosensor applications. However, conventional BLM formation techniques have several drawbacks and they often require specific expertise and laborious processes. In particular, conventional BLMs suffer from low formation success rates and inconsistent membrane formation time. Here, we demonstrate a storable and transportable BLM formation system with controlled thinning-out time and enhanced BLM formation rate by replacing conventionally used films (polytetrafluoroethylene, polyoxymethylene, polystyrene) to polydimethylsiloxane (PDMS). In this experiment, a porous-structured polymer such as PDMS thin film is used. In addition, as opposed to conventionally used solvents with low viscosity, the use of squalene permitted a controlled thinning-out time via slow solvent absorption by PDMS, prolonging membrane lifetime. In addition, by using a mixture of squalene and hexadecane, the freezing point of the lipid solution was increased (~16 &#176;C), in addition, membrane precursors were produced that can be indefinitely stored and readily transported. These membrane precursors have reduced BLM formation time of &#60; 1 hr and achieved a BLM formation rate of ~80%. Moreover, ion channel experiments with gramicidin A demonstrated the feasibility of the membrane system.

We demonstrate a storable, transportable lipid bilayer formation system. A lipid bilayer membrane can be formed within 1 hr with over 80% success rate when a frozen membrane precursor is brought to ambient temperature. This system will reduce laborious processes and expertise associated with ion channels.

폴리 디메틸 실록산 박막을 사용하여 자동화 지질 이중층 제막

An artificial lipid bilayer, or black lipid membrane (BLM), is a powerful tool for studying ion channels and protein interactions, as well as for ...

A Microcontroller Operated Device for the Generation of Liquid Extracts from Conventional Cigarette Smoke and Electronic Cigarette Aerosol

Electronic cigarettes are the most popular tobacco product among middle and high schoolers and are the most popular alternative tobacco product among adults. High quality, reproducible research on the consequences of electronic cigarette use is essential for understanding emerging public health concerns and crafting evidence based regulatory policy. While a growing number of papers discuss electronic cigarettes, there is little consistency in methods across groups and very little consensus on results. Here, we describe a programmable laboratory device that can be used to create extracts of conventional cigarette smoke and electronic cigarette aerosol. This protocol details instructions for the assembly and operation of said device, and demonstrates the use of the generated extract in two sample applications: an in vitro cell viability assay and gas-chromatography mass-spectrometry. This method provides a tool for making direct comparisons between conventional cigarettes and electronic cigarettes, and is an accessible entry point into electronic cigarette research.

Here, we describe a programmable laboratory device that can be used to create extracts of conventional cigarette smoke and electronic cigarette aerosol. This method provides a useful tool for making direct comparisons between conventional cigarettes and electronic cigarettes, and is an accessible entry point into electronic cigarette research.

전통적인 담배 연기와 전자 담배 연 무질에서 액체 추출의 세대에 대 한 장치를 운영 하는 마이크로컨트롤러

Electronic cigarettes are the most popular tobacco product among middle and high schoolers and are the most popular alternative tobacco product among ...

Immobilization of Live Caenorhabditis elegans Individuals Using an Ultra-thin Polydimethylsiloxane Microfluidic Chip with Water Retention

Radiation is widely used for biological applications and for ion-beam breeding, and among these methods, microbeam irradiation represents a powerful means of identifying radiosensitive sites in living organisms. This paper describes a series of on-chip immobilization methods developed for the targeted microbeam irradiation of live individuals of Caenorhabditis elegans. Notably, the treatment of the polydimethylsiloxane (PDMS) microfluidic chips that we previously developed to immobilize C. elegans individuals without the need for anesthesia is explained in detail. This chip, referred to as a worm sheet, is resilient to allow the microfluidic channels to be expanded, and the elasticity allows animals to be enveloped gently. Also, owing to the self-adsorption capacity of the PDMS, animals can be sealed in the channels by covering the surface of the worm sheet with a thin cover film, in which animals are not pushed into the channels for enclosure. By turning the cover film over, we can easily collect the animals. Furthermore, the worm sheet shows water retention and allows C. elegans individuals to be subjected to microscopic observation for long periods under live conditions. In addition, the sheet is only 300 &#181;m thick, allowing heavy ions such as carbon ions to pass through the sheet enclosing the animals, thus allowing the ion particles to be detected and the applied radiation dose to be measured accurately. Because selection of the cover films used for enclosing the animals is very important for successful long-term immobilization, we conducted the selection of the suitable cover films and showed a recommended one among some films. As an application example of the chip, we introduced imaging observation of muscular activities of animals enclosing the microfluidic channel of the worm sheet, as well as the microbeam irradiation. These examples indicate that the worm sheets have greatly expanded the possibilities for biological experiments.

Immobilization of Live Caenorhabditis elegans Individuals Using an Ultra-thin Polydimethylsiloxane Microfluidic Chip with Water Retention

A series of immobilization methods has been established to allow the targeted irradiation of live Caenorhabditis elegans individuals using a recently developed ultra-thin polydimethylsiloxane microfluidic chip with water retention. This novel on-chip immobilization is also adequate for imaging observations. The detailed treatment and application examples of the chip are explained.

물 유지와 울트라입니다 미세 칩을 사용 하 여 라이브 꼬마 선 충 의 동원 정지

Radiation is widely used for biological applications and for ion-beam breeding, and among these methods, microbeam irradiation represents a powerful means ...

Generation of Transgenic Rats using a Lentiviral Vector Approach

Transgenic animal models are fundamentally important for modern biomedical research. The incorporation of foreign genes into early mouse or rat embryos is an invaluable tool for gene function analysis in living organisms. The standard transgenesis method is based on microinjecting foreign DNA fragments into a pronucleus of a fertilized oocyte. This technique is widely used in mice but remains relatively inefficient and technically demanding in other animal species. The transgene can also be introduced into one-cell-stage embryos via lentiviral infection, providing an effective alternative to standard pronuclear injections, especially in species or strains with a more challenging embryo structure. In this approach, a suspension that contains lentiviral vectors is injected into the perivitelline space of a fertilized rat embryo, which is technically less demanding and has a higher success rate. Lentiviral vectors were shown to efficiently incorporate the transgene into the genome to determine the generation of stable transgenic lines. Despite some limitations (e.g., Biosafety Level 2 requirements, DNA fragment size limits), lentiviral transgenesis is a rapid and efficient transgenesis method. Additionally, using female rats that are mated with a fertile male strain with a different dominant fur color is presented as an alternative to generate pseudopregnant foster mothers.

This article aims to provide the methodology for lentiviral transgenesis in rat embryos using multiple injections of a virus suspension into the zygote perivitelline space. Female rats that are mated with a fertile male strain with a different dominant fur color is used to generate pseudopregnant foster mothers.

렌티바이러스 벡터 접근법을 이용한 형질전환 쥐 생성

Transgenic animal models are fundamentally important for modern biomedical research. The incorporation of foreign genes into early mouse or rat embryos is ...

Generation of Dynamical Environmental Conditions using a High-Throughput Microfluidic Device

Mimicking in vivo environmental conditions is crucial for in vitro studies on complex life machinery. However, current techniques targeting live cells and organs are either highly expensive, like robotics, or lack nanoliter volume and millisecond time accuracy in liquid manipulation. We herein present the design and fabrication of a microfluidic system, which consists of 1,500 culture units, an array of enhanced peristaltic pumps and an on-site mixing modulus. To demonstrate the capacities of the microfluidic device, neural stem cell (NSC) spheres are maintained in the proposed system. We observed that when the NSC sphere is exposed to CXCL in day 1 and EGF in day 2, the round-shaped conformation is well maintained. Variation in the input order of 6 drugs causes morphological changes to the NSC sphere and the expression level representative marker for NSC stemness (i.e., Hes5 and Dcx). These results indicate that dynamic and complex environmental conditions have great effects on NSC differentiation and self-renewal, and the proposed microfluidic device is a suitable platform for high throughput studies on the complex life machinery.

We present a microfluidic system for high throughput studies on complex life machinery, which consists of 1500 culture units, an array of enhanced peristaltic pumps and an on-site mixing modulus. The microfluidic chip allows for the analysis of the highly complex and dynamic micro-environmental conditions in vivo.

고처리량 미세 유체 장치를 사용하여 동적 환경 조건의 생성

Mimicking in vivo environmental conditions is crucial for in vitro studies on complex life machinery. However, current techniques targeting live cells and ...

Contrast-Enhanced Subharmonic Aided Pressure Estimation (SHAPE) Using Ultrasound Imaging with a Focus on Identifying Portal Hypertension

Noninvasive, accurate measurement of pressures within the human body has long been an important but elusive clinical goal. Contrast agents for ultrasound imaging are gas-filled, encapsulated microbubbles (diameter &#60; 10 &#956;m) that traverse the entire vasculature and enhance signals by up to 30 dB. These microbubbles also produce nonlinear oscillations at frequencies ranging from the subharmonic (half of the transmit frequency) to higher harmonics. The subharmonic amplitude has an inverse linear relationship with the ambient hydrostatic pressure. Here an ultrasound system capable of performing real-time, subharmonic aided pressure estimation (SHAPE) is presented. During ultrasound contrast agent infusion, an algorithm for optimizing acoustic outputs is activated. Following this calibration, subharmonic microbubble signals (i.e., SHAPE) have the highest sensitivity to pressure changes and can be used to noninvasively quantify pressure. The utility of the SHAPE procedure for identifying portal hypertension in the liver is the emphasis here, but the technique has applicability across many clinical scenarios.

Contrast-Enhanced Subharmonic Aided Pressure Estimation SHAPE Using Ultrasound Imaging with a Focus on Identifying Portal Hypertension

A protocol for noninvasively estimating ambient pressures utilizing subharmonic ultrasound imaging of infused contrast microbubbles (following appropriate calibration) is described with examples from human patients with chronic liver disease.

문맥 고혈압 식별에 중점을 둔 초음파 영상을 사용한 조영 강화 저고조파 보조 압력 추정(SHAPE)

Noninvasive, accurate measurement of pressures within the human body has long been an important but elusive clinical goal. Contrast agents for ultrasound ...