마커스 Pajtler과 로버트 Bohrer의 지원과 조언이 프로토콜의 시각화를위한 귀중한되었습니다.

동물 실험 기관에 의해 규정된 동물 실험 절차와 규정에 대한 동물의 보호 및 사용에 관한 NIH의 지침에 따라, 유럽 커뮤니티 이사회 지침 (86/609/EEC)에 따라 수행다고 저자 상태 관리 및 뒤스부르크 - 에센 대학 (독일)에서위원회 (IACUC)를 사용합니다. 1 부 : (비디오에 표시되지 않음) 설정 (인큐베이터 밖으로 알을 복용하기 전에) 자료 및 솔루션 준비 <ul><li> 계란은 37 11일위한 humidified 인큐베이터에 incubated해야 ° C (100 ° F). </li><li> 70 % 에탄올 스프레이, 깨끗한 접시, 정밀 청소 집게, 4 ° C (40 ° F) 추운 살균 얼음 DMEM 60 ML 무균 주사기 </li><li> 50 ML 무균 코닝 튜브 ° C 감기 DMEM가 1시 1분의 비율 (DMEM 분해 질량)에서 분해 배아를 섞어 4 반 가득해야합니다. 즉시 계란이 여자는 태아의 인큐베이터의 해부 밖으로 가져옵니다 같은 것은 시작되어야한다. </li><li> 압력솥이나 해부 당일 해부 도구를 소독, 20 분 70 %의 에탄올에있는 도구를 잠그다. </li></ul> 2 부 : 칙 태아의 해부 (비디오에서 보여준) <ol><li> 최소한 30 초 동안 70 % 에탄올 스프레이로 incubated 알을 습식 및 과도하게 계란을 흔들림없는 동안 깨끗하고 부드러운 종이 타올로 신중하게 건조. 참고 : 우리는 동시에 60 개 이상의 여자는 배아를 해부하지 권장합니다. </li><li> 날카로운 모서리로부터적으로하여 신중하게 계란을 열고 노른자의 자루 또는 여자는 배아 자체를 파괴하지주의와 배아를 꺼내. </li><li> 빨리 탯줄의 시작에 대한 검색 및 노른자의 자루 또는 작은 혈관을 파괴하지 않고 맑은 포장을 엽니다. 그런 다음 정확한 깨끗한 포셉과 탯줄을 해부하다. </li><li> 난황 자루의 배아를 꺼내주세요. 참고 : 태아가 (비디오에 표시되지 않음) 신속하게 참수해야합니다. </li><li> 4 최소한의 필수적인 매체 배아를 수집 ° C. </li></ol> 파트 3 : 칙 태아의 물에 담가서 부드럽게 함 <ol><li> 60 ML 멸균 주사기 밖으로 플런저를 가져가라. </li><li> 한 60 ML 주사기로 약 10 배아를 전송합니다. </li><li> 분해 물질을 잃을 위험하지 위해 신중하게 다시 주사기로 플런저를 넣어. </li><li> 플런저를 밀어 물에 담가서 부드럽게. </li><li> 준비 코닝 튜브 1시 1분 (DMEM 분해 질량)의 비율로 DMEM과 절반 채워진에서 분해 덩어리를 수집합니다. 10 배아에게 약 25 ML의 볼륨을 사용하는 것은 생산이다. </li><li> 4 45 분 ° C. 위해 쉐이크에 혼합 플레이스 </li></ol> 파트 4 : 칙 배의 원심 분리 - DMEM 서스펜션 <ol><li> 여자의 배아를 전송 - DMEM 서스펜션을 원심 분리 튜브에. </li><li> 배아의 25 MG 당 1 밀리그램의 농도에서 살균 hyaluronidase를 추가합니다. </li><li> 매우 정확하게 테어 원심 분리 튜브. 참고 : 원심 분리가 매우 높은 G - 포스 및 재료 또는 불균형 튜브를 사용하는 경우 원심 분리기가 발생할 수의 손상에서 실행되는이 단계는주의해서 수행해야합니다. 모든 튜브에 대한 소수점은 DMEM으로 누락된 금액을 조정하여 접수해야 다음 세 위치에 대한 기계에 따라 무게 높은 정확도. </li><li> 4 원심 분리기의 온도를 줄이기 ° C. 참고 : 또한 원심 분리의 회전자는 밤 동안 냉각 챔버 또는 냉장고에 보관해야합니다. </li><li> 적어도 180.000 xgx H. 6 시간 원심 분리기 참고 : 단일 단계의 분리에 관한 뛰어난 결과를 얻으려면이 266.000 xgx H.를 사용하여 가능하면 진공 조정을 사용합니다. </li></ol> 5 부 : 칙의 배아 추출의 여과 원심 분리 후, 세 단계가 발견하실 수 있습니다 지방산 조각, 중간에 투명 액상 및 원심 분리 관의 하단에있는 펠렛과 함께 상위 거무죽죽한 액상 참고 사항 :. 떨려 또는 활발한 centrifuged 혼합물의 이동하면되어야합니다 이것은 명확한 중앙 액체 단계를 잃는하여 원심 분리 단계의 결과를 파괴처럼 엄격히 피해야. 참고 : 모든 다음 단계는 층류 공기 흐름 벤치를 사용하여 무균 조건 하에서 수행되어야 : <ol><li> 0.45 μm의의 모공과 필터 시스템에 투명한 액체 단계를 피펫. 다음 연결된 진공 펌프 스위치. 참고 : 하단에있는 펠렛를 만지지 마십시오. 이것은 추출하고 필터 시스템의 차단의 오염으로 이어질 것입니다. </li><li> 0.22 μm의와 연결된 진공 펌프의 스위치를 다시의 모공과 필터 시스템에 사전 추출물을 전송합니다. </li><li> 나누어지는 획득 여자는 배아 추출물에게무균 15 ML 코닝 튜브 인치 </li><li> 신속 -80에서 aliquots를 저장할 ° C (-62 ° F). 참고 : -80에 저장된 경우 최대 2 년 동안 마지막시 ° C. </li></ol>

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관심 없음 충돌 선언하지 않습니다.

이 프로토콜은 과거 10,11에서 설명했습니다 절차의 변경에 따라 달라집니다. 전 출판물 이외에도 우리는 여기 개별 서로 다른 프로세스의 단계 더 확장을 보여줍니다. 이것은 올바른 절차와 신뢰할 수있는 결과를 확보 중요한 세부 실험자을 지원합니다. 해부 과정시 추출의 필수적인 부분이므로 나아가, 우리는 모든 세부 사항에 노른자의 자루의 배아를 복용의 절차를 설명합니다. 아래의 논의로 참고하기 위해 몇 가지 중요한 포인트가 있습니다. 부화가 끝난 후 여자는 태아의 해부는 가능한 빨리 시작되어야합니다. 이전 해부 죽은되고 태아도 부정적인 결과에 영향을 미칠 수 효소 활성화 및 단백질 저하로이 프로토콜의 모든 단계를 사용하지 않아야합니다. 시작하기 전에 몇 가지 수작업에 대한 교육 신속하고 정확하게 실시해야 해부 프로세스의 모든 단계로 수행되어야합니다.을 그것은 부정적인 간섭 단백질의 주목할만한 양의 동행했다 노른자위와 오염을 피하기 위해, 전체 과정의 성공을 위해 중요합니다. 배아는 노른자의 자루 밖으로 찍은 후 그들은 (비디오에 표시되지 않음) 신속하게 참수해야합니다. 온도뿐만 아니라 회전자의 민첩함도 매우 중요합니다. 효소 활성화 및 NCSCM에 필요한 요소의 품질 및 수량의 감소를 방지하기 위해 원심 분리기 및 로터의 온도가 4 감소되어야 ° C. 원심 분리 단계 후에 취소 중앙 액체 단계 NCSCM 위해 필수적인 성장 요소를 포함한 중요한 사전 추출물입니다. 다른 분수의 상당한 분리 및 요소의 농축을 달성하기 위해 우리는보다 낮은 180.000 xgx H.는 민첩함하지 권장 필요 원심 분리 튜브의 상단에있는 맑은 액상 및 거무죽죽한 액체 상 사이의 밀도의 차이는 떨고는 피할 수있는 두 부분으로 remixing을 방지하지 않으므로. 그것의 지방 부분이 곧 필터 시스템의 막히는로 이어질 것처럼 거무죽죽한 액체 단계의 구성 요소, 0.45 μm의 필터 시스템을 통해 filtrated 수 없습니다. 그것의 부분은 필터 시스템의 막히는로 이어질 것처럼 원심의 하단에있는 또한 펠렛은 전혀 손도 안됩니다. 최종 단계에서 NCSCs의 재배에 대한시의 잠재력은 검증되어야한다. 줄기 세포의이 multipotent 과도 인구는 충분한 생산시를 추가하지 않고 미디어에 신속하게 국제 경쟁력을 잃고. 따라서 transgene - 활동에 의존 방식으로 초기 NCSC 마커를 표현하고 NCSC 생물학을 연구하는 데 사용되는 유전자 변형 마우스 라인 (JoMa1)는시 10 때마다 새로 얻은 담당 재배됩니다. NCSCs의 성공적인 재배는시의 적절한 검증을 나타냅니다.

<table><tbody><tr><td>11 days incubated chicken eggs</td><td>Sorries-Trockels Vermehrungszucht GmbH</td><td></td><td></td></tr><tr><td>Alcoholic disinfection spray</td><td>Sch&amp;#252;lke Mayr GmbH</td><td></td><td></td></tr><tr><td>DMEMGlutamax</td><td>GIBCO, by Life Technologies</td><td></td><td></td></tr><tr><td>Syringe sterile (60 ml)</td><td>BD Biosciences</td><td></td><td></td></tr><tr><td>Tubes sterile (50; 15 ml)</td><td>Greiner Bio-One</td><td></td><td></td></tr><tr><td>Pipet tips sterile</td><td>Starlab GmbH</td><td></td><td></td></tr><tr><td>Hyaluronidase typeIV-S</td><td>Sigma-Aldrich</td><td></td><td></td></tr><tr><td>Milipore stericups sterile (0.45; 0.22 &amp;mu;l)</td><td>Fisher Scientific</td><td></td><td></td></tr><tr><td>Pipettes sterile (25; 10; 5 ml)</td><td>Greiner Bio-One</td><td></td><td></td></tr><tr><td>Centrifuge L5-50</td><td>Rotor type Ti-45</td><td></td><td></td></tr></tbody></table>

production of chick embryo extract for the cultivation of murine neural crest stem cells

신경 능선은 embryogenesis 동안 신경 ectoderm에서 발생에만 일시적으로 지속. 초기 실험은 이미 신경 문장 하나의 불가분의 일부로 pluripotent 전구 세포를 침대. Phenotypically, 신경 크레스트 줄기 세포 (NCSC)는 동시에 신경 문장 2,3,4,5으로부터 마이 그 레이션하는 동안 p75 (낮은 affine 신경 성장 인자 수용체, LNGFR)와 SOX10 표현에 의해 정의됩니다. 이러한 전구 세포는 평활근 세포, chromaffin 세포, 뉴런과 glial 세포뿐만 아니라, melanocytes, 연골과 뼈 6,7,8,9로 구별하실 수 있습니다. 시험 관내, 특별한 신경 크레스트 줄기 세포 매체 (NCSCM)에 NCSC를 육성하기 위해서는 10 필요합니다. NCSCM의 가장 복잡한 부분은 NCSC뿐만 아니라 신경 explants 다른 유형의 성장 요인의 필수 소스를 대표하는 여자는 배아 추출물 (시)의 준비입니다. 시 옆에 다른 NCSCM 재료는 상업적으로 사용할 수 있습니다. 그것은 고립, 물에 담가서 부드럽게 함, 원심 분리 및 여과 공정으로 도전하는 세부 사항에 대해 알고 높은 중요 CCE 사용하여 실험실 표준 장비를 생산. 이 프로토콜에서는 순수하고 고품질시의 최대 금액을 생산하는 정확한 방법을 설명합니다.

Watch this Scientific Journal Video about Production of Chick Embryo Extract for the Cultivation of Murine Neural Crest Stem Cells at JoVE.com

Production of Chick Embryo Extract for the Cultivation of Murine Neural Crest Stem Cells

체외에서 신경 크레스트 줄기 세포 (NCSC)을 육성하기 위해 특수 매체 (NCSCM)가 필요합니다. NCSCM의 필수적인 부분은 여자의 배아 추출물 (시)입니다. 우리는 여기서 고립, 물에 담가서 부드럽게 함, 원심 분리 및 여과 공정과 같은 세부 사항을 포함하여 순수하고 고품질시의 최대 금액을 생산하는 정확한 방법을 설명합니다.

Murine 신경 크레스트 줄기 세포의 배양 칙의 배아 추출 생산

The neural crest arises from the neuro-ectoderm during embryogenesis and persists only temporarily. Early experiments already proofed pluripotent ...

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Nanyang Technological University

Research

JoVE Journal

Neuroscience

12.7K 조회수.  University Children's Hospital Essen. 체외에서 신경 크레스트 줄기 세포 (NCSC)을 육성하기 위해 특수 매체 (NCSCM)가 필요합니다. NCSCM의 필수적인 부분은 여자의 배아 추출물 (시)입니다. 우리는 여기서 고립, 물에 담가서 부드럽게 함, 원심 분리 및 여과 공정과 같은 세부 사항을 포함하여 순수하고 고품질시의 최대 금액을 생산하는 정확한 방법을 설명합니다.

비디오: Production of Chick Embryo Extract for the Cultivation of Murine Neural Crest Stem Cells

Culture of Mouse Neural Stem Cell Precursors

Primary neural stem cell cultures are useful for studying the mechanisms underlying central nervous system development. Stem cell research will increase our understanding of the nervous system and may allow us to develop treatments for currently incurable brain diseases and injuries. In addition, stem cells should be used for stem cell research aimed at the detailed study of mechanisms of neural differentiation and transdifferentiation and the genetic and environmental signals that direct the specialization of the cells into particular cell types.  This video demonstrates a technique used to disaggregate cells from the embryonic day 12.5 mouse dorsal forebrain. The dissection procedure includes harvesting E12.5 mouse embryos from the uterus, removing the "skin" with fine dissecting forceps and finally isolating pieces of cerebral cortex. Following the dissection, the tissue is digested and mechanically dissociated. The resuspended dissociated cells are then cultured in "stem cell" media that favors growth of neural stem cells.

This video describes the method used for isolation of neuroprecursors from the developing cortex of embryonic mice. The procedure for removing embryos from the uterus, dissecting the cortical tissue, and digesting the isolated cerebral cortex is shown.

마우스 신경 줄기 세포 엽 성의 전구 물질의 문화

Primary neural stem cell cultures are useful for studying the mechanisms underlying central nervous system development. Stem cell research will increase ...

연구

생물학

Neural Differentiation of Mouse Embryonic Stem Cells in Serum-free Monolayer Culture

The ability to differentiate mouse embryonic stem cells (ESC) to neural progenitors allows the study of the mechanisms controlling neural specification as well as the generation of mature neural cell types for further study. In this protocol we describe a method for the differentiation of ESC to neural progenitors using serum-free, monolayer culture. The method is scalable, efficient and results in production of ~70% neural progenitor cells within 4 - 6 days. It can be applied to ESC from various strains grown under a variety of conditions. Neural progenitors can be allowed to differentiate further into functional neurons and glia or analyzed by microscopy, flow cytometry or molecular techniques. The differentiation process is amenable to time-lapse microscopy and can be combined with the use of reporter lines to monitor the neural specification process. We provide detailed instructions on media preparation and cell density optimization to allow the process to be applied to most ESC lines and a variety of cell culture vessels.

This protocol describes in detail the method for generating neural progenitors from embryonic stem cells using a serum-free monolayer method. These progenitors can be used to derive mature neural cell types or to study the process of neural specification and is amenable to multiwell format scaling for compound screening.

혈청이없는 단층 문화 마우스 배아 줄기 세포의 신경 분화

The ability to differentiate mouse embryonic stem cells (ESC) to neural progenitors allows the study of the mechanisms controlling neural specification as ...

발생학

Generation of Neural Stem Cells from Discarded Human Fetal Cortical Tissue

Neural stem cells (NSCs) reside along the ventricular zone neuroepithelium during the development of the cortical plate. These early progenitors ultimately give rise to intermediate progenitors and later, the various neuronal and glial cell subtypes that form the cerebral cortex. The capacity to generate and expand human NSCs (so called neurospheres) from discarded normal fetal tissue provides a means with which to directly study the functional aspects of normal human NSC development 1-5. This approach can also be directed toward the generation of NSCs from known neurological disorders, thereby affording the opportunity to identify disease processes that alter progenitor proliferation, migration and differentiation 6-9. We have focused on identifying pathological mechanisms in human Down syndrome NSCs that might contribute to the accelerated Alzheimer's disease phenotype 10,11. Neither in vivo nor in vitro mouse models can replicate the identical repertoire of genes located on human chromosome 21. 
Here we use a simple and reliable method to isolate Down syndrome NSCs from aborted human fetal cortices and grow them in culture. The methodology provides specific aspects of harvesting the tissue, dissection with limited anatomical landmarks, cell sorting, plating and passaging of human NSCs. We also provide some basic protocols for inducing differentiation of human NSCs into more selective cell subtypes.

A simple and reliable method on isolation and culture of neural stem cells from discarded human fetal cortical tissue is described. Cultures derived from known human neurological disorders can be used for characterization of pathological cellular and molecular processes, as well as provide a platform to assess pharmacological efficacy.

폐기 인간 태아 두피 조직에서 신경 줄기 세포의 생성

Neural stem cells (NSCs) reside along the ventricular zone neuroepithelium during the development of the cortical plate. These early progenitors ...

신경과학

Assay for Neural Induction in the Chick Embryo

The chick embryo is a valuable tool in the study of early embryonic development. Its transparency, accessibility and ease of manipulation, make it an ideal tool for studying the formation and initial patterning of the nervous system. This video demonstrates how to graft organizer tissue into a host, a method by which Hensen s node (the organizer in the chick embryo) is grafted to a host competent ectoderm. The organizer graft instructs overlying na ve tissue to adopt a neural fate via neural inducing signals. This mechanism is referred to as neural induction, and constitutes the initial step in the formation of brain and spinal cord in amniotes. This method is essentially used for the characterization of putative neural inducing molecules in chick. This video demonstrates the different steps in the assay for neural induction; First, the donnor embryo is explanted and pinned on a dish. Then, the host embryo is prepared for New culture. The graft is excised and transplanted to the host area pellucida margin. The host is cultured for 18-22 hrs. The assembly is fixed and processed for further applications (e.g. in situ hybridization). This method was originally devised by Waddington 1,2 and Gallera 3,4.

Neural induction is the first step in the formation of the brain. It is a mechanism by which Hensen's node (organizer), instructs adjacent tissue to adopt a neural fate, i.e. to give rise to the nervous system. This video demonstrates an assay for neural induction in chick embryo.

칙의 배아에서 신경 유도에 대한 분석

The chick embryo is a valuable tool in the study of early embryonic development. Its transparency, accessibility and ease of manipulation, make it an ...

Isolation and Culture of Neural Crest Cells from Embryonic Murine Neural Tube

The embryonic neural crest (NC) is a multipotent progenitor population that originates at the dorsal aspect of the neural tube, undergoes an epithelial to mesenchymal transition (EMT) and migrates throughout the embryo, giving rise to diverse cell types 1-3. NC also has the unique ability to influence the differentiation and maturation of target organs4-6. When explanted in vitro, NC progenitors undergo self-renewal, migrate and differentiate into a variety of tissue types including neurons, glia, smooth muscle cells, cartilage and bone. 
NC multipotency was first described from explants of the avian neural tube7-9. In vitro isolation of NC cells facilitates the study of NC dynamics including proliferation, migration, and multipotency. Further work in the avian and rat systems demonstrated that explanted NC cells retain their NC potential when transplanted back into the embryo10-13. Because these inherent cellular properties are preserved in explanted NC progenitors, the neural tube explant assay provides an attractive option for studying the NC in vitro. 
To attain a better understanding of the mammalian NC, many methods have been employed to isolate NC populations. NC-derived progenitors can be cultured from post-migratory locations in both the embryo and adult to study the dynamics of post-migratory NC progenitors11,14-20, however isolation of NC progenitors as they emigrate from the neural tube provides optimal preservation of NC cell potential and migratory properties13,21,22. Some protocols employ fluorescence activated cell sorting (FACS) to isolate a NC population enriched for particular progenitors11,13,14,17. However, when starting with early stage embryos, cell numbers adequate for analyses are difficult to obtain with FACS, complicating the isolation of early NC populations from individual embryos. Here, we describe an approach that does not rely on FACS and results in an approximately 96% pure NC population based on a Wnt1-Cre activated lineage reporter23. 
The method presented here is adapted from protocols optimized for the culture of rat NC11,13. The advantages of this protocol compared to previous methods are that 1) the cells are not grown on a feeder layer, 2) FACS is not required to obtain a relatively pure NC population, 3) premigratory NC cells are isolated and 4) results are easily quantified. Furthermore, this protocol can be used for isolation of NC from any mutant mouse model, facilitating the study of NC characteristics with different genetic manipulations. The limitation of this approach is that the NC is removed from the context of the embryo, which is known to influence the survival, migration and differentiation of the NC2,24-28.

Isolation of embryonic neural crest from the neural tube facilitates the use of in vitro methods for studying migration, self-renewal, and multipotency of neural crest.

배아 Murine 신경 튜브에서 신경 크레스트 세포의 분리 및 문화

The embryonic neural crest (NC) is a multipotent progenitor population that originates at the dorsal aspect of the neural tube, undergoes an epithelial to ...

Xenotransplantation of Human Stem Cells into the Chicken Embryo

The chicken embryo is a classical animal model for studying normal embryonic and fetal development and for xenotransplantation experiments to study the behavior of cells in a standardized in vivo environment. The main advantages of the chicken embryo include low cost, high accessibility, ease of surgical manipulation and lack of a fully developed immune system. Xenotransplantation into chicken embryos can provide valuable information about cell proliferation, differentiation and behavior, the responses of cells to signals in defined embryonic tissue niches, and tumorigenic potential. Transplanting cells into chicken embryos can also be a step towards transplantation experiments in other animal models. Recently the chicken embryo has been used to evaluate the neurogenic potential of human stem and progenitor cells following implantation into neural anlage1-6. In this video we document the entire procedure for transplanting human stem cells into the developing central nervous system of the chicken embryo. The procedure starts with incubation of fertilized eggs until embryos of the desired age have developed. The eggshell is then opened, and the embryo contrasted by injecting dye between the embryo and the yolk. Small lesions are made in the neural tube using microsurgery, creating a regenerative site for cell deposition that promotes subsequent integration into the host tissue. We demonstrate injections of human stem cells into such lesions made in the part of the neural tube that forms the hindbrain and the spinal cord, and into the lumen of the part of the neural tube that forms the brain. Systemic injections into extraembryonic veins and arteries are also demonstrated as an alternative way to deliver cells to vascularized tissues including the central nervous system. Finally we show how to remove the embryo from the egg after several days of further development and how to dissect the spinal cord free for subsequent physiological, histological or biochemical analyses.

In this paper we present a method for transplanting human stem cells into various regions of the central nervous system of the chicken embryo. This provides an in vivo model for assessing the proliferation and differentiation of various types of human stem cells in embryonic tissue environments.

치킨의 배아에 인간 줄기 세포의 Xenotransplantation

The chicken embryo is a classical animal model for studying normal embryonic and fetal development and for xenotransplantation experiments to study the ...

Preparation and Morphological Analysis of Chick Cranial Neural Crest Cell Cultures

During vertebrate development, neural crest cells (NCCs) migrate extensively and differentiate into various cell types that contribute to structures like the craniofacial skeleton and the peripheral nervous system. While it is critical to understand NCC migration in the context of a 3D embryo, isolating migratory cells in 2D culture facilitates visualization and functional characterization, complementing embryonic studies. The present protocol demonstrates a method for isolating chick cranial neural folds to generate primary NCC cultures. Migratory NCCs emerge from neural fold explants plated onto a fibronectin-coated substrate. This results in dispersed, adherent NCC populations that can be assessed by staining and quantitative morphological analyses. This simplified culture approach is highly adaptable and can be combined with other techniques. For example, NCC emigration and migratory behaviors can be evaluated by time-lapse imaging or functionally queried by including inhibitors or experimental manipulations of gene expression (e.g., DNA, morpholino, or CRISPR electroporation). Because of its versatility, this method provides a powerful system for investigating cranial NCC development.

This versatile protocol describes the isolation of premigratory neural crest cells (NCCs) through the excision of cranial neural folds from chick embryos. Upon plating and incubation, migratory NCCs emerge from neural fold explants, allowing for assessment of cell morphology and migration in a simplified 2D environment.

병아리 두개골 신경 볏 세포 배양의 준비 및 형태학적 분석

During vertebrate development, neural crest cells (NCCs) migrate extensively and differentiate into various cell types that contribute to structures like ...

Isolation and Culture of Embryonic Mouse Neural Stem Cells

Neural stem cells (NSCs) are multipotent and can give rise to the three major cell types of the central nervous system (CNS). In vitro culture and expansion of NSCs provide a suitable source of cells for neuroscientists to study the function of neurons and glial cells along with their interactions. There are several reported techniques for the isolation of neural stem cells from adult or embryo mammalian brains. During the microsurgical operation to isolate NSCs from different regions of the embryonic CNS, it is very important to reduce the damage to the brain cells to obtain the highest ratio of live and expandable stem cells. A possible technique for stress reduction during isolation of these cells from the mouse embryo brain is the reduction of surgical time. Here, we demonstrate a developed technique for rapid isolation of these cells from the E13 mouse embryo ganglionic eminence. Surgical procedures include harvesting E13 mouse embryos from the uterus, cutting the frontal fontanelle of the embryo with a bent needle tip, extracting the brain from the skull, microdissection of the isolated brain to harvest the ganglionic eminence, dissociation of the harvested tissue in NSC medium to gain a single cell suspension, and finally plating cells in suspension culture to generate neurospheres.

Here, we introduce a novel microsurgical technique for the isolation of neural stem cells from the E13 mouse embryo ganglionic eminence.

절연 및 마우스 배아 신경 줄기 세포의 문화

Neural stem cells (NSCs) are multipotent and can give rise to the three major cell types of the central nervous system (CNS). In vitro culture and ...

A Technique for Generating Neural Progenitor Cells from Mouse Embryonic Stem Cells

Source: Mao, X., et al. <a href="https://app-jove-com.remotexs.ntu.edu.sg/v/61190">Neuronal Differentiation from Mouse Embryonic Stem Cells In vitro.</a> J. Vis. Exp. (2020).
This video demonstrates a method for the in vitro generation of neural progenitor cells (NPCs) from mouse embryonic stem cells (mESCs). The mESCs are cultured in suspension to form embryoid bodies (EBs), which are then guided to differentiate into NPCs via retinoic acid treatment in an adherent culture.

마우스 배아 줄기 세포에서 신경 전구 세포를 생성하는 기술

1. Mouse embryonic stem cell culture

	Prepare 0.1% gelatin coated cell culture dishes or plates.
	
		Add 2 mL of sterilized 0.1% gelatin (0.1% w/v in ...

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Neuronal Culture Techniques

Advanced Neuronal Models

Isolating Cranial Neural Folds From a Chick Embryo for a Neural Crest Cell Culture

Source: Jacques-Fricke, B. T., et al. <a href="https://app-jove-com.remotexs.ntu.edu.sg/v/63799">Preparation and Morphological Analysis of Chick Cranial Neural Crest Cell Cultures.</a> J. Vis. Exp. (2022).
This video illustrates a technique for isolating cranial neural folds from the midbrain region of chick embryos. It presents a detailed dissection process of the neural folds, their placement, and attachment on fibronectin-coated coverslips, followed by the observation of neural crest cell migration from the neural folds.

신경 능선 세포 배양을 위해 병아리 배아에서 두개골 신경 주름 분리

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. ...