우리는 공 촛점 이미징에 대한 자신의 훈련과 조언을 닥터 제프리 캐플란 감사합니다. 우리는 또한 SEM의 지원 중사 전자 현미경 연구소 박사 Chaoying 니켈 감사합니다. 이 작품은 건강 (NIDCD, R01DC008965 및 R01DC011377)의 국립 연구소에 의해 자금 지원됩니다. ABZ은 NSF 통합 대학원 교육 및 연구 훈련생 자금 (IGERT) 프로그램을 인정합니다.

1. 생물 반응기 조립 (비디오 1) <ol><li> 미리 정해진 내부 및 외부 치수 알루미늄 형 (원형 다이 + 스페이서 핀) (그림 1) 확인합니다. </li><li> 에서 (그림 1의 스페이서 핀에 의해 형성 12mm, 두께 ~ 0.25 mm, 직경) 입지를 굳힌 슬리브 단계 1.1에서 금형을 사용하여, 실리콘 막 (1.5 mm, 그림 1 : 42mm, 두께 직경) 제작 시판되는 실리콘 엘라스토머 키트를 사용하여 중간. </li><li> 일치하는 능선과 홈과 블록 (0.9 cm 두께) 상단을 조각 (1.8 cm 두께)과 하단, 중간에 : 원형 개구 (24mm 직경)와 아크릴 블록 쌍 (그림 2 (4, 5)) 확인 . 10 </li><li> 페어링 아크릴계 블록 사이 실리콘 막을 샌드위치. 일정한 힘 (35 CN · m)로 설정 마이크로 토크 드라이버를 사용하여 네 모서리의 나사를 사용하여 어셈블리를 고정합니다. 결과적으로, 방수, 24mm 폭 18mm 깊이 진동 챔버 (그림 2C)가 생성됩니다. </li><li> 바닥 아크릴 블록에 각 나사의 또 다른 세트를 통해 진동 실 아래 3 &quot;확장 된 범위의 미니 우퍼 (그림 2D, 8 Ω/20 W)를 설치합니다. 이 시점에서, 개별 진동 모듈 조립된다. </li><li> 7 개의 추가 진동 모듈을 복제합니다. 바로 절단 균등하게 원형 구멍 (: 7cm, 두께 2cm 직경)의 스피커 기지를 배치하여 두 개의 고정 된 알루미늄 바 (40cm X 10cm X 2.5 cm) 중 하나에 4 개를 붙입니다. 각각의 원형 구멍에 알루미늄 막대의 측면을 통해 나사를 삽입하여 각 스피커를 안정. </li><li> 개별 스피커를 선택하여 스피커를 제어 할 수 있습니다. 선택의 해당 출력에 다음 스피커 본체의 양극과 음극 입력에 와이어를 연결하여 선택 개별 스피커를 연결합니다. 스피커의 선택은 수 t의 신호그 함수 발생기는, 전력 증폭기를 통과 한 후, 일단 (도 2E)에 모두 팔 스피커에 도달한다. </li><li> 반대로 습도가 인클로저에 두 개의 챔버 어레이 스피커 선택 및 관련 전자 제품을 놓습니다. 상업 세포 배양 인큐베이터에서 전체 어셈블리를 집. </li><li> 인큐베이터의 뒤쪽에 필터 조립체를 통해 파워 앰프와 스피커를 연결 선택기 (의료용 PVC 튜빙을 통해) 메인 케이블 피드. </li></ol> 2. 비계 제조 및 특성 <ol><li> 15 중량 %의 농도로 클로로포름 PCL 펠릿을 용해. 21 G의 무딘 엔드 바늘로 덮인 10 ML의 주사기에 솔루션을로드합니다. </li><li> 프로그램 주사기 펌프에 주사기를 잠그고 / 시간 1 ㎖에 유량을 설정합니다. </li><li> ~ 18cm의 바늘 끝 - 투 - 컬렉터 거리를 수평으로 바늘 맞은 편에 알루미늄 호일 덮인 집 놓습니다. </li><li> 긍정적 인 알리 클램프바늘의 중간, 알루미늄 집 전체에 접지 악어 클립 게이터 클립은, 다음에 15 kV의 고전압 전원 장치에 전압을 설정한다. 주의 : 고전압, 바늘에서 거리를 유지. </li><li> 순차적으로 주사기 펌프와 전원 공급 장치의 전원을 켭니다; 신속 클린 / 안정 섬유 제트 전에 건조 종이 타월을 이용하여 바늘의 선단부를 둘러싸는 잔류 중합체 용액을 제거하고 테일러 콘 (19)을 형성한다. </li><li> 섬유는 250 ~ 300 μm의 ~의 두께로 알 수집기에 축적 할 수 있도록 (~ 현재 회전 상태에서 7 시간). 잔류 용매를 제거 1-2 일 동안 진공 건조기에서 얻어진 지지체를 저장한다. </li><li> 화상 지지체가 일관된 섬유 형태를 보여주기 위하여 주사 전자 현미경을 사용하여, 금으로 스퍼터 코팅. 10 </li></ol> 3. 생물 반응기 조립 및 특성 <ol><li> 네 팔 (길이 : 원통형 디스크 (8mm 직경) : 펀치 2우선 디스크의 외경을 잘라 12mm 직경 생검 펀치를 사용하여 같은 회전 PCL 매트 (도 2a) 중 mm). 그런 균등 팔이 절단 어디 점수를 원형 블레이드 주위에 간격 네 2mm 긴 홈을 만들기 위해 두 번째, 8mm 생검 펀치를 사용합니다. 8 개 mm 펀치로 득점 한 후, 바깥쪽으로 팔의 가장자리를 잘라 메스 블레이드를 사용합니다. 연장 아암을 통해 실리콘 막의 홈에 지지체를 삽입 (비디오 1). 온화 납작한 핀셋을 사용하여 표면을 눌러 삽입 비계 평평. </li><li> 레이저 반사를 돕기 위해 PCL 지지체에 얇은 알루미늄 포일의 작은 조각 (8mm × 2 mm의 수직 모양, 그림 2B)를 연결합니다. </li><li> 진동 실에서 조립 된 실리콘 막 / PCL 발판을 (단계 1.4에 설명 된대로)에 고정합니다. 진동 전에 PCL의 발판을 수화하기 위해 챔버에 1.5 ㎖의 물을 추가합니다. </li><li> 함수 발생기를 이용하여 진동 신호 소개ALS 개재 아크릴 챔버 (0.1 V의 피크 - 피크 전압 Vpp로 가진 예를 들면, 200Hz의 사인파). 정확하게 각 스피커로의 입력 전압을 측정하기위한 전압계를 사용한다. 참고 : 함수 발생기에서 Vpp로 판독 스피커에 전달되는 최종 전압이 다를 것입니다. </li><li> 단일 지점 LDV를 조립하고 팬 틸트 헤드 삼각대에 광섬유 레이저 센서 헤드를 고정합니다. 각도 센서 헤드는 테이블 탑에 수직을 가리 키도록. 동축 케이블을 통해 데이터 수집 모듈 LDV 센서 헤드를 연결 한 후 USB를 통해 노트북에 모듈. </li><li> 실리콘 막 (도 2b 및도 3)에 다양 소정 지점에서 수직으로 레이저 빔을 집중. </li><li> 데이터 수집 소프트웨어를 이용하여, 중간 막 변위를 기록한다. &quot;옵션&quot;메뉴에서 &quot;획득 설정&quot;을 클릭합니다; 다음 측정 모드를 &quot;FFT 변경221;. 다음으로, 기본 도구 모음에서 &quot;연속 측정&quot;을 클릭 한 후 변위를 기록 선택된 주파수 (그림 6D)에 형성하는 피크를 클릭합니다. </li><li> 기판에 걸쳐 상대적 위치의 함수로서 정상 미드 막 변위 (0 w)를 그린다. 원래 8.5 데이터 분석 소프트웨어를 사용하여 표면 진동 프로파일의 3D 컬러 맵을 구축. </li></ol> 4. 진동 세포 배양 <ol><li> MSC 유지 보수 미디어 / ㎠ 4,000 ~ 5,000 세포의 초기 시딩 밀도 T150 조직 배양 플라스크에서 하위 문화 인간 골수 유래 중간 엽 줄기 세포. </li><li> 세포 배양 7-8 일 후 (~ 85 % 컨 플루로) 등 accutase 같은 세포 해리 시약과 세포를 Trypsinize, 혈구, 원심 분리 (5 분 440 XG)를 사용하여 계산하고, 다시 중단 세포 펠릿 4.5 × 106 세포 / ml의 농도로 신선한 MSC 유지 매체. </li><li> PCL의 scaffo를 담가70 % 에탄올 O / N.에 LD 용매를 증발 한 후 5-8 분 동안 UV 광 (254 ㎚ 인) 살균하는 지지체의 양면을 노출. </li><li> 1 시간 동안 37 ° C에서 20 ㎍ / ml의 피브로넥틴 솔루션의 PCL의 발판을 적신다. 실리콘 막에 피브로넥틴 코팅 된 지지체를 삽입합니다. 1 단계에서 설명하는대로 생물 반응기를 조립합니다. </li><li> 보안 PCL 발판에 균등하게 세포 현탁액의 40 μl를 배포합니다. 세포가 진동 챔버에 추가 1.5 ML 신선한 매체를 추가하기 전에 1 ~ 1.5 시간 동안 부착 할 수 있습니다. </li><li> 문화 MSC 라덴 PCL 3 일간 정적으로 비계와 정적 문화가 완료되면 미디어를 새로 고칩니다. </li><li> 세포 구조에 선정 진동 정권을 부과. 참고 : 예를 들어, 세포는 최대 7 일 동안 하루에 12 시간 ~ 40 ㎛, 아 0에서 200 Hz에서 진동 (의) 1 - 시간 -에 - 1 - 시간 - 오프를 실시한다. 진동 자극을받는 구조는 VIB 샘플 및 그 cultu로 지정정적으로 동일한 진동 챔버 빨간색은 정적 컨트롤 (합계) 역할을한다. </li></ol> 5. 생물 대한 평가 <ol><li> 매일 각 챔버로부터 200 ㎕의 세포 배양 배지를 수집 (일 1, 3, 5, 7) 및 (800 μL 각) 함께 동일 샘플로부터 분취 풀. </li><li> 매트릭스 메탈-1 (MMP1)와 제조 업체의 절차에 따라, 각각 MMP1 ELISA 개발 키트 및 히알루 론산 경쟁 ELISA 키트를 사용하여 HA의 세포 생산을 정량화. 한편, 이전에보고 된 ELISA 절차에 따라 수용성 엘라스틴 전구체의 생산을 분석 실험. 8 </li><li> 일 7시에 마지막 진동주기가 완료되면, 신속하게 날카로운 핀셋을 사용하여 진동 챔버에서 세포 구조를 제거하고 간단하게 감기 인산염 완충 생리 식염수 (PBS, 4 ° C)로 헹구어. </li><li> 라이브 / 죽은 염색의 경우, 요오드화 프로피 디움 (PBS에서 1:2,000)와 구문을 배양하고SYTO-13 RT에서 5 분을 동시에 (PBS에서 1:1,000). 이미지 다 광자 공 초점 현미경으로 얼룩진 구조. </li><li> 이와는 별도로, 드라이 아이스의 PBS-씻어서 세포 구조를 스냅인 동결 및 유전자 분석을위한 이전에보고 된 프로토콜 다음 전체 세포 RNA를 추출 할 수 있습니다. 9 </li><li> UV-VIS 분광 광도계를 사용하여 추출 된 RNA의 수량과 품질을 확인합니다. 1.8 ~ 2.2의 A260/A280과 A260/A230 비율이 RNA 샘플은 다음의 qPCR 분석을 위해 사용됩니다. </li><li> 역 시판 역전사 키트를 사용하여 cDNA를에 RNA (500 NG / 샘플)를 표기. </li><li> 이전에 설명 된 절차에 따라 시판 PCR 마스터 믹스를 이용한 실시간 연속 검출 시스템에 대한 PCR 반응을 수행한다. 8 </li><li> 상업적 qPCR의 데이터 분석 소프트웨어를 사용 qPCR의 결과를 분석한다. 데이터 분석의 신뢰도를 보장하기 위해, 여러 표준 발현 유전자 (YWHAZ, TBP, PPIA)는 INT로서 이용된다ernal 컨트롤 및 특정 프라이머 효율성의 분산을 고려해야한다. 8 </li></ol>

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더 경쟁하는 금융 이익은 존재하지 않습니다.

체외에서 기능 성대 조직의 성공적인 설계는 다 능성 세포의 행동을 중재하기 위해 성대와 같은 미세 환경의 휴양이 필요합니다. 그것은 일반적으로 조직 또는 장기 구조 그들이 수행하는 데 필요한 기능을 반영하는 것으로 받아 들여진다. 성대 조직 22는, 발성 중에 발생 고주파 진동은 조직의 성숙을위한 중요한 것으로 제시된다. 성대 생물 반응기 배양 된 중간 엽 줄기 세포에 생...

상피 층으로 구성된 인간의 성대는 고유 층 (LP) 및 vocalis 근육, 사운드 제작을위한 음향 파에 폐에서 공기 흐름을 변환하는 전문 부드러운 조직이다. 1 성대가 정상 발성을하는 동안 정기적으로 진동, 에 이르기까지 기본 주파수에서 최대 30 %의 변종을 나타내는 100 ~ 300 Hz에서에서. 2 성인 성대 LP는 표면 (SLP)로 구성된 그라데이션 구조, 중간 (ILP)과 깊이 (DLP) 층이다. 또한 분류 그룹 상피 세포와 점막 층으로 SLP, 그리고 보컬 인대에 ILP 및 DLP를 결합합니다. 3 인대가 성숙 콜라겐과 엘라스틴 섬유가 풍부하면서 SLP 층, 띄엄 띄엄 분산 된 콜라겐 섬유와 주로 비정질 매트릭스를 포함 충분한 강도를 제공 할 수 있습니다. 4의 구조와 신생아 성대의 역학은 성숙한 대응 크게 다릅니다. 비록 메커니즘성대의 발전과 성숙을 조절의는 아직 완전히 실험적인 증거가 발성 파생 기계적 스트레스의 정의 역할을 지적했다, 이해되지 않습니다. 음성 남용, 감염, 화학적 자극 및 수술 등 여러 가지 건강 상태는 성대가 손상 될 수 있습니다. 성대 장애는 미국 인구의 추정 3-9% 영향을 미친다. 성대 질환에 대한 현재의 치료 방법은 5 개 제한하고 줄기 세포 기반 조직 공학 방법은 성대 기능 복원을위한 유망한 전략으로 떠오르고있다. 중간 엽 줄기 세포 (MSC) 성대 조직 공학에 대한 기본 성대 섬유 아 세포에 대한 적절한 대안입니다. 6-9 줄기 세포의 운명 사양 및 이후의 조직 개발은 기계 상태가되는가에있는 특정 틈새 시장에 의해 매개되는 중요한 요소. 10 기계적 힘은 조직의 형태 형성에 필수적인 레귤레이터이다ND 특히 일상적으로 조직 공학의 관점에서로드. 11을 실시하고 조직에 대한 항상성은, 그것은 노출 생리학 관련 기계적 자극이 세포 분화 및 조직 별 매트릭스 리모델링 줄기 촉진하는 것으로 입증되었다. 12-15 조직 배양 용 바이오 리액터는 시험 관내 세포 또는 조직 성장을위한 바람직한 생리적 환경을 시뮬레이션하기 위해 설계된다. 성대 조직 공학의 경우, phonating 성대의 기계적인 환경을 재현하는 것이 특히 중요합니다. 이상적인 성대 생물 반응기 효과적으로 주파수, 진폭과 진동의 기간 동안 손쉬운 제어를 허용, 배양 세포에 진동 신호를 제공한다. Titze 및 동료 기질 단백질의 세포 생산을 자극하는 높은 주파수 (20 ~ 200 Hz에서) 진동과 정적 스트레칭을 결합한 성대 생물 반응기 (T1 생물 반응기) 16을 고안했습니다. 저기서g이 생물 반응기, 웹의 동료 (17)은 히알루 론산 (HA) 기반의 하이드로 겔에서 배양 피부 섬유 아세포에서 10 일, 100 Hz에서 진동의 영향을 연구했다. 진동이 구문은 HA 합성 효소-2 (HAS2) 정적 컨트롤을 기준으로, decorin, fibromodulin 매트릭스 메탈-1 (MMP1)의 높은 발현을 보였다. 자극 효과가 따라 시간이 될 것으로 밝혀졌다. 더 최근에는, 단체 (18)는 전력 증폭기, 함수 발생기, 봉입 시끄러운 스피커 및 세포 부착에 진동 공기 전송 원주 정박 실리콘 막을 사용 성대 생물 반응기 (J1 생물 반응기)를 조립했다. J1 생물 반응기에서 배양 신생아 포피의 섬유 아 세포는 최대 0.05 %의 평면 변형으로, 60, 110 또는 300 Hz에서 진동이 1 시간을 실시 하였다. qPCR의 결과는 몇 가지 ECM 유전자의 발현이 적당히 다양한 진동 주파수에 대한 응답으로 변경된 것을 제안및 진폭. 이러한 생물 반응기 설계, 음모를 꾸미는 동안, 몇 가지 제한이 있습니다. 예를 들어, T1 시스템이 달성 가능한 최대 주파수를 제한하는 기계적 결합을위한 커넥터와 바의 다수를 필요로한다. 또한, 세포는 데이터 해석을 복잡 바람직 기계적 교반 및 액체 섭동을 거치게된다. J1 생물 반응기는, 반면에, 상대적으로 낮은 에너지 변환 효율을 나타내고 사용자 친화적 아니다. 또, 진동이 빈번 내부 실리콘 막에서 세포 함유 구조물을 분리. J1 버전과 동일한 원칙에 따라 설계 여기에보고 J2 성대 생물 반응기는, 일관성과 재현성에 최적화되어 있습니다. 발성을 흉내 낸 진동은 MSC-채워 섬유 폴리 (ε-카프로 락톤) (PCL) 발판이 effectiv 있습니다 개별적으로 장착되어 진동 챔버에 공기 역학적으로 생성됩니다엘리 확보했다. 레이저 도플러 진동계 (LDV)는 사용자가 멤브레인 / 지지체 조립체의 진동 프로파일을 확인한다. 우리의 데모에서는 중간 엽 줄기 세포 7 일 동안 12 시간의 매일의 총 1 시간 -에 - 1 - 시간 - 오프 (의) 패턴으로 200 Hz의 정현파 진동에 노출되어 있습니다. 부과 된 진동 신호에 대한 세포 반응을 체계적으로 조사하고 있습니다. 전반적으로, J2 성대 동적 세포 배양 연구는 높은 처리량과 재생 가능한 방식으로 수행 할 수 있도록, 가장 사용자 친화적 인 기능을 제공합니다.

<table border="0" cellpadding="0" cellspacing="0" width="853">
	<tbody>
		<tr height="16">
			<td height="16" style="height:17px;width:248px;">silicone elastomer kit</td>
			<td style="width:188px;">Dow Corning</td>
			<td style="width:124px;">Sylgard 184</td>
			<td style="width:293px;">cure the membrane at 100 C for 2 hr</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">PCL</td>
			<td>Sigma Aldrich</td>
			<td>440744-500G</td>
			<td>Mn ~ 80 kDa, dissolve overnight</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">chloroform</td>
			<td>Sigma Aldrich</td>
			<td>C7559-5VL</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">human bone marrow-derived MSCs</td>
			<td>Lonza</td>
			<td>PT-2501</td>
			<td>received with passage 2</td>
		</tr>
		<tr height="33">
			<td height="33" style="height:33px;">MSC maintenance media</td>
			<td>Lonza</td>
			<td>PT-3001</td>
			<td style="width:293px;">10% FBS in basal media supplemented 
			with L-glutamine, gentamicin and amphotericin</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">Accutase cell dissociation reagent</td>
			<td>Life Technologies</td>
			<td>A11105-01</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">ethanol</td>
			<td>Sigma Aldrich</td>
			<td>E7023-500ML</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">fibronectin</td>
			<td>Sigma Aldrich</td>
			<td>F2006-1MG</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">MMP1 DuoSet ELISA kit</td>
			<td>R&#38;D systems</td>
			<td>DY901</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">HA ELISA kit</td>
			<td>Echelon Biosciences</td>
			<td>&#160;K-1200&#160;&#160;</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">PBS</td>
			<td>Life Technologies</td>
			<td>14190-136</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">propidium iodide&#160;</td>
			<td>Life Technologies</td>
			<td>P1304MP</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">Syto-13&#160;</td>
			<td>Life Technologies</td>
			<td>S7575</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">QuantiTect reverse transcription kit&#160;</td>
			<td>Qiagen</td>
			<td>205311</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">SYBR Green PCR master mix</td>
			<td>Life Technologies</td>
			<td>4309155</td>
			<td></td>
		</tr>
		<tr height="33">
			<td height="33" style="height:33px;">replacement speaker</td>
			<td style="width:188px;">DAYTON audio 
			(via Parts Express)</td>
			<td>DS90-8</td>
			<td>paper cone, full range (80-13000 Hz), 85dB</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">Ergo Micro torque screwdriver</td>
			<td>Mountz</td>
			<td># 020377</td>
			<td>torque range: 20-120 cN.m</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">stereo speaker selector</td>
			<td>RadioShack</td>
			<td>40-244</td>
			<td>maximum power handling 50 W</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">function generator&#160;</td>
			<td>Agilent&#160;</td>
			<td>33220A</td>
			<td>frequency range 1 &#181;Hz- 20 MHz</td>
		</tr>
		<tr height="33">
			<td height="33" style="height:33px;">power amplifier&#160;</td>
			<td>PYLE audio</td>
			<td>PylePro PT2400</td>
			<td style="width:293px;">frequency response: 10 Hz-50 kHz, two speaker 
			channels</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">cell culture incubator&#160;</td>
			<td>Thermo Fisher&#160;</td>
			<td>Steri-Cult 3307</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">syringe pump&#160;</td>
			<td>New Era Pump Systems</td>
			<td>NE-300</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">High voltage power supply</td>
			<td>Spellman</td>
			<td>CZE 1000R</td>
			<td>output voltage: 0-30 kV</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">scanning electron microscope&#160;</td>
			<td>JEOL-USA</td>
			<td>JSM-7400F</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">desk gold sputter coater</td>
			<td>Denton Vacuum</td>
			<td>DSK00V-0013</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">Doppler laser vibrometer&#160;</td>
			<td>Polytec</td>
			<td>PDV-100</td>
			<td>non-contact velocity measurement (0-22 kHz)</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">PCR sequence detection system&#160;</td>
			<td>Applied Biosystems</td>
			<td>ABI7300</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">multiphoton confocal microscope</td>
			<td>Zeiss</td>
			<td>Zeiss 510Meta NLO</td>
			<td></td>
		</tr>
		<tr height="33">
			<td height="33" style="height:33px;">UV-VIS Spectrophotometer&#160;</td>
			<td style="width:188px;">NanoDrop Products 
			via Thermo Scientific</td>
			<td>ND-2000</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">VibSoft Data Acquisition Software</td>
			<td>Polytec</td>
			<td></td>
			<td>acquisition bandwidth up to 40 MHz</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">Origin 8.5 data analysis software&#160;</td>
			<td>OriginLab</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">qbasePlus qPCR data analysis software&#160;</td>
			<td>Biogazelle</td>
			<td>V2.3</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">aluminium alloy&#160;</td>
			<td>McMaster-Carr</td>
			<td>Alloy 6061</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">acrylic blocks</td>
			<td>McMaster-Carr</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="27">
			<td height="27" style="height:27px;">polycarbonate anti-humidity chamber</td>
			<td>McMaster-Carr</td>
			<td></td>
			<td style="width:293px;">Impact-Resistant&#160;Polycarbonate</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">screws&#160;</td>
			<td>McMaster-Carr</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">electronic cable/wire</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">medical grade PVC tubing</td>
			<td>US Plastic Corp.</td>
			<td>Tygon S-50-HL</td>
			<td>clear, biocompatible</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">10 mL syringe&#160;</td>
			<td>Becton Dickinson</td>
			<td>309604</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">21 G blunt ended needle</td>
			<td>Small Parts</td>
			<td>NE-213PL-25</td>
			<td>1-1/2&#34; length</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">Alligator clip adapters&#160;</td>
			<td>RadioShack</td>
			<td>270-354</td>
			<td>fully insulated</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">8 mm biopsy punch</td>
			<td><a href="http://www.sklarcorp.com/ProductListUS.aspx?DepartmentID=10&amp;CategoryID=96">Sklar&#160;Surgical Instruments</a></td>
			<td>96-1152</td>
			<td>sterile, disposable</td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">12 mm biopsy punch</td>
			<td>Acuderm (via Fisher Scientific)</td>
			<td style="width:124px;">NC9998681</td>
			<td></td>
		</tr>
		<tr height="16">
			<td height="16" style="height:17px;">tissue culture flasks</td>
			<td>Corning</td>
			<td></td>
			<td>cell culture treated</td>
		</tr>
	</tbody>
</table>

construction and characterization of a novel vocal fold bioreactor

In vitro engineering of mechanically active tissues requires the presentation of physiologically relevant mechanical conditions to cultured cells. To emulate the dynamic environment of vocal folds, a novel vocal fold bioreactor capable of producing vibratory stimulations at fundamental phonation frequencies is constructed and characterized. The device is composed of a function generator, a power amplifier, a speaker selector and parallel vibration chambers. Individual vibration chambers are created by sandwiching a custom-made silicone membrane between a pair of acrylic blocks. The silicone membrane not only serves as the bottom of the chamber but also provides a mechanism for securing the cell-laden scaffold. Vibration signals, generated by a speaker mounted underneath the bottom acrylic block, are transmitted to the membrane aerodynamically by the oscillating air. Eight identical vibration modules, fixed on two stationary metal bars, are housed in an anti-humidity chamber for long-term operation in a cell culture incubator. The vibration characteristics of the vocal fold bioreactor are analyzed non-destructively using a Laser Doppler Vibrometer (LDV). The utility of the dynamic culture device is demonstrated by culturing cellular constructs in the presence of 200-Hz sinusoidal vibrations with a mid-membrane displacement of 40 &#181;m. Mesenchymal stem cells cultured in the bioreactor respond to the vibratory signals by altering the synthesis and degradation of vocal fold-relevant, extracellular matrix components. The novel bioreactor system presented herein offers an excellent in vitro platform for studying vibration-induced mechanotransduction and for the engineering of functional vocal fold tissues.

전기 방사에 의해 제조 된 PCL 발판은 마이크론 크기의 중간 세공 및 4.7 μm의 (그림 4A)의 평균 직경이 무작위로 얽혀 섬유가 포함되어 있습니다. 높은 배율에서, 나노 그루브와 모공 개별 섬유 (그림 4B)에서 볼 수 있습니다. 피브로넥틴과 비계의 코팅은 친수성을 향상시키고 초기 세포 부착 / PCL 지지체 (게시되지 않은 관찰)의 확산을 용이하게한다. 원...

Watch this Scientific Journal Video about Construction and Characterization of a Novel Vocal Fold Bioreactor at JoVE.com

Construction and Characterization of a Novel Vocal Fold Bioreactor

A novel vocal fold bioreactor capable of delivering physiologically relevant, vibratory stimulation to cultured cells is constructed and characterized. This dynamic culture device, when combined with a fibrous poly(&#949;-caprolactone) scaffold, creates a vocal fold-mimetic environment that modulates the behaviors of mesenchymal stem cells.

소설 보컬 폴드 생물 반응기의 구조 및 특성

In vitro engineering of mechanically active tissues requires the presentation of physiologically relevant mechanical conditions to cultured cells. To ...

construction-and-characterization-of-a-novel-vocal-fold-bioreactor

Research

JoVE Journal

Bioengineering

9.1K 조회수.  University of Delaware. A novel vocal fold bioreactor capable of delivering physiologically relevant, vibratory stimulation to cultured cells is constructed and characterized. This dynamic culture device, when combined with a fibrous poly(ε-caprolactone) scaffold, creates a vocal fold-mimetic environment that modulates the behaviors of mesenchymal stem cells.

비디오: Construction and Characterization of a Novel Vocal Fold Bioreactor

Engineering Biological-Based Vascular Grafts Using a Pulsatile Bioreactor

Much effort has been devoted to develop and advance the methodology to regenerate functional small-diameter arterial bypasses. In the physiological environment, both mechanical and chemical stimulation are required to maintain the proper development and functionality of arterial vessels1,2.
Bioreactor culture systems developed by our group are designed to support vessel regeneration within a precisely controlled chemo-mechanical environment mimicking that of native vessels. Our bioreactor assembly and maintenance procedures are fairly simple and highly repeatable3,4. Smooth muscle cells (SMCs) are seeded onto a tubular polyglycolic acid (PGA) mesh that is threaded over compliant silicone tubing and cultured in the bioreactor with or without pulsatile stimulation for up to 12 weeks. There are four main attributes that distinguish our bioreactor from some predecessors. 1) Unlike other culture systems that simulate only the biochemical surrounding of native blood vessels, our bioreactor also creates a physiological pulsatile environment by applying cyclic radial strain to the vessels in culture. 2) Multiple engineered vessels can be cultured simultaneously under different mechanical conditions within a controlled chemical environment. 3) The bioreactor allows a mono layer of endothelial cells (EC) to be easily coated onto the luminal side of engineered vessels for animal implantation models. 4) Our bioreactor can also culture engineered vessels with different diameter size ranged from 1 mm to 3 mm, saving the effort to tailor each individual bioreactor to fit a specific diameter size. 
The engineered vessels cultured in our bioreactor resemble native blood vessels histologically to some degree. Cells in the vessel walls express mature SMC contractile markers such as smooth muscle myosin heavy chain (SMMHC)3. A substantial amount of collagen is deposited within the extracellular matrix, which is responsible for ultimate mechanical strength of the engineered vessels5. Biochemical analysis also indicates that collagen content of engineered vessels is comparable to that of native arteries6. Importantly, the pulsatile bioreactor has consistently regenerated vessels that exhibit mechanical properties that permit successful implantation experiments in animal models3,7. Additionally, this bioreactor can be further modified to allow real-time assessment and tracking of collagen remodeling over time, non-invasively, using a non-linear optical microscopy (NLOM)8. To conclude, this bioreactor should serve as an excellent platform to study the fundamental mechanisms that regulate the regeneration of functional small-diameter vascular grafts.

Our group has developed a bioreactor culture system that mimics the physiological pulsatile stresses of the cardiovascular system to regenerate implantable small-diameter vascular grafts.

타악기 생물 반응기를 사용하여 생물 공학 기반 혈관의 이식

Much effort has been devoted to develop and advance the methodology to regenerate functional small-diameter arterial bypasses. In the physiological ...

연구

생체공학

Synthetic, Multi-Layer, Self-Oscillating Vocal Fold Model Fabrication

Sound for the human voice is produced via flow-induced vocal fold vibration. The vocal folds consist of several layers of tissue, each with differing material properties 1. Normal voice production relies on healthy tissue and vocal folds, and occurs as a result of complex coupling between aerodynamic, structural dynamic, and acoustic physical phenomena. Voice disorders affect up to 7.5 million annually in the United States alone 2 and often result in significant financial, social, and other quality-of-life difficulties. Understanding the physics of voice production has the potential to significantly benefit voice care, including clinical prevention, diagnosis, and treatment of voice disorders.
Existing methods for studying voice production include in vivo experimentation using human and animal subjects, in vitro experimentation using excised larynges and synthetic models, and computational modeling. Owing to hazardous and difficult instrument access, in vivo experiments are severely limited in scope. Excised larynx experiments have the benefit of anatomical and some physiological realism, but parametric studies involving geometric and material property variables are limited. Further, they are typically only able to be vibrated for relatively short periods of time (typically on the order of minutes).
Overcoming some of the limitations of excised larynx experiments, synthetic vocal fold models are emerging as a complementary tool for studying voice production. Synthetic models can be fabricated with systematic changes to geometry and material properties, allowing for the study of healthy and unhealthy human phonatory aerodynamics, structural dynamics, and acoustics. For example, they have been used to study left-right vocal fold asymmetry 3,4, clinical instrument development 5, laryngeal aerodynamics 6-9, vocal fold contact pressure 10, and subglottal acoustics 11 (a more comprehensive list can be found in Kniesburges et al. 12)
Existing synthetic vocal fold models, however, have either been homogenous (one-layer models) or have been fabricated using two materials of differing stiffness (two-layer models). This approach does not allow for representation of the actual multi-layer structure of the human vocal folds 1 that plays a central role in governing vocal fold flow-induced vibratory response. Consequently, one- and two-layer synthetic vocal fold models have exhibited disadvantages 3,6,8 such as higher onset pressures than what are typical for human phonation (onset pressure is the minimum lung pressure required to initiate vibration), unnaturally large inferior-superior motion, and lack of a "mucosal wave" (a vertically-traveling wave that is characteristic of healthy human vocal fold vibration).
In this paper, fabrication of a model with multiple layers of differing material properties is described. The model layers simulate the multi-layer structure of the human vocal folds, including epithelium, superficial lamina propria (SLP), intermediate and deep lamina propria (i.e., ligament; a fiber is included for anterior-posterior stiffness), and muscle (i.e., body) layers 1. Results are included that show that the model exhibits improved vibratory characteristics over prior one- and two-layer synthetic models, including onset pressure closer to human onset pressure, reduced inferior-superior motion, and evidence of a mucosal wave.

The methodology for fabricating synthetic vocal fold models is described. The models are life-sized and mimic the multi-layer structure of the human vocal folds. Results show the models to self-oscillate at pressures comparable to lung pressure and demonstrate flow-induced vibratory responses that are similar to those of human vocal folds.

합성, 멀티 레이어, 셀프 진동 보컬 접어 모델 제작

Sound for the human voice is produced via flow-induced vocal fold vibration. The vocal folds consist of several layers of tissue, each with differing ...

Design of a Biaxial Mechanical Loading Bioreactor for Tissue Engineering

We designed a loading device that is capable of applying uniaxial or biaxial mechanical strain to a tissue engineered biocomposites fabricated for transplantation. While the device primarily functions as a bioreactor that mimics the native mechanical strains, it is also outfitted with a load cell for providing force feedback or mechanical testing of the constructs. The device subjects engineered cartilage constructs to biaxial mechanical loading with great precision of loading dose (amplitude and frequency) and is compact enough to fit inside a standard tissue culture incubator. It loads samples directly in a tissue culture plate, and multiple plate sizes are compatible with the system. The device has been designed using components manufactured for precision-guided laser applications. Bi-axial loading is accomplished by two orthogonal stages. The stages have a 50 mm travel range and are driven independently by stepper motor actuators, controlled by a closed-loop stepper motor driver that features micro-stepping capabilities, enabling step sizes of less than 50 nm. A polysulfone loading platen is coupled to the bi-axial moving platform. Movements of the stages are controlled by Thor-labs Advanced Positioning Technology (APT) software. The stepper motor driver is used with the software to adjust load parameters of frequency and amplitude of both shear and compression independently and simultaneously. Positional feedback is provided by linear optical encoders that have a bidirectional repeatability of 0.1 &mu;m and a resolution of 20 nm, translating to a positional accuracy of less than 3 &mu;m over the full 50 mm of travel. These encoders provide the necessary position feedback to the drive electronics to ensure true nanopositioning capabilities. In order to provide the force feedback to detect contact and evaluate loading responses, a precision miniature load cell is positioned between the loading platen and the moving platform. The load cell has high accuracies of 0.15% to 0.25% full scale.

We designed a novel mechanical loading bioreactor that can apply uniaxial or biaxial mechanical strain to a cartilage biocomposite prior to transplantation into an articular cartilage defect.

조직 공학을위한 이축 기계 로딩 생물 반응기의 설계

We  designed a loading device that is capable of applying uniaxial or biaxial  mechanical strain to a tissue engineered biocomposites fabricated for  ...

Operation of a Benchtop Bioreactor

Fermentation systems are used to provide an optimal growth environment for many different types of cell cultures. The ability afforded by fermentors to carefully control temperature, pH, and dissolved oxygen concentrations in particular makes them essential to efficient large scale growth and expression of fermentation products. This video will briefly describe the advantages of the fermentor over the shake flask. It will also identify key components of a typical benchtop fermentation system and give basic instruction on setup of the vessel and calibration of its probes. The viewer will be familiarized with the sterilization process and shown how to inoculate the growth medium in the vessel with culture. Basic concepts of operation, sampling, and harvesting will also be demonstrated. Simple data analysis and system cleanup will also be discussed.

Fermentors are used to increase culture yield and productivity of bioengineered cells. After screening multiple microbial or animal cell culture candidates in shake flasks, the next logical step is to increase the selected culture&#x2019;s biomass with the fermentor. This video demonstrates the setup and operation of a typical benchtop bioreactor system.

벤치 탑 생물 반응기의 운전

Fermentation systems are used to provide an optimal growth environment for many different types of cell cultures. The ability afforded by fermentors to ...

Nonhuman Primate Lung Decellularization and Recellularization Using a Specialized Large-organ Bioreactor

There are an insufficient number of lungs available to meet current and future organ transplantation needs. Bioartificial tissue regeneration is an attractive alternative to classic organ transplantation. This technology utilizes an organ&#39;s natural biological extracellular matrix (ECM) as a scaffold onto which autologous or stem/progenitor cells may be seeded and cultured in such a way that facilitates regeneration of the original tissue. The natural ECM is isolated by a process called decellularization. Decellularization is accomplished by treating tissues with a series of detergents, salts, and enzymes to achieve effective removal of cellular material while leaving the ECM intact. Studies conducted utilizing decellularization and subsequent recellularization of rodent lungs demonstrated marginal success in generating pulmonary-like tissue which is capable of gas exchange in vivo. While offering essential proof-of-concept, rodent models are not directly translatable to human use. Nonhuman primates (NHP) offer a more suitable model in which to investigate the use of bioartificial organ production for eventual clinical use.
The protocols for achieving complete decellularization of lungs acquired from the NHP rhesus macaque are presented. The resulting acellular lungs can be seeded with a variety of cells including mesenchymal stem cells and endothelial cells. The manuscript also describes the development of a bioreactor system in which cell-seeded macaque lungs can be cultured under conditions of mechanical stretch and strain provided by negative pressure ventilation as well as pulsatile perfusion through the vasculature; these forces are known to direct differentiation along pulmonary and endothelial lineages, respectively. Representative results of decellularization and cell seeding are provided.

Whole-organ decellularization produces natural biological scaffolds that may be used for regenerative medicine. The description of a nonhuman primate model of lung regeneration in which whole lungs are decellularized and then seeded with adult stem cells and endothelial cells in a bioreactor that facilitates vascular circulation and liquid media ventilation is presented.

There are an insufficient number of lungs available to meet current and future organ transplantation needs. Bioartificial tissue regeneration is an ...

Investigating the Three-dimensional Flow Separation Induced by a Model Vocal Fold Polyp

The fluid-structure energy exchange process for normal speech has been studied extensively, but it is not well understood for pathological conditions. Polyps and nodules, which are geometric abnormalities that form on the medial surface of the vocal folds, can disrupt vocal fold dynamics and thus can have devastating consequences on a patient&#39;s ability to communicate. Our laboratory has reported particle image velocimetry (PIV) measurements, within an investigation of a model polyp located on the medial surface of an in&#160;vitro driven vocal fold model, which show that such a geometric abnormality considerably disrupts the glottal jet behavior. This flow field adjustment is a likely reason for the severe degradation of the vocal quality in patients with polyps. A more complete understanding of the formation and propagation of vortical structures from a geometric protuberance, such as a vocal fold polyp, and the resulting influence on the aerodynamic loadings that drive the vocal fold dynamics, is necessary for advancing the treatment of this pathological condition. The present investigation concerns the three-dimensional flow separation induced by a wall-mounted prolate hemispheroid with a 2:1 aspect ratio in cross flow, i.e. a model vocal fold polyp, using an oil-film visualization technique. Unsteady, three-dimensional flow separation and its impact of the wall pressure loading are examined using skin friction line visualization and wall pressure measurements.

Vocal fold polyps can disrupt vocal fold dynamics and thus can have devastating consequences on a patient&#39;s ability to communicate. Three-dimensional flow separation induced by a wall-mounted model polyp and its impact on the wall pressure loading are examined using particle image velocimetry, skin friction line visualization, and wall pressure measurements.

모델 보컬 폴드 폴립에 의해 유도된 3차원 유동 분리 조사

The fluid-structure energy exchange process for normal speech has been studied extensively, but it is not well understood for pathological conditions. ...

Characterization of Leukocyte-platelet Rich Fibrin, A Novel Biomaterial

Autologous platelet concentrates represent promising innovative tools in the field of regenerative medicine and have been extensively used in oral surgery. Unlike platelet rich plasma (PRP) that is a gel or a suspension, Leukocyte-Platelet Rich Fibrin (L-PRF) is a solid 3D fibrin membrane generated chair-side from whole blood containing no anti-coagulant. The membrane has a dense three dimensional fibrin matrix with enriched platelets and abundant growth factors. L-PRF is a popular adjunct in surgeries because of its superior handling characteristics as well as its suturability to the wound bed. The goal of the study is to demonstrate generation as well as provide detailed characterization of relevant properties of L-PRF that underlie its clinical success.

Leucocyte-Platelet Rich Fibrin (L-PRF) represents an FDA cleared preparation of autologous platelet concentrates that possesses unique fibrin architecture, enriched platelets and abundant growth factors. Here, we present a protocol for chair-side generation of L-PRF as well as evaluate its mechanical properties including uniaxial testing and suture retention strength testing.

백혈구 - 혈소판 풍부한 섬유소, 소설 생체 재료의 특성

Autologous platelet concentrates represent promising innovative tools in the field of regenerative medicine and have been extensively used in oral ...

A Novel Bioreactor for High Density Cultivation of Diverse Microbial Communities

A novel reactor design, coined a high density bioreactor (HDBR), is presented for the cultivation and study of high density microbial communities. Past studies have evaluated the performance of the reactor for the removal of COD1 and nitrogen species2-4 by heterotrophic and chemoautotrophic bacteria, respectively. The HDBR design eliminates the requirement for external flocculation/sedimentation processes while still yielding effluent containing low suspended solids. In this study, the HDBR is applied as a photobioreactor (PBR) in order to characterize the nitrogen removal characteristics of an algae-based photosynthetic microbial community. As previously reported for this HDBR design, a stable biomass zone was established with a clear delineation between the biologically active portion of the reactor and the recycling reactor fluid, which resulted in a low suspended solid effluent. The algal community in the HDBR was observed to remove 18.4% of total nitrogen species in the influent. Varying NH4+ and NO3- concentrations in the feed did not have an effect on NH4+ removal (n=44, p=0.993 and n=44, p=0.610 respectively) while NH4+ feed concentration was found to be negatively related with NO3- removal (n=44, p=0.000) and NO3- feed concentration was found to be positively correlated with NO3- removal (n=44, p=0.000). Consistent removal of NH4+, combined with the accumulation of oxidized nitrogen species at high NH4+ fluxes indicates the presence of ammonia- and nitrite-oxidizing bacteria within the microbial community.

A novel reactor design, coined a high density bioreactor (HDBR), is presented for the cultivation and study of high density microbial communities. Here, the HDBR is successfully applied in a photobioreactor (PBR) configuration for the study of nitrogen metabolism by a mixed high density algal community.

다양한 미생물 커뮤니티의 고밀도 배양위한 새로운 생물 반응기

A novel reactor design, coined a high density bioreactor (HDBR), is presented for the cultivation and study of high density microbial communities. Past ...

Construction and Setup of a Bench-scale Algal Photosynthetic Bioreactor with Temperature, Light, and pH Monitoring for Kinetic Growth Tests

The optimal design and operation of photosynthetic bioreactors (PBRs) for microalgal cultivation is essential for improving the environmental and economic performance of microalgae-based biofuel production. Models that estimate microalgal growth under different conditions can help to optimize PBR design and operation. To be effective, the growth parameters used in these models must be accurately determined. Algal growth experiments are often constrained by the dynamic nature of the culture environment, and control systems are needed to accurately determine the kinetic parameters. The first step in setting up a controlled batch experiment is live data acquisition and monitoring. This protocol outlines a process for the assembly and operation of a bench-scale photosynthetic bioreactor that can be used to conduct microalgal growth experiments. This protocol describes how to size and assemble a flat-plate, bench-scale PBR from acrylic. It also details how to configure a PBR with continuous pH, light, and temperature monitoring using a data acquisition and control unit, analog sensors, and open-source data acquisition software.

This paper describes the assembly process and operation of a bench-scale photosynthetic bioreactor that can be used, in conjunction with other methods, to estimate pertinent kinetic growth parameters. This system continuously monitors the pH, light, and temperature using sensors, a data acquisition and control unit, and open-source data acquisition software.

키네 성장 시험을위한 온도, 빛 및 pH 모니터링을 갖춘 벤치 규모의 조류 광합성 생물 반응기의 건설 및 설치

The optimal design and operation of photosynthetic bioreactors (PBRs) for microalgal cultivation is essential for improving the environmental and economic ...

Preparation and Characterization of Novel HDL-mimicking Nanoparticles for Nerve Growth Factor Encapsulation

The objective of this article is to introduce preparation and characterization methods for nerve growth factor (NGF)-loaded, high-density, lipoprotein (HDL)-mimicking nanoparticles (NPs). HDLs are endogenous NPs and have been explored as vehicles for the delivery of therapeutic agents. Various methods have been developed to prepare HDL-mimicking NPs. However, they are generally complicated, time consuming, and difficult for industrial scale-up. In this study, one-step homogenization was used to mix the excipients and form the prototype NPs. NGF is a water-soluble protein of 26 kDa. To facilitate the encapsulation of NGF into the lipid environment of HDL-mimicking NPs, protamine USP was used to form an ion-pair complex with NGF to neutralize the charges on the NGF surface. The NGF/protamine complex was then introduced into the prototype NPs. Apolipoprotein A-I was finally coated on the surface of the NPs. NGF HDL-mimicking NPs showed preferable properties in terms of particle size, size distribution, entrapment efficiency, in vitro release, bioactivity, and biodistribution. With the careful design and exploration of homogenization in HDL-mimicking NPs, the procedure was greatly simplified, and the NPs were made scalable. Moreover, various challenges, such as separating unloaded NGF from the NPs, conducting reliable in vitro release studies, and measuring the bioactivity of the NPs, were overcome.

Simple homogenization was used to prepare novel, high-density, lipoprotein-mimicking nanoparticles to encapsulate nerve growth factor. Challenges, detailed protocols for nanoparticle preparation, in vitro characterization, and in vivo studies are described in this article.

신경 성장 인자 인 캡슐 레이션을위한 새로운 HDL 모방 나노 입자의 제조 및 특성 규명

The objective of this article is to introduce preparation and characterization methods for nerve growth factor (NGF)-loaded, high-density, lipoprotein ...