作者要感谢詹姆斯Oesterling流式细胞仪的帮助。这项研究是由美国国家癌症研究所（R01-CA178380，ZY）和美国国家研究院​​博士后培训补助（5T32CA009035-37，LWY）的资金支持。

已按照批准的Fox Chase癌症中心的动物护理和使用委员会的程序在执行该协议利用动物。 1，准备工具，解决方案和盖玻片<ol><li>清洁两套＃5细镊子，一组7号细弯钳，一个大的手术剪刀，1 microdissecting剪一小铲和一个带孔的勺子。将仪器的自密封消毒袋和高压锅。请眼袋仪器，直到准备使用。 </li><li>制备3％低熔点琼脂糖溶液。 <ol><li>添加1.5克琼脂糖至50毫升无菌贝科的磷酸盐缓冲液（DPBS）中，然后放入微波炉，加热至泡沫出现。漩涡烧瓶中，并再加热，直到所有的琼脂糖溶解。混合与搅拌棒的解决方案之前，如果加热团块出现。 </li><li>在37℃水浴商店解决方案，直到需要。 </li><li>对于长期存储媒体E，存储以供将来使用的琼脂糖溶液，在室温或4℃下进行几个月。再热的微波液化。在琼脂糖溶液中反复加热，加热以保持其与温暖无菌蒸馏水初始重量之前称量烧瓶中。 </li></ol></li><li>准备如前所述2木瓜蛋白酶类细胞分离通过以下方法解决: <ol><li>木瓜蛋白酶溶液，木瓜蛋白酶（100单位/毫升），半胱氨酸（0.2毫克/毫升）和DNA酶（250 U / ml）的无菌酚红含-DPBS。 </li><li>在无菌的无酚红的含DPBS，大豆胰蛋白酶抑制剂（8毫克/毫升））和DNA酶（250单位/毫升），用牛血清白蛋白的卵类粘蛋白溶液（8毫克/毫升BSA）。 过滤消毒，这两种解决方案，并调节pH值，返回酚红含的DPBS原来的颜色。 </li></ol></li><li>制备的细胞培养基（NB / B27）:补充基础培养基用1mM丙酮酸钠，2mM的L-谷氨酰胺，1％青霉素 - 链霉素（P / S）和B27补充剂。滤清器ř消毒和避光。在使用前平衡在37℃的介质和5％的CO 2。 </li><li>制备FACS缓冲液:5％的胎牛血清（FBS）的DPBS中。添加2.5毫升胎牛血清47.5毫升DPBS。 </li><li>制备聚-D-赖氨酸（PDL）包被的盖玻片上。 <ol><li>新高压灭菌盖玻片。 </li><li>外套与PDL中的盖玻片（在无菌蒸馏水中100微克/毫升）并孵育2小时，在37℃或室温下放置过夜。 </li><li>洗涤盖玻片，用无菌蒸馏水。采用NB / B27传媒洗一次。让盖玻片在使用前组织培养罩完全干燥。 </li></ol></li></ol> 2，解剖组织切片和准备<ol><li>上解剖当天，擦拭用70％乙醇和盖的吸水垫的解剖区域。 </li><li>从灭菌袋中取出解剖工具。添加冰冷，无菌DPBS / 1％P / S的小培养皿，置于冰上。 </li><li>杀头的P4 MATH1-GFP / 巢蛋白CFP小鼠的幼崽有大的手术剪刀然后按住低着头用细弯钳。取出皮肤，轻轻剥离用＃5的头骨从头骨使用刮刀细钳子然后舀出脑，并将其转移至含有DPBS / 1％P / S的培养皿中。 <ol><li>使用5号细镊子，小心地从大脑的其余部分分离小脑。一定要尽快解剖1小狗在同一时间成为可能。 </li><li>收集至少8小脑，以获得足够的非符合资格人士的分子和功能分析。 </li></ol></li><li>填充嵌入模具测量2×2×2厘米，3％低熔点琼脂糖溶液。确保琼脂糖的温度不超过37℃。 </li><li>通过在实验室组织轻轻涂抹去除小脑多余的液体。将小脑成模在垂直位置。放置在冰上的模具以允许其快速固化。使用每块〜4小脑。 </li><li>获得李咏组织切片用振动叶片vibratome。修剪含小脑，用刀片琼脂糖块。胶琼脂糖块到vibratome板与小脑定向在垂直位置。 </li><li>填补vibratome的用冰冷却的无菌DPBS / 1％P / S和地方冰下方的托盘上。 <ol><li>切成600微米的部分通过小脑的整个长度。收集使用穿孔勺子从冰盘的部分。地方的部分在培养皿中填充用DPBS / 1％P / S在冰上。 </li></ol></li></ol> 3，显微切割和细胞分离<ol><li>将含有荧光解剖显微镜下组织切片的培养皿中。 <ol><li>小心地用细镊子小脑节的其余部分由CFP +分子层和GFP +东瀛之间的夹层中（参见虚线图1）分离东瀛。小心不包括任何部分的分子层的，这将导致在污染的巢蛋白表达伯格曼胶质细胞。 </li><li>尽可能快地完成这个步骤，并保持组织在冰上，以避免细胞死亡。 </li><li>在进行下一步骤之前，除去周围的组织的琼脂糖。 </li></ol></li><li>消化使用如前所述2，得到单细胞悬浮液中的木瓜蛋白酶的基于协议的显微EGL组织。 <ol><li>重悬的细胞在FACS缓冲液用于收集通过FACS CFP-阳性细胞。 </li></ol></li></ol> 4，细胞分选及镀层<ol><li>使用无菌，高速流式细胞仪包含适当的CFP过滤器和收集细胞，流式细胞仪缓冲冰上排序细胞CFP荧光。获得每个P4小脑大约100,000非符合资格人士。 </li><li>离心细胞，在300×g离心5分钟。立即进行分子分析，或文化进行进一步的实验用的细胞。 </li><li>向培养的细胞，重新悬浮细胞在预热NB / B27米EDIA和盘上的PDL涂覆的盖玻片如前所述2。 </li></ol>

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作者什么都没有透露。

此处所描述的显微切割的方法是能够分离的活细胞中的分子和功能分析使用的纯群体的第一个过程。这表现在Li 等人9， 通过该方法得到的NEP可以培养并掺入了各种实验，并由于其高纯度的，也可用于微阵列分析。 花时间去精通这种显微切割技术，这是非常重要的。的NEP位于EGL的深区域中的非常接近的分子层，其中巢蛋白表达伯格曼胶质细胞可能污染NEP...

小脑由多个细胞层，每层含有不同的细胞类型。在开发过程中，东瀛包含增殖颗粒神经元前体（国民生产总值），而分子层和浦肯野层包含伯格曼胶质细胞和浦肯野神经元，分别。小脑深处位于脑白质，其中包含神经干细胞（NSCs）和少突胶质细胞1。 音猬蛋白（Shh的）在调节小脑发育的关键作用，特别是，它促进的GNP的增殖通过与其受体结合补丁（PTC），对Shh信号2-4的负调节物。的Shh信号传导的异常活化产生髓母细胞瘤（MB），在孩子5,6中最常见的恶性脑肿瘤。 PTC突变MB的转基因小鼠模型是MB的学习功能强大的工具，开发肿瘤类似人类MB 7,8。使用这些小鼠，它被发现的GNP是起源于刺猬型MB 7的细胞。此外，除的GNP，我们最近发现的显影小脑也可以产生MB的EG​​L内的神经元祖细胞的独特群体。这些细胞表达高水平的VI型中间丝蛋白，巢蛋白，并且被称为网络设备供应商9。的NEP位于EGL的深部内并仅在新生儿小脑发育存在短暂。他们致力于颗粒细胞系作为国民生产总值却是不同的，因为它们是静态和不表达MATH1，一个完善的标志传统国民生产总值10。此外，网络设备供应商产生MB比更有效地活化Shh信号通路9，这使得它们的新型原点为MB的肿瘤发生后的GNP。 我们以前的显微切割技术分离出小脑东瀛非符合资格人士在4（P4）日龄这里说明9。通过组织直接镜检可视显微允许小脑东瀛的具体解剖。这是必要的，因为巢是由神经干细胞在小脑白质和伯格曼胶质细胞中的分子层1,7,11,12也表示，这是至关重要的，这些细胞不被包括在内，因为他们混淆分析。细胞显微组织分离可立即用于分子分析，或者它们可以被培养用于进一步的应用。 为了有助于具体microdissecting的EGL和进一步纯化的NEP，MATH1-GFP（绿色荧光蛋白）的小鼠进行杂交以巢蛋白-CFP（青色荧光蛋白）的小鼠。转录因子MATH1通过的GNP和GFP表达特异性表达是在EGL 9,13清晰可见。 巢蛋白-CFP小鼠表达CFP的核形式，并且可以很容易地可视化的分子层9,14。总之，GFP和CFP的表达创造边界东瀛的显微切割（ 见图1）。 CFP-阳性的NEP，然后通过FACS分离，继解剖EGLS的酶解。 目前，使用最多的方法也从东瀛分离细胞是整个小脑组织15,16 Percoll密度梯度离心法。这个方法，但是，不能完全排除从分子层和白质巢蛋白+细胞群中，因此不能被用于网络设备供应商的研究。激光捕获显微切割，其使用激光能量的传递，以除去所关注的细胞，是一种用于多相组织17,18内分离特定的细胞，但捕获的细胞仅可用于DNA，RNA和蛋白质的回收和不是另一种方法能够进行培养。 该协议提供了一种专门隔离东瀛非符合资格人士的活着，纯粹的人口。这种技术也可以应用于具有任何可识别的解剖结构或荧光标记细胞/区域不同的组织类型。因此，将这一新颖显微切割技术进入细胞的分离协议的主要优点是，细胞可以从特定组织区域进行分离，以消除污染邻近的组织和细胞可以立即被收集用于分析或培养的其他应用程序。

<table><tbody><tr><td>Neurobasal media</td><td>Gibco</td><td>21103-049</td><td></td></tr><tr><td>PDL</td><td>Millipore</td><td>A-003-E</td><td></td></tr><tr><td>Ultra pure low melting temperature agarose</td><td>Invitrogen</td><td>16520-050</td><td></td></tr><tr><td>DPBS</td><td>Gibco</td><td>14190144</td><td></td></tr></tbody></table>

isolation of distinct cell populations from the developing cerebellum by microdissection

显微切割是一种新技术，可以分离组织的特定区域，消除污染邻近地区的细胞来源。在这项研究是第一个使用这种方法巢蛋白表达祖商（NEP），细胞在小脑外部生发层（EGL）的新发现的种群。在组合使用显微切割用荧光激活细胞分选（FACS），棉结的纯群体是从在EGL常规颗粒神经元前体和从其它污染巢蛋白表达细胞在小脑分开收集。未经显微切割，棉结的功能分析将不会有可能与可用的电流的方法，如Percoll密度梯度离心法和激光捕获显微解剖。该技术还可以应用于用于与包含任一可识别的区域或荧光标记细胞的各种组织中的使用。最重要的是，这主要microdissectio优势Ñ​​技术是分离的细胞是活的，并且可以被培养了进一步的实验，这是目前不可能与其他所描述的方法。

如图1A所示 ，小脑切片从P4 MATH1-GFP / 巢蛋白-CFP小鼠制备的，其中，所述小脑EGL主要是由绿色荧光蛋白表达的GNP和分子层中富集由CFP-阳性的神经胶质细胞。 （; 图1A中沿虚线）以分离的NEP位于EGL的深部的，切片的EGL和分子层之间的微切割。剖分EGLS（ 图1B）然后收集用酶解随后通过FACS。 正如预期的那样，细胞在解剖东瀛的大?...

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Isolation of Distinct Cell Populations from the Developing Cerebellum by Microdissection

巢蛋白表达祖细胞是在发展中国家小脑的神经元祖细胞的新发现的种群。用在这里用荧光激活细胞分选组合呈现的显微切割技术中，该细胞群可与任何其他小脑区域污染被纯化和可以培养用于进一步研究。

从开发小脑显微切割分离的不同的细胞群

Microdissection is a novel technique that can isolate specific regions of a tissue and eliminate contamination from cellular sources in adjacent areas. ...

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9.5K 次观看.  Temple University Health System. 巢蛋白表达祖细胞是在发展中国家小脑的神经元祖细胞的新发现的种群。用在这里用荧光激活细胞分选组合呈现的显微切割技术中，该细胞群可与任何其他小脑区域污染被纯化和可以培养用于进一步研究。 

视频: Isolation of Distinct Cell Populations from the Developing Cerebellum by Microdissection

Ex Vivo Culture of Chick Cerebellar Slices and Spatially Targeted Electroporation of Granule Cell Precursors

The cerebellar external granule layer (EGL) is the site of the largest transit amplification in the developing brain, and an excellent model for studying neuronal proliferation and differentiation. In addition, evolutionary modifications of its proliferative capability have been responsible for the dramatic expansion of cerebellar size in the amniotes, making the cerebellum an excellent model for evo-devo studies of the vertebrate brain. The constituent cells of the EGL, cerebellar granule progenitors, also represent a significant cell of origin for medulloblastoma, the most prevalent paediatric neuronal tumour. Following transit amplification, granule precursors migrate radially into the internal granular layer of the cerebellum where they represent the largest neuronal population in the mature mammalian brain. In chick, the peak of EGL proliferation occurs towards the end of the second week of gestation. In order to target genetic modification to this layer at the peak of proliferation, we have developed a method for genetic manipulation through ex vivo electroporation of cerebellum slices from embryonic Day 14 chick embryos. This method recapitulates several important aspects of in vivo granule neuron development and will be useful in generating a thorough understanding of cerebellar granule cell proliferation and differentiation, and thus of cerebellum development, evolution and disease.

Ex Vivo Culture of Chick Cerebellar Slices and Spatially Targeted Electroporation of Granule Cell Precursors

The cerebellar external granule layer is the site of the largest transit amplification in the developing brain. Here, we present a protocol to target genetic modification to this layer at the peak of proliferation using ex vivo electroporation and culture of cerebellar slices from embryonic Day 14 chick embryos.

鸡小脑切片的体外培养和空间上有针对性的颗粒细胞前体电穿孔

The cerebellar external granule layer (EGL) is the site of the largest transit amplification in the developing brain, and an excellent model for studying ...

科研

发育生物学

Isolation and Culture of Post-Natal Mouse Cerebellar Granule Neuron Progenitor Cells and Neurons 

The cerebellar cortex is a well described structure that provides unique opportunities for studying neuronal properties and development1,2. Of the cerebellar neuronal types (granule cells, Purkinje cells and inhibitory interneurons), granule neurons are by far the most numerous and are the most abundant type of neurons in the mammalian brain. In rodents, cerebellar granule neurons are generated during the first two post-natal weeks from progenitor cells in the outermost layer of the cerebellar cortex, the external granule layer (EGL). The protocol presented here describes techniques to enrich and culture granule neurons and their progenitor cells from post-natal mouse cerebellum. We will describe procedures to obtain cultures of increasing purity3,4, which can be used to study the differentiation of proliferating progenitor cells into granule neurons5,6. Once the progenitor cells differentiate, the cultures also provide a homogenous population of granule neurons for experimental manipulation and characterization of phenomena such as synaptogenesis, glutamate receptor function7, interaction with other purified cerebellar cells8,9 or cell death7.

Isolation and Culture of Post-Natal Mouse Cerebellar Granule Neuron Progenitor Cells and Neurons

Here we present a method to isolate and culture cerebellar granule neuron progenitor cells and cerebellar granule neurons from postnatal mouse.

产后小鼠小脑颗粒神经元祖细胞和神经元的分离和培养

The cerebellar cortex is a well described structure that provides unique opportunities for studying neuronal properties and development1,2. Of the ...

生物学

Isolation of Cortical Microglia with Preserved Immunophenotype and Functionality From Murine Neonates

Isolation of microglia from CNS tissue is a powerful investigative tool used to study microglial biology ex vivo. The present method details a procedure for isolation of microglia from neonatal murine cortices by mechanical agitation with a rotary shaker. This microglia isolation method yields highly pure cortical microglia that exhibit morphological and functional characteristics indicative of quiescent microglia in normal, nonpathological conditions in vivo. This procedure also preserves the microglial immunophenotype and biochemical functionality as demonstrated by the induction of morphological changes, nuclear translocation of the p65 subunit of NF-&#954;B (p65), and secretion of the hallmark proinflammatory cytokine, tumor necrosis factor-&#945; (TNF-&#945;), upon lipopolysaccharide (LPS) and Pam3CSK4 (Pam) challenges. Therefore, the present isolation procedure preserves the immunophenotype of both quiescent and activated microglia, providing an experimental method of investigating microglia biology in ex vivo conditions.

One key to successful investigation of microglial biology is the preservation of microglial immunofunction ex vivo during isolation from CNS tissue. Isolating microglia via rotary shaking results in highly pure and immunofunctional cell cultures as assessed by fluorescent imaging, immunocytochemistry, and ELISA following microglia activation with the proinflammatory stimuli lipopolysaccharide (LPS) and Pam3CSK4 (Pam).

皮层小胶质细胞的分离与保存免疫表型和功能从新生儿鼠

Isolation of microglia from CNS tissue is a powerful investigative tool used to study microglial biology ex vivo. The present method details a procedure ...

免疫与感染

Preparation and Immunostaining of Myelinating Organotypic Cerebellar Slice Cultures

In the nervous system, myelin is a complex membrane structure generated by myelinating glial cells, which ensheathes axons and facilitates fast electrical conduction. Myelin alteration has been shown to occur in various neurological diseases, where it is associated with functional deficits. Here, we provide a detailed description of an ex vivo model consisting of mouse organotypic cerebellar slices, which can be maintained in culture for several weeks and further be labeled to visualize myelin.

Here we present a method to prepare organotypic slice cultures from mouse cerebellum and myelin sheath staining by immunohistochemistry suitable for investigating mechanisms of myelination and remyelination in the central nervous system.

髓鞘有机小脑片培养的制备及免疫染色

In the nervous system, myelin is a complex membrane structure generated by myelinating glial cells, which ensheathes axons and facilitates fast electrical ...

神经科学

Primary Culture of Neurons Isolated from Embryonic Mouse Cerebellum

The use of primary cell cultures has become one of the major tools to study the nervous system in vitro. The ultimate goal of using this simplified model system is to provide a controlled microenvironment and maintain the high survival rate and the natural features of dissociated neuronal and nonneuronal cells as much as possible under in vitro conditions. In this article, we demonstrate a method of isolating primary neurons from the developing mouse cerebellum, placing them in an in vitro environment, establishing their growth, and monitoring their viability and differentiation for several weeks. This method is applicable to embryonic neurons dissociated from cerebellum between embryonic days 12&ndash;18.

Conducting in vitro experiments to reflect in vivo conditions as adequately as possible is not an easy task. The use of primary cell cultures is an important step toward understanding cell biology in a whole organism. The provided protocol outlines how to successfully grow and culture embryonic mouse cerebellar neurons.

从胚胎小鼠小脑分离的神经元的主要培养

The use of primary cell cultures has become one of the major tools to study the nervous system in vitro. The ultimate goal of using this simplified model ...

Dissection and Isolation of Murine Glia from Multiple Central Nervous System Regions

The methods presented here demonstrate laboratory procedures for the dissection of four different regions of the central nervous system (CNS) from murine neonates for the isolation of glial subpopulations. The purpose of the procedure is to dissociate microglia, oligodendrocyte progenitor cells (OPCs), and astrocytes from cortical, cerebellar, brainstem, and spinal cord tissue to facilitate further in vitro analysis. The CNS region isolation procedures allow for the determination of regional heterogeneity among glia in multiple cell culture systems. Rapid CNS region isolation is performed, followed by the mechanical removal of meninges to prevent meningeal cell contamination of glia. This protocol combines gentle tissue dissociation and plating on a specified matrix designed to preserve cell integrity and adherence. Isolating mixed glia from multiple CNS regions provides a comprehensive analysis of potentially heterogenous glia while maximizing the use of individual experimental animals. Additionally, following dissociation of regional tissue, mixed glia are further divided into multiple cell types including microglia, OPCs, and astrocytes for use in either single cell type, cell culture plate inserts, or co-culture systems. Overall, the demonstrated techniques provide a comprehensive protocol of broad applicability for careful dissection of four individual CNS regions from murine neonates and includes methods for the isolation of three individual glia cell types to examine regional heterogeneity in any number of in vitro cell culture systems or assays.

Here we present a protocol for in vitro isolation of multiple glial cell populations from a mouse CNS. This method allows for the segregation of regional microglia, oligodendrocyte precursor cells, and astrocytes to study the phenotypes of each in a variety of culture systems.

从多个中枢神经系统区域解剖和分离小鼠神经胶质细胞

The methods presented here demonstrate laboratory procedures for the dissection of four different regions of the central nervous system (CNS) from murine ...

Cerebellar Regional Dissection for Molecular Analysis

Cerebellum plays an important role in several key functions including control of movement, balance, cognition, reward, and affect. Imaging studies indicate that distinct cerebellar regions contribute to these different functions. Molecular studies examining regional cerebellar differences are lagging as they are mostly done on whole cerebellar extracts thereby masking any distinctions across specific cerebellar regions. Here we describe a technique to reproducibly and quickly dissect four different cerebellar regions: the deep cerebellar nuclei (DCN), anterior and posterior vermal cerebellar cortex, and the cerebellar cortex of the hemispheres. Dissecting out these distinct regions allows for the exploration of molecular mechanisms that may underlie their unique contributions to balance, movement, affect and cognition. This technique may also be used to explore differences in pathological susceptibility of these specific regions across various mouse disease models.

Different cerebellar regions have been implicated to play a role in distinct behavioral outputs, yet the underlying molecular mechanisms remain unknown. This work describes a method to reproducibly and quickly dissect cerebellar cortex of the hemispheres, anterior and posterior regions of the vermis, and the deep cerebellar nuclei in order to probe for molecular differences by isolating RNA and testing for differences in gene expression.&#160;

分子分析的谷类区域解剖

Cerebellum plays an important role in several key functions including control of movement, balance, cognition, reward, and affect. Imaging studies ...

Efficient and Cost Effective Electroporation Method to Study Primary Cilium-Dependent Signaling Pathways in the Granule Cell Precursor

The primary cilium is a critical signaling organelle found on nearly every cell that transduces Hedgehog (Hh) signaling stimuli from the cell surface. In the granule cell precursor (GCP), the primary cilium acts as a pivotal signaling center that orchestrates precursor cell proliferation by modulating the Hh signaling pathway. The investigation of primary cilium-dependent Hh signaling machinery is facilitated by in vitro genetic manipulation of the pathway components to visualize their dynamic localization to the primary cilium. However, transfection of transgenes in the primary cultures of GCPs using the currently known electroporation methods is generally costly and often results in low cell viability and undesirable transfection efficiency. This paper introduces an efficient, cost-effective, and simple electroporation protocol that demonstrates a high transfection efficiency of ~80-90% and optimal cell viability. This is a simple, reproducible, and efficient genetic modification method that is applicable to the study of the primary cilium-dependent Hedgehog signaling pathway in primary GCP cultures.

Here, we present a reproducible in vitro electroporation protocol for genetic manipulation of primary cerebellar granule cell precursors (GCPs) that is cost-effective, efficient, and viable. Moreover, this protocol also demonstrates a straightforward method for the molecular study of primary cilium-dependent Hedgehog signaling pathways in primary GCP cells.

高效且具有成本效益的电穿孔方法研究颗粒细胞前体中的原代纤毛依赖性信号通路

The primary cilium is a critical signaling organelle found on nearly every cell that transduces Hedgehog (Hh) signaling stimuli from the cell surface. In ...

Isolation and Culture of Mouse Primary Cerebellar Granule Neurons

Source: Laaper, M., et al. <a href="https://app.jove.com/v/55871">Modeling Neuronal Death and Degeneration in Mouse Primary Cerebellar Granule Neurons.</a> J. Vis. Exp. (2017)
This video demonstrates the technique for isolating and culturing primary mouse cerebellar granule neurons. Cells are treated with an antimetabolite to eliminate proliferating glial cells and produce a pure culture of cerebellar granule neurons.

小鼠原代小脑颗粒神经元的分离和培养

All procedures involving sample collection have been performed in accordance with the institute&#39;s IRB guidelines.
1. Experimental Preparation
NOTE: ...

JoVE Encyclopedia of Experiments

Neuronal Culture Techniques

Primary Neuronal Culture

Isolation of Stem Cells from Postnatal Mouse Cerebellum and Differentiation into Neural Cells

Source: Edamakanti, C. R., et al. <a href="https://app.jove.com/v/60554">Purification of Prominin-1+ Stem Cells from Postnatal Mouse Cerebellum.</a> J. Vis. Exp. (2020).
This video demonstrates a method for isolating stem cells from the postnatal mouse cerebellum and differentiating them into neural cells. Stem cells from the isolated cell suspension are purified using antibody-coated magnetic beads and are cultured to form neurospheres. The neurospheres are then maintained in a differentiation medium to generate neural cells.

从出生后小鼠小脑中分离干细胞并分化为神经细胞

1. Preparation of Solutions

	Prepare tissue dissociation solution made from sterile phenol red-containing Dulbecco&#39;s phosphate-buffered saline (DPBS) ...