This work was partly supported by NSF CBET 1227766, NSF CBET 1511759, and Byars-Tarnay Endowment. We gratefully acknowledge use of the West Virginia University Shared Research Facilities which are supported, in part, by NSF EPS-1003907.

EBL 금형에서 PDMS 금형 1. 복제 <ol><li> (29) 실리콘 몰드를 제조 <ol><li> 스핀 코팅은 200 ㎕의 폴리 메틸 메타 크릴 레이트 (PMMA) 1 분 동안 2,500 rpm에서 1 × 1cm의 실리콘 (Si) 기판 상에 용액의 박막을 형성한다. </li><li> 2 분 동안 180 ° C에서 Si 기판의 PMMA 필름을 굽는다. </li><li> 300 μC / ㎠의 면적 도즈 집속 전자 빔을 이용하여 PMMA 필름의 설계 나노 패턴을 작성한다. </li><li> 80 초 동안 개발자의 PMMA의 나노 패턴을 개발합니다. </li><li> 10 kV로, 0.5 mA의 방출 전류는 0.5 Å / sec의 증착 속도의 출력 전압 전자빔 증착기를 이용하여 두께 50 nm의 니켈 층과 PMMA의 나노 패턴을 증착. </li><li> 20 분 동안 80 ° C에서 20 ㎖ 제거제의 PMMA 부 리프트 오프. </li><li> 반응성 이온 에칭 (RIE) Si 기판에 나노 패턴은 원하는 깊이의 실리콘 몰드 도착. 참고 : tetr의 가스 혼합물afluoromethane (CF 4) 150 W 400 W와 RIE 전력의 유도 결합 플라즈마 (ICP) 전원으로 / 산소 (O 2) (90 % / 10 %)이 560 nm의 깊이까지 실리콘 기판을 에칭하기 위해 사용된다. </li></ol></li><li> silanization합니다 실리콘 몰드 <ol><li> 100mm의 PS 페트리 접시에서 유리 커버 슬립하고 실리콘 주형을 넣어 흄 후드에 위치한 유리 데시 케이 터 내에서 전송할. </li><li> 커버 슬립에 10 μL의 1H, 1H, 2H, 2H-Perfluorooctyltrichlorosilane을 삭제합니다. 주의 : 1H는, 1H는, 2H, 2H-Perfluorooctyltrichlorosilane 피부 부식 및 눈에 심한 손상을 일으킬 수 있습니다. 적절한 개인 보호 장비 (PPE)를 착용 할 것. </li><li> 부분적으로 페트리 접시를 커버. </li><li> Si를 몰드의 실란 화를 완료하는 흄 후드에서 5 시간 동안 진공하에 데시 케이 유지. </li></ol></li><li> PDMS 예비 중합체를 준비 <ol><li> 일회용 계량 보트 10 g의 PDMS 수지 1.05 g 경화제를 단다. </li><li> 플라스틱 숟가락을 사용하여 철저하게 PDMS 예비 중합체를 섞는다. </li><li> 맑은 혼합물이 관찰 될 때까지 약 20 분 동안 진공 하에서 플라스틱 건조기에서 PDMS의 예비 중합체를 탈기. </li></ol></li><li> PDMS 몰드 복제 <ol><li> 60mm 페트리 접시에 실란 화시 금형을 넣습니다. </li><li> 페트리 접시의시 금형에 PDMS 예비 중합체를 따르십시오. </li><li> 모든 거품이 사라질 때까지 약 10 분 동안 플라스틱 건조기 및 탈기의 페트리 접시를 놓습니다. </li><li> 열판을 페트리 접시로 이동시켜, 4 시간 동안 70 ° C에서 PDMS의 예비 중합체를 경화. </li><li> 신중 핀셋을 이용하여 Si를 몰드에서 PDMS 몰드를 떼어. 참고 : PDMS 금형은 일주까지 주위 조건에서 저장 될 수있다. 경화 후, PDMS 몰드 (30)의 일부 미가 교 PDMS 수지 분자 잔류 경화제가있다. 저 분자량 분자들은 점차 확산 시간 동안 표면에 축적된다. 이 PDMS 표면 (31)의 지형 및 기계적 특성에 영향을 미친다. DIF융합 1 주일 내 중요하지 않습니다. </li></ol></li></ol> 대형, 단일 금형에 PDMS 금형 2. 바느질 <ol><li> 단계 1.4을 반복하여 여러 PDMS 금형을 준비합니다. 참고 : PDMS 혼합물의 동일한 금액을 동일한 두께의 PDMS 금형을 얻을 때마다 체중. </li><li> 이러한 광학 현미경으로 nanogratings 이방성 PDMS 나노 패턴의 방향을 결정하고, 마커 펜으로 PDMS 몰드의 뒷면에 마크. 참고 : 이러한 나노 기둥으로 등방성 나노 형상의 방향을 표시 할 필요가 없습니다. </li><li> 흄 후드에서 에탄올 Si 기판을 청소하고 압축 공기로 건조. </li><li> 블레이드 각 PDMS 몰드의 패턴 화되지 않은 영역을 낸다. 참고 : 스티치 금형의 둘레에 배치되는 PDMS 금형의 경우, 다른 사람과의 접촉 만이 패턴 화되지 않은 영역이 떨어져 손질해야한다. </li><li> 에 나노 패턴면이 아래로 트리밍 된 PDMS 몰드를 배치미러 Si 기판의면 다음은 가까이 있지만 주변 금형 (들)을 터치하지 다른 금형을 맞 춥니 다. </li><li> PDMS 접착층 준비 <ol><li> 0.5 mm 두께의 층을 형성하기 위해 깨끗한 유리 슬라이드 (2.5 cm × 7.5 cm)에 1g는 PDMS 프리폴리머 (1.05 : 10 PDMS 수지와 경화제의 비) 탈기 전송. </li><li> 3-5 분 동안 핫 플레이트상에서 100 ℃에서 PDMS 층 굽는다. 레이어를 터치 레이어가 부분적으로하지만 완치는 것을 보장하기 위해 바늘을 사용합니다. 참고 : 부분적으로 경화 된 PDMS는 경화 PDMS 예비 중합체처럼 흐를 수 없지만 경화 PDMS에 비해 스티커입니다. </li></ol></li><li> 정렬 PDMS 몰드의 뒷면에 PDMS 층을 배치하고 신속하게 조립이 반전하고, 핫 플레이트로 옮긴다. </li><li> PDMS의 접착제 층 및 PDMS 몰드의 배면 사이의 양호한 접촉을 보장하고, 1 시간 동안 100 ° C에서 PDMS를 접착제 층을 경화 조립체의 상부에 금속 블록을 사용 압축력 (5 kPa로)를 적용한다. &lt;BR /&gt; 주 : 조심 조립체의 경사를 방지하기 위해 상기 금속 블록의 위치를 ​​조정한다. </li><li> 금속 블록을 제거하고 실리콘 기판의 단일 스티치 PDMS 몰드를 떼어. </li></ol> PS 기판에 마스터 몰드의 3 세대 주 : 유리 슬라이드에 고정화 스티칭 PDMS 몰드가 PS 판 또는 작동 나노 패턴 기판을 제조 할 수있는에서 PS 박막의 마스터 몰드를 생성하는데 사용될 수있다. <ol><li> 이 PS 판에 마스터 몰드를 생성 <ol><li> 이 PS 판을 준비 <ol><li> 이틀 동안 80 ℃에서 진공 오븐에서 PS 펠렛을 건조. </li><li> 230 ° C에서 프레스 기계를 예열. </li><li> 아래에서 위로 순서대로 알루미늄 판, 폴리 테트라 플루오로 에틸렌 (PTFE) 시트와 알루미늄 스페이서를 조립한다. </li><li> 3cm (L) × 3 cm (W) × 0.3 cm (H)의 사각형 구멍에 알루미늄 스페이서에서로드 3.5 g의 PS 펠릿. 참고 : 스페이서는 약 0입니다.1 PDMS 몰드보다 두꺼운 cm, 따라서 최종 나노 패턴의 PS 기판은 약 0.1 cm의 두께이다. </li><li> 알루미늄 스페이서의 다른 PTFE 시트 다음 다른 알루미늄 판을 놓습니다. </li><li> 프레스 기계 어셈블리를 놓습니다. </li><li> 30 분 동안 230 ° C에서 PS 펠렛을 예열. </li><li> 5 분 동안 어셈블리에 압축 압력 (0.1 MPa의)를 적용합니다. </li><li> 압력을 해제 한 후 조립 0.5 MPa의 압축 압력을 다시 적용합니다. </li><li> 1.5 MPa의 압력의 바람직한까지, 0.5 MPa의 압력 증가 단계를 반복 3.1.1.9 진행한다. </li><li> 프레스 기계의 히터를 끄고 1.5 MPa의 일정한 압력에서 70 ° C 이하로 냉각. </li><li> 어셈블리를 타고 PS 판을 다시 들어가는 습기를 방지하기 위해 80 ℃의 진공 오븐에서 PS 판을 저장한다. </li></ol></li><li> PS 판에 나노 임프린트 스티칭 PDMS 몰드 <ol><li> 알루미늄 스페이서의 PS 판을 놓고3 인치 실리콘 웨이퍼에 설정합니다. 주 : 상기 스페이서의 내부 치수는 PS 판과 동일하다 그래서 PS 판 바로 스페이서 맞는다. </li><li> 30 분 동안 250 ° C에서 열판에 PS 판을 가열한다. </li><li> 용융 PS 판에 나노 패턴면이 아래로 스티치 PDMS 몰드를 넣습니다. 주 : PDMS 몰드의 일측 먼저 PS 판의 표면과 접촉에 배치되고, 다른 쪽의 계면에 기포의 형성을 방지하기 위해 PS 표면과 접촉하여 서서히 저하된다. 주의 : 핫 플레이트의 표면이 뜨겁습니다. 나노 임 프린팅 과정에서 thermogloves을 착용 할 것. </li><li> 스티칭 PDMS 금형의 유리 슬라이드에 알루미늄 판을 놓습니다. </li><li> 알루미늄 판에 금속 블록을 사용하여 압축 압력 (12.5 kPa의)을 적용하고 3 분 동안 기다립니다. 주 : 알루미늄 플레이트가 기울어지지되어 있는지 확인합니다. </li><li> 들어 올려 알루미늄 판에서 금속 블록을 교체하고 난25 kPa의에 압축 압력을 ncrease. </li><li> 압력과 단계를 반복 3.1.2.6는 kPa의 50로 증가. 주의 :이 단계는 PDMS 몰드와 PS 판 사이에 포획 된 공기를 제거하는 것이다. </li><li> 15 분 동안 50 kPa로의 일관된 압력을 받고 240, 250 °의 C 사이의 열판의 온도를 유지한다. </li><li> 열판의 전원을 끄고 전체 설치를 냉각. 주 : 팬은 냉각 과정을 촉진하기 위해 사용될 수있다. </li><li> 의 PS 판에서 스티치 PDMS 몰드 박리 신중 온도가 50 ° C 이하로 된 후, 금속 블록을 제거. 주 : PS 기판 역방향 나노 패턴을 가지고 작업 PDMS 기판을 제조하여 마스터 몰드로서 사용될 수있다. </li></ol></li></ol></li><li> 이 PS 박막에 마스터 몰드를 생성 <ol><li> 에 PS 박막을 준비 <ol><li> 흄 후드에서 10 ml의 톨루엔에 1g의 PS를 녹여. 주의 : 톨루엔은 피부 irritatio가 발생할 수 있습니다n 및 심각한 눈 손상, 그리고 장기간 또는 반복 노출되면 장기에 손상을 줄 수 있습니다. 적절한 개인 보호구를 착용 할 것. </li><li> 1 분 동안 2,500 rpm에서 웨이퍼 2 인에 스핀 코트 1 ML의 PS 솔루션은 ~ 1 μm의 두께 PS 박막을 형성한다. </li><li> 3 일간 흄 후드에서 Si 웨이퍼에 PS 필름을 설정하여, 필름으로부터 톨루엔을 증발시켰다. </li><li> 밤새 80 ℃에서 진공 오븐에서 PS 박막을 어닐링. </li></ol></li><li> 나노 임프린트는 PS 박막의 PDMS 몰드 <ol><li> 열판에 설정되어있는 PS 박막에 나노 형상면이 아래로 스티치 PDMS 몰드를 넣습니다. </li><li> PDMS의 주형의 유리 측에 금속 블록을 사용하여 PDMS 몰드 12 kPa로의 압축 압력을 적용한다. </li><li> 180 ° C에 열판의 온도를 높이고 15 분 정도 유지한다. 주의 : 용융 PS 필름 윤활제로서 기능 할 수있다. 미끄러에서 금속 블록 않도록주의. </li><li> 열판의 전원을 끄고전체 설치를 냉각. 주 : 팬은 냉각 과정을 촉진하기 위해 사용될 수있다. </li><li> 의 PS 필름으로부터 스티치 PDMS 몰드 박리 신중 온도가 50 ° C 이하로 떨어지면 후 금속 블록을 제거. 참고 : 나노 패턴 PS 필름 작업 PDMS 기판을 생산하는 마스터 몰드 될 것입니다. </li></ol></li></ol></li></ol> 세포 행동 4. Nanotopographical 변조 참고 : 대표 nanotopographies 셀 확산의 nanotopographical 변조를 설명하기에 인간의 상피 세포가 배양. <ol><li> 애플리케이션에 따라 3.1 또는 3.2 단계 중 하나에서 생성 된 마스터 주형으로부터 작업 PDMS 기판을 전송. </li><li> 중공 강철 아치 펀치를 이용하여, 특정 멀티 웰 플레이트 (예를 들면, 24- 웰 플레이트)의 구성에 맞도록 디스크에 나노 패턴 PDMS 기판을 절단. </li><li> 오 우물에 PDMS 디스크를 배치 핀셋을 사용하여FA 멀티 웰 플레이트. </li><li> 70 % 에탄올 다음 UV 노출 30 분 동안 각을 이용하여 PDMS 기판을 소독. </li><li> 1 배 멸균 인산 완충 식염수 (PBS)를 세 번으로 PDMS 기판을 세척 할 것. </li><li> 코팅을 실온에서 30 분 동안 세포 외 기질 단백질 (즉, 20 μg의 / ml의 피브로넥틴)와 PDMS 기판. </li><li> 5 분 동안 PDMS 기판을 멸균 PBS로 3 회, 각 린스. </li><li> 10 % 소 태아 혈청 둘 베코 변형 이글 배지에 인간 폐암 세포 A549를 중단하고 혈구 세포를 사용하여 계산. </li><li> 하루 동안 5 % 이산화탄소를 함유하는 가습 분위기하에 37 ℃에서 PDMS 기판 및 배양을 2,000 세포 / ㎠의 밀도로 시딩 세포 접시. </li><li> PBS로 3 회 세포를 씻으십시오. </li><li> 4 시간 동안 PBS 중의 4 % 파라 포름 알데히드 및 2 % 글루 타르 알데히드의 혼합물에 세포를 고정 및 CO 2 임계 포를 사용하여 세포를 탈수전자 현미경 관찰 (29)를 스캔 INT 건조기. 주의 : 파라 포름 알데히드와 글루 타르 알데히드는 피부에 심한 화상과 눈에 손상을 줄 수 있습니다. 화학 후드에서 작동하고 적절한 PPE를 착용하십시오. </li></ol>

<ol>
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The authors have nothing to disclose.

우리는 나노 패턴 기판의 넓은 영역을 생성하기 위해, 단순 저렴하면서도 다목적 방법을 제시한다. 충실하게 높은 정의 나노 패턴을 확장하려면 큰 관심은 몇 가지 중요한 단계에 지불해야한다. 첫 번째 다중 PDMS 금형을 트리밍하는 것이다. PDMS의 주형의 패터닝되지 않은 부분을 제거 할 필요가있다. 또한, 주형의 측벽 금형 사이의 갭을 최소화하기 위해 가능한 한 수직으로 완벽 절단한다. 총체적 최종 스티치 몰드에 비 패턴 영역의 일부가 감소 될 수있다. 둘째,이 PDMS 몰드의 나노 패턴 표면은 실리콘 기판 상에 변형이없는 정렬 될 필요가있다. PDMS의 나노 구조물을 변형하는 경향이 있기 때문에합니다 (PDMS 몰드와 실리콘 표면 사이의 공기를 포획하지 않도록) 부드럽고 균일하게 실리콘 기판의 거울면에 대하여 나노 패턴 표면을 배치하는 것이 중요하다. PDMS 몰드가 있지만 네이 닿지 않으면 서 최대한 가까이 정렬 될 것이다주형 ghboring하면 상기 최종 스티치 주형의 패터닝되지 않은 부분을 최소화한다. 그렇지 않으면, 터치 나노 구조는 나노 임 프린팅 동안 변형됩니다. 셋째, PDMS 몰드의 두께는 생산 로트에 따라 달라질 수 있으며, 따라서 완벽 PDMS 주조 전에 작은 실리콘 몰드 레벨링 각 몰드 균일의 두께를 결정하는 것 외에도, 두께를 균일하게하는 것이 중요하다. PDMS의 금형에 걸쳐 막 두께의 변화는 유리 슬라이드 상에 캐스팅 층 PDMS 프리폴리머 (접착제)의 두께를 조정함으로써 보상 될 수 있지만, 두꺼운 예비 중합체 층을 문제가 될 수있다. 예비 중합체는 모세관 력으로 패턴 화 된 표면에 PDMS 몰드 사이의 틈을 통해 뽑아 결과적으로 나노 패턴을 손상시킬 수있다. 두께 변화는 EBL 몰드에서 전송할 때 PDMS 혼합물을 동일한 양을 제조함으로써 최소화 될 수있다. 결과적으로, 얇은 프리폴리머 PDMS 층을 사용할 수있다. 대안 적으로, 부분적으로 경화 prepolymeR 층의 점도를 증가시키고 따라서 그 인상을 줄이고 결국 나노 패턴 표면의 손상을 제거한다. 스티치 기술은 PDMS의 탄성 특성에 의해 제한된다. 소프트 리소그래피 원칙적으로 0.5 미만 내지 (18)의 해상도에 도달 할 수있는 작은 32 내지 2로 및 피처 크기를 복제 적용 하였지만 높이 대 폭의 종횡비가 너무되면 나노 PDMS 기능을 완벽하게 복제 될 수없는 높은 (&gt; 2) 또는 (0.2 &lt;) 너무 낮습니다. nanofeatures 종횡비가 너무 높은 경우 축소 또는 &lt;0.2 비율의 PDMS 스탬프 (33)를 사용할 경우 충분한 완화 될 수있다. 또한, 여러 PDMS 금형 원활 때문에 틈새와 PDMS 금형의 불완전 트리밍의 스티치 할 수 없습니다, 따라서 (예 : nanogratings 등 지속적인 나노 패턴에 대한 특히) 비 패턴과 잘못 정렬 영역이 있습니다. 불량의 작은 비율을 감안할 때영역 전체 표면적 위에 스티치 기법은 여전히 ​​나노 패턴 기판의 넓은 면적을 생성하는 간단하고 저렴한 방법을 제공한다. 또한, 스티치 금형 폴리머 기판으로 nanoimprinted하면, 용융 중합체는 요철면 (도 1a), 그 결과 간극으로 흘러 것이다. 요철면은 도전 세포 또는 분자 생물학적 분석을 위해 샘플을 수집 할 수있다. 마이크로은 패턴 화 된 기판에 밀봉 할 때 미세 유체 응용 프로그램에서, 인상은 불완전한 밀봉됩니다. 요철의 문제는 용이하게 막 두께를 조정 통해 인상을 최소화 중합체 박막 기술을 적용함으로써 해결 될 수있다 (도 1B). 스티치 기술은 확장 정의 마스터 몰드를 필요로하지만, 이러한 스텝 앤드 플래시 리소그래피 및 롤 - 투 - 롤 lithogra 나노 임 프린팅과 같은 다른 기술에 비해 간단하고 저렴PHY. 스티치 기법은 뜨거운 접시와 스티치 나노 임 프린팅 과정에서 압축 힘을 발휘하는 방법,하지만 고가의 장비 만이 필요합니다. 또한, 스티치 처리는 필수적인 것은 아니지만 크린룸의 청정 환경에서 수행 될 수있다. 스티치 기술은 다용도. 큰 면적에 동일한 나노 패턴을 확대하는 것 외에도, 스티치 방법은 다양한 형태, 크기 및 배열의 ​​마이크로 - 및 / 또는 나노 스케일 특징 이루어지는 금형에 적용 할 수있다. 이 점까지, 마이크로 / nanotopographies의 조합 적 라이브러리 셀 지형 상호 작용을 조사하기 위해 높은 처리량 플랫폼을 제공하기 위해 구축 될 수있다. 이 간단한 저렴하고 다용도 스티치 기술은 잠재적으로 복합 성분과 마이크로 스케일 / 나노 스케일 디바이스를 생성하기 위해 확장 될 수있다.

A number of recent findings reveal that substrate nanotopography has pronounced influence on cell behavior, from cell adhesion, spreading and migration, to proliferation and differentiation1-6. For instance, a smaller cell size and lower proliferation rate have been observed in cells cultured on deep nanogratings, even leading to apoptosis although the cell alignment, elongation and migration were enhanced, compared with the flat controls2,7-10. Moreover, nanotopography has been shown to facilitate the differentiation of stem cells into certain lineages such as neuron2,11,12, muscle13, and bone3,4. In addition, because of increasing concerns on the toxicity of engineered nanomaterials14,15, there is a need to incorporate nanotopography into physiologically relevant in vitro models for accurate risk assessment of nanomaterials. To fulfil the biochemical and molecular biology analyses, enough cells are needed to be grown on a large area of nanopatterned substrate. However, conventional nanofabrication techniques have limitations to generate highly defined nanopatterns over a large area.
Self-assembly including colloid lithography16 and polymer demixing17 can readily generate large-area nanostructures at low costs. Because self-assembly relies on interactions between the assembling elements such as colloidal particles and macromolecules, and possible interactions between these elements and substrate, it cannot be a stand-alone method of producing nanostructures with precise spatial positioning and arbitrary shapes18. The accompanied high density of defects is also a drawback. Precise spatial control of nanopatterns can be achieved by employing templated self-assembly, which uses top-down lithographic approaches to provide the topographical and/or chemical template to guide the bottom-up assembly of the assembling elements19-21. Alternative nanofabrication techniques such as step-and-flash lithography22 and a roll-to-roll nanoimprinting lithography23 have been developed but have limited use because of their sophisticated process or the requirement of specialized equipment. Nevertheless, a template or a master mold with defined nanoscale patterns is needed for templated or alternative nanofabrication techniques. 
Such templates and master molds are conventionally generated by using focused electron, ion, or photon beam lithography. For instance, electron beam lithography (EBL)24 and focused ion beam lithography25 can generate defined patterns with a sub-5 nm resolution. Two-photon lithography has demonstrated a feature size as small as 30 nm26. Although the focused beam lithography techniques enable generation of well-defined nanoscale structures, the capital investment and the time-consuming, costly process restrict their widespread use in academic research27. Therefore, it is highly desirable to develop enabling yet affordable techniques to produce a large area of nanopatterned surfaces with high fidelity.
We have reported a simple, cost-effective stitch technique for generating a large area of nanopatterned surface from a small well-defined mold28. This protocol provides step-by-step procedure from replication of poly(dimethylsiloxane) (PDMS) molds using an EBL-written pattern, to assembly of multiple PDMS molds into a single large mold, to generation of a master mold on polymeric such as polystyrene (PS) substrates, to production of working substrates. With the expanded nanopatterned substrates, we demonstrated nanotopographical modulation of cell spreading.

<table><tbody><tr><td>JEOL field emission SEM</td><td>JEOL</td><td>JSM-7600F</td><td>EBL</td></tr><tr><td>E-beam evaporator</td><td>Kurt J. Lesker</td><td>Model: LAB 18 e-beam evaporator</td><td>nickel deposition</td></tr><tr><td>Trion Minilock III ICP/RIE</td><td>Trion technology</td><td>Model: Minilock-phantom III</td><td></td></tr><tr><td>Press machine</td><td>PHI Hydraulic Press</td><td>Molde: SQ-230H</td><td></td></tr><tr><td>Spin coater</td><td>Laurell Technologies</td><td>Modle: WS-400A-6NPP-LITE</td><td></td></tr><tr><td>CO&lt;sub&gt;2&lt;/sub&gt; critical dryer</td><td>Tousimis</td><td>Modle: Autosamdri-815</td><td></td></tr><tr><td>Silicon wafer</td><td>University Wafer</td><td>1080</td><td></td></tr><tr><td>Aluminum plates</td><td>McMaster-carr</td><td>9057K123</td><td></td></tr><tr><td>Teflon sheets</td><td>McMaster-carr</td><td>8711K92</td><td></td></tr><tr><td>100 mm Petri dish</td><td>FALCON</td><td>353003</td><td></td></tr><tr><td>60 mm Petri dish</td><td>FALCON</td><td>353004</td><td></td></tr><tr><td>Glass coverslip</td><td>Fisher Scientific</td><td>12-542-B</td><td></td></tr><tr><td>Glass slide</td><td>Fisher Scientific</td><td>12-550-34</td><td></td></tr><tr><td>Disposable weighing boats</td><td>Fisher Scientific</td><td>13-735-743</td><td></td></tr><tr><td>Glass desiccator</td><td>Fisher Scientific</td><td>02-913-360</td><td></td></tr><tr><td>Plastic desiccator</td><td>Bel-Art Products</td><td>F42025-000</td><td></td></tr><tr><td>Hotplate</td><td>Fisher Scientific</td><td>1110049SH</td><td></td></tr><tr><td>Tweezer</td><td>Ted Pella, inc.</td><td>5726</td><td></td></tr><tr><td>Blade</td><td>Fisher Scientific</td><td>S17302</td><td></td></tr><tr><td>Metal blocks</td><td>McMaster-carr</td><td></td><td></td></tr><tr><td>Punch</td><td>Brettuns Village Leather Craft Supplies</td><td>Arch punch</td><td></td></tr><tr><td>Poly(methyl methacrylate)</td><td>MicroChem</td><td>495 PMMA A4</td><td></td></tr><tr><td>PDMS</td><td>Dow Corning</td><td>Sylgard 184 kit</td><td></td></tr><tr><td>Polystyrene</td><td>Dow Chemical</td><td>Styron 685D</td><td></td></tr><tr><td>1H,1H,2H,2H-perfluorooctylmethyldichlorosilane</td><td>Oakwood Chemical</td><td>7142</td><td></td></tr><tr><td>Developer</td><td>MicroChem</td><td>MIBK/IPA at 1: 3 ratio</td><td></td></tr><tr><td>Remover</td><td>MicroChem</td><td>Remover PG</td><td></td></tr><tr><td>Ethanol</td><td>Fisher Scientific</td><td>BP2818500</td><td></td></tr><tr><td>Toluene</td><td>Fisher Scientific</td><td>T324-500</td><td></td></tr><tr><td>Phosphate buffered saline</td><td>Sigma Aldrich</td><td>D8537</td><td></td></tr><tr><td>Dulbecco&amp;rsquo;s modified eagle medium</td><td>Sigma Aldrich</td><td>D5796</td><td></td></tr><tr><td>Fetal bovine serum</td><td>Atlanta Biologicals</td><td>S11550</td><td></td></tr><tr><td>Paraformaldehyde</td><td>Electron Microsopy Science</td><td>15712-S</td><td></td></tr><tr><td>Glutaraldehyde&amp;nbsp;</td><td>Fisher Chemical</td><td>G151-1</td><td></td></tr><tr><td>Fibronectin</td><td>Corning</td><td>356008</td><td></td></tr><tr><td>A549 cells</td><td>ATCC</td><td>ATCC CCL-185</td><td></td></tr></tbody></table>

expanding nanopatterned substrates using stitch technique for nanotopographical modulation of cell behavior

Substrate nanotopography has been shown to be a potent modulator of cell phenotype and function. To dissect nanotopography modulation of cell behavior, a large area of nanopatterned substrate is desirable so that enough cells can be cultured on the nanotopography for subsequent biochemical and molecular biology analyses. However, current nanofabrication techniques have limitations to generate highly defined nanopatterns over a large area. Herein, we present a method to expand nanopatterned substrates from a small, highly defined nanopattern to a large area using stitch technique. The method combines multiple techniques, involving soft lithography to replicate poly(dimethylsiloxane) (PDMS) molds from a well-defined mold, stitch technique to assemble multiple PDMS molds to a single large mold, and nanoimprinting to generate a master mold on polystyrene (PS) substrates. With the PS master mold, we produce PDMS working substrates and demonstrate nanotopographical modulation of cell spreading. This method provides a simple, affordable yet versatile avenue to generate well-defined nanopatterns over large areas, and is potentially extended to create micro-/nanoscale devices with hybrid components.

<html> 스티치 기법은 높은 정확도와 나노 패턴 기판의 넓은 영역을 생성 할 수있다. 도 1a는 각각, Si 기판 상에 PS 판과 PS 박막 스티칭 PDMS 몰드로부터 전송 된 나노 패턴의 넓은 영역을 표시 1B. 원래 EBL-기록 금형 (그림 1C) 및 기판 (그림 1D)를 작업의 최종 PDMS 사이의 비교는 EBL-기록 된 나노 패턴이 충실하게 작업 기판에 전달 될 수 있음을 확인합니다. 다양한 형상 및 크기의 나노 토포 그래피는 셀의 동작을 조절하는데 사용될 수있다. A549과도 2에서 설명한 바와 같이, 모델 셀로서 adenocarcinomic 기저 상피 세포주 이방성 nanogratings는 A549 세포 등방성 나노 기둥에 표시 다극 형태에 비해 nanograting 방향으로 셀을 길게 할 수있다. NT를 &quot;FO : 유지-together.within 페이지 =&quot;1 &quot;&gt; <img alt="그림 1" src="/files/ftp_upload/54840/54840fig1.jpg" /> 스티치 기술을 이용하여 나노 패턴 기판의 대 면적의 1 세대 도표. (A, B)는 각각 PS 판과 PS 박막에 전송 된 나노 패턴의 광학 이미지. 화살표는 스티치 PDMS 금형의 간극에서 폴리머 레이즈를 나타냅니다. (c, d) EBL 금형 및 각각 기판을 작업의 최종 PDMS에 나노 패턴의 SEM 이미지. 스케일 바는 1 ㎛이다. <a href="https://ecsource-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/54840/54840fig1large.jpg" target="_blank">이 그림의 더 큰 버전을 보려면 여기를 클릭하십시오.</a> <img alt="그림 2" src="/files/ftp_upload/54840/54840fig2.jpg" /> 도 2는 A549 세포의 세포 확산 변조 Nanotopographical. </strong&gt; (a) 선폭이 500 nm 인 nanogratings, 간격 500 nm이고, 높이가 560 nm이고, 직경이 500 nm 인 (b) 나노 기둥 450 nm의 가장자리까지의 간격과 높이가 560 nm의 성장 A549 세포 각각. 스케일 바는 10 μm의 수 있습니다. <a href="https://ecsource-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/54840/54840fig2large.jpg" target="_blank">이 그림의 더 큰 버전을 보려면 여기를 클릭하십시오.</a>

Watch this Scientific Journal Video about Expanding Nanopatterned Substrates Using Stitch Technique for Nanotopographical Modulation of Cell Behavior at JoVE.com

Expanding Nanopatterned Substrates Using Stitch Technique for Nanotopographical Modulation of Cell Behavior

A protocol for producing a large area of nanopatterned substrate from small nanopatterned molds for study of nanotopographical modulation of cell behavior is presented.

셀 동작의 Nanotopographical 변조를위한 나노 패턴 기판 사용 스티치 기법을 확장

Substrate nanotopography has been shown to be a potent modulator of cell phenotype and function. To dissect nanotopography modulation of cell behavior, a ...

expanding-nanopatterned-substrates-using-stitch-technique-for

Universidad San Sebastian

Research

JoVE Journal

Bioengineering

6.5K 조회수.  West Virginia University. A protocol for producing a large area of nanopatterned substrate from small nanopatterned molds for study of nanotopographical modulation of cell behavior is presented.

비디오: Expanding Nanopatterned Substrates Using Stitch Technique for Nanotopographical Modulation of Cell Behavior

Stencil Micropatterning of Human Pluripotent Stem Cells for Probing Spatial Organization of Differentiation Fates

Human pluripotent stem cells (hPSCs), including embryonic stem cells and induced pluripotent stem cells, have the intrinsic ability to differentiate into all three germ layers. This makes them an attractive cell source for regenerative medicine and experimental modeling of normal and diseased organogenesis. However, the differentiation of hPSCs in vitro is heterogeneous and spatially disordered. Cell micropatterning technologies potentially offer the means to spatially control stem cell microenvironments and organize the resultant differentiation fates. Micropatterning hPSCs needs to take into account the stringent requirements for hPSC survival and maintenance. Here, we describe stencil micropatterning as a method that is highly compatible with hPSCs. hPSC micropatterns are specified by the geometries of the cell stencil through-holes, which physically confine the locations where hPSCs can access and attach to the underlying extracellular matrix-coated substrate. Due to this mode of operation, there is greater flexibility to use substrates that can adequately support hPSCs as compared to other cell micropatterning methods. We also highlight critical steps for the successful generation of hPSC micropatterns. As an example, we demonstrate that stencil micropatterning of hPSCs can be used to modulate spatial polarization of cell-cell and cell-matrix adhesions, which in turn determines mesoendoderm differentiation patterns. This simple and robust method to micropattern hPSCs widens the prospects of establishing experimental models to investigate tissue organization and patterning during early embryonic development.

Human pluripotent stem cells (hPSCs) have the intrinsic ability to differentiate and self-organize into distinct tissue patterns; although this requires the presentation of spatial environmental gradients. We present stencil micropatterning as a simple and robust method to generate biochemical and mechanical gradients for controlling hPSC differentiation patterns.

차별화 운명의 공간 조직을 프로빙에 대한 인간 다 능성 줄기 세포의 스텐실 미세화

Human pluripotent stem cells (hPSCs), including embryonic stem cells and induced pluripotent stem cells, have the intrinsic ability to differentiate into ...

연구

발생학

Capillary Force Lithography for Cardiac Tissue Engineering

Cardiovascular disease remains the leading cause of death worldwide1. Cardiac tissue engineering holds much promise to deliver groundbreaking medical discoveries with the aims of developing functional tissues for cardiac regeneration as well as in vitro screening assays. However, the ability to create high-fidelity models of heart tissue has proven difficult. The heart&#8217;s extracellular matrix (ECM) is a complex structure consisting of both biochemical and biomechanical signals ranging from the micro- to the nanometer scale2. Local mechanical loading conditions and cell-ECM interactions have recently been recognized as vital components in cardiac tissue engineering3-5.
A large portion of the cardiac ECM is composed of aligned collagen fibers with nano-scale diameters that significantly influences tissue architecture and electromechanical coupling2. Unfortunately, few methods have been able to mimic the organization of ECM fibers down to the nanometer scale. Recent advancements in nanofabrication techniques, however, have enabled the design and fabrication of scalable scaffolds that mimic the in vivo structural and substrate stiffness cues of the ECM in the heart6-9.
Here we present the development of two reproducible, cost-effective, and scalable nanopatterning processes for the functional alignment of cardiac cells using the biocompatible polymer poly(lactide-co-glycolide) (PLGA)8 and a polyurethane (PU) based polymer. These anisotropically nanofabricated substrata (ANFS) mimic the underlying ECM of well-organized, aligned tissues and can be used to investigate the role of nanotopography on cell morphology and function10-14.
Using a nanopatterned (NP) silicon master as a template, a polyurethane acrylate (PUA) mold is fabricated. This PUA mold is then used to pattern the PU or PLGA hydrogel via UV-assisted or solvent-mediated capillary force lithography (CFL), respectively15,16. Briefly, PU or PLGA pre-polymer is drop dispensed onto a glass coverslip and the PUA mold is placed on top. For UV-assisted CFL, the PU is then exposed to UV radiation (&#955; = 250-400 nm) for curing. For solvent-mediated CFL, the PLGA is embossed using heat (120 &#176;C) and pressure (100 kPa). After curing, the PUA mold is peeled off, leaving behind an ANFS for cell culture. Primary cells, such as neonatal rat ventricular myocytes, as well as human pluripotent stem cell-derived cardiomyocytes, can be maintained on the ANFS2.

In this protocol, we demonstrate the fabrication of biomimetic cardiac cell culture substrata made from two distinct polymeric materials using capillary force lithography. The described methods provide a scalable, cost-effective technique to engineer the structure and function of macroscopic cardiac tissues for in vitro and in vivo applications.

심장 조직 공학을위한 모세관 력 리소그래피

Cardiovascular disease remains the leading cause of death worldwide1. Cardiac tissue engineering holds much promise to deliver groundbreaking medical ...

생체공학

Microfabrication of Nanoporous Gold Patterns for Cell-material Interaction Studies

Nanostructured materials with feature sizes in tens of nanometers have enhanced the performance of several technologies, including fuel cells, biosensors, biomedical device coatings, and drug delivery tools. Nanoporous gold (np-Au), produced by a nano-scale self-assembly process, is a relatively new material that exhibits large effective surface area, high electrical conductivity, and catalytic activity. These properties have made np-Au an attractive material to scientific community. Most studies on np-Au employ macro-scale specimens and focus on fundamental science of the material and its catalytic and sensor applications. The macro-scale specimens limit np-Au's potential in miniaturized systems, including biomedical devices. In order to address these issues, we initially describe two different methods to micropattern np-Au thin films on rigid substrates. The first method employs manually-produced stencil masks for creating millimeter-scale np-Au patterns, while the second method uses lift-off photolithography to pattern sub-millimeter-scale patterns. As the np-Au thin films are obtained by sputter-deposition process, they are compatible with conventional microfabrication techniques, thereby amenable to facile integration into microsystems. These systems include electrically-addressable biosensor platforms that benefit from high effective surface area, electrical conductivity, and gold-thiol-based surface bioconjugation. We describe cell culture, immunostaining, and image processing techniques to quantify np-Au's interaction with mammalian cells, which is an important performance parameter for some biosensors. We expect that the techniques illustrated here will assist the integration of np-Au in platforms at various length-scales and in numerous applications, including biosensors, energy storage systems, and catalysts.

We report on techniques to micropattern nanoporous gold thin films via stencil printing and photolithography, as well as methods to culture cells on the microfabricated patterns. In addition, we describe image analysis methods to characterize morphology of the material and the cultured cells using scanning electron and fluorescence microscopy techniques.

세포 물질의 상호 작용 연구를위한 나노 기공 골드 패턴의 미세

Nanostructured materials with feature sizes in tens of nanometers have enhanced the performance of several technologies, including fuel cells, biosensors, ...

Cell Patterning on Photolithographically Defined 
Parylene-C: SiO2 Substrates

Cell patterning platforms support broad research goals, such as construction of predefined in vitro neuronal networks and the exploration of certain central aspects of cellular physiology. To easily combine cell patterning with Multi-Electrode Arrays (MEAs) and silicon-based &#8216;lab on a chip&#8217; technologies, a microfabrication-compatible protocol is required. We describe a method that utilizes deposition of the polymer parylene-C on SiO2&#160;wafers. Photolithography enables accurate and reliable patterning of parylene-C at micron-level resolution. Subsequent activation by immersion in fetal bovine serum (or another specific activation solution) results in a substrate in which cultured cells adhere to, or are repulsed by, parylene or SiO2 regions respectively. This technique has allowed patterning of a broad range of cell types (including primary murine hippocampal cells, HEK 293 cell line, human neuron-like teratocarcinoma cell line, primary murine cerebellar granule cells, and primary human glioma-derived stem-like cells). Interestingly, however, the platform is not universal; reflecting the importance of cell-specific adhesion molecules. This cell patterning process is cost effective, reliable, and importantly can be incorporated into standard microfabrication (chip manufacturing) protocols, paving the way for integration of microelectronic technology.

Cell Patterning on Photolithographically Defined 
Parylene-C: SiO2 Substrates

This protocol describes a microfabrication-compatible method for cell patterning on SiO2. A predefined parylene-C design is photolithographically printed on SiO2 wafers. Following incubation with serum (or other activation solution) cells adhere specifically to (and grow according to the conformity of) underlying parylene-C, whilst being repulsed by SiO2 regions.

포토 리소그래피 정의 파릴 렌-C에 대한 세포 패터닝 : 그런가<sub&gt; 2</sub&gt; 기판

Cell patterning platforms support broad research goals, such as construction of predefined in vitro neuronal networks and the exploration of certain ...

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 ...

Simple Lithography-Free Single Cell Micropatterning using Laser-Cut Stencils

Micropatterning techniques have been widely used in cell biology to study effects of controlling cell shape and size on cell fate determination at single cell resolution. Current state-of-the-art single cell micropatterning techniques involve soft lithography and micro-contact printing, which is a powerful technology, but requires trained engineering skills and certain facility support in microfabrication. These limitations require a more accessible technique. Here, we describe a simple alternative lithography-free method: stencil-based single cell patterning. We provide step-by-step procedures including stencil design, polyacrylamide hydrogel fabrication, stencil-based protein incorporation, and cell plating and culture. This simple method can be used to pattern an array of as many as 2,000 cells. We demonstrate the patterning of cardiomyocytes derived from single human induced pluripotent stem cells (hiPSC) with distinct cell shapes, from a 1:1 square to a 7:1 adult cardiomyocyte-like rectangle. This stencil-based single cell patterning is lithography-free, technically robust, convenient, inexpensive, and most importantly accessible to those with a limited bioengineering background.

This protocol introduces a lithography-free micropatterning method that is simple and accessible to those with a limited bioengineering background. This method utilizes customized laser-cut stencils to micropattern extracellular matrix proteins in a shape of interest for modulating cell morphologies. The procedure for micropatterning is demonstrated using induced pluripotent stem cell derived cardiomyocytes.

레이저 컷 스텐실을 사용한 간단한 리소그래피 없는 단일 셀 마이크로패터닝

Micropatterning techniques have been widely used in cell biology to study effects of controlling cell shape and size on cell fate determination at single ...

A Versatile Method of Patterning Proteins and Cells

Substrate and cell patterning techniques are widely used in cell biology to study cell-to-cell and cell-to-substrate interactions. Conventional patterning techniques work well only with simple shapes, small areas and selected bio-materials. This article describes a method to distribute cell suspensions as well as substrate solutions into complex, long, closed (dead-end) polydimethylsiloxane (PDMS) microchannels using negative pressure. This method enables researchers to pattern multiple substrates including fibronectin, collagen, antibodies (Sal-1), poly-D-lysine (PDL), and laminin. Patterning of substrates allows one to indirectly pattern a variety of cells. We have tested C2C12 myoblasts, the PC12 neuronal cell line, embryonic rat cortical neurons, and amphibian retinal neurons. In addition, we demonstrate that this technique can directly pattern fibroblasts in microfluidic channels via brief application of a low vacuum on cell suspensions. The low vacuum does not significantly decrease cell viability as shown by cell viability assays. Modifications are discussed for application of the method to different cell and substrate types. This technique allows researchers to pattern cells and proteins in specific patterns without the need for exotic materials or equipment and can be done in any laboratory with a vacuum.

This report describes a simple, easy to perform technique, using low pressure vacuum, to fill microfluidic channels with cells and substrates for biological research.

패터닝 단백질과 세포의 다목적 방법

Substrate and cell patterning techniques are widely used in cell biology to study cell-to-cell and cell-to-substrate interactions. Conventional patterning ...

Fabrication of Gradient Nanopattern by Thermal Nanoimprinting Technique and Screening of the Response of Human Endothelial Colony-forming Cells

Nanotopography can be found in various extracellular matrices (ECMs) around the body and is known to have important regulatory actions upon cellular reactions. However, it is difficult to determine the relation between the size of a nanostructure and the responses of cells owing to the lack of proper screening tools. Here, we show the development of reproducible and cost-effective gradient nanopattern plates for the manipulation of cellular responses. Using anodic aluminum oxide (AAO) as a master mold, gradient nanopattern plates with nanopillars of increasing diameter ranges [120-200&#8239;nm (GP 120/200), 200-280&#8239;nm (GP 200/280), and 280-360&#8239;nm (GP 280/360)] were fabricated by a thermal imprinting technique. These gradient nanopattern plates were designed to mimic the various sizes of nanotopography in the ECM and were used to screen the responses of human endothelial colony-forming cells (hECFCs). In this protocol, we describe the step-by-step procedure of fabricating gradient nanopattern plates for cell engineering, techniques of cultivating hECFCs from human peripheral blood, and culturing hECFCs on nanopattern plates.

Here, we present a protocol for the fabrication of gradient nanopattern plates via thermal nanoimprinting and the method of screening responses of human endothelial progenitor cells to the nanostructures. By using the described technology, it is possible to produce a scaffold that can manipulate cell behavior by physical stimuli.

열 Nanoimprinting 기술과 인간의 내 피 식민지 형성 세포의 응답의 심사에 의해 그라데이션 Nanopattern의 제조

Nanotopography can be found in various extracellular matrices (ECMs) around the body and is known to have important regulatory actions upon cellular ...

Control of Cell Geometry through Infrared Laser Assisted Micropatterning

Micropatterning is an established technique in the cell biology community used to study connections between the morphology and function of cellular compartments while circumventing complications arising from natural cell-to-cell variations. To standardize cell shape, cells are either confined in 3D molds or controlled for adhesive geometry through adhesive islands. However, traditional micropatterning techniques based on photolithography and deep UV etching heavily depend on clean rooms or specialized equipment. Here we present an infrared laser assisted micropatterning technique (microphotopatterning) modified from Doyle et al. that can be conveniently set up with commercially available imaging systems. In this protocol, we use a Nikon A1R MP+ imaging system to generate micropatterns with micron precision through an infrared (IR) laser that ablates preset regions on poly-vinyl alcohol coated coverslips. We employ a custom script to enable automated pattern fabrication with high efficiency and accuracy in systems not equipped with a hardware autofocus. We show that this IR laser assisted micropatterning (microphotopatterning) protocol results in defined patterns to which cells attach exclusively and take on the desired shape. Furthermore, data from a large number of cells can be averaged due to the standardization of cell shape. Patterns generated with this protocol, combined with high resolution imaging and quantitative analysis, can be used for relatively high throughput screens to identify molecular players mediating the link between form and function.

The protocol presented here enables automated fabrication of micropatterns that standardizes cell shape to study cytoskeletal structures within mammalian cells. This user-friendly technique can be set up with commercially available imaging systems and does not require specialized equipment inaccessible to standard cell biology laboratories.

적외선 레이저 보조 마이크로패터닝을 통한 세포 지오메트리 제어

Micropatterning is an established technique in the cell biology community used to study connections between the morphology and function of cellular ...

Pattern Generation for Micropattern Traction Microscopy

Micropattern traction microscopy allows control of the shape of single cells and cell clusters. Furthermore, the ability to pattern at the micrometer length scale allows the use of these patterned contact zones for the measurement of traction forces, as each micropatterned dot allows for the formation of a single focal adhesion that then deforms the soft, underlying hydrogel. This approach has been used for a wide range of cell types, including endothelial cells, smooth muscle cells, fibroblasts, platelets, and epithelial cells. 
This review describes the evolution of techniques that allow the printing of extracellular matrix proteins onto polyacrylamide hydrogels in a regular array of dots of prespecified size and spacing. As micrometer-scale patterns are difficult to directly print onto soft substrates, patterns are first generated on rigid glass coverslips that are then used to transfer the pattern to the hydrogel during gelation. First, the original microcontact printing approach to generate arrays of small dots on the coverslip is described. A second step that removes most of the pattern to leave islands of small dots is required to control the shapes of cells and cell clusters on such arrays of patterned dots. 
Next, an evolution of this approach that allows for the generation of islands of dots using a single subtractive patterning step is described. This approach is greatly simplified for the user but has the disadvantage of a decreased lifetime for the master mold needed to make the patterns. Finally, the computational approaches that have been developed for the analysis of images of displaced dots and subsequent cell-generated traction fields are described, and updated versions of these analysis packages are provided.

We describe improvements to a standard method for measuring cellular traction forces, based on microcontact printing with a single subtractive patterning step of dot arrays of extracellular matrix proteins on soft hydrogels. This method allows for simpler and more consistent fabrication of island patterns, essential for controlling cell cluster shape.

마이크로패턴 견인 현미경을 위한 패턴 생성

Micropattern traction microscopy allows control of the shape of single cells and cell clusters. Furthermore, the ability to pattern at the micrometer ...