Este trabajo fue apoyado por el NIH subvención EB007278.

Hacer una BFC BFC son hechas por moldeo polidimetilsiloxano (PDMS) en un 4 &quot;oblea de silicio maestro. Hacer una oblea de maestro <ol><li> Comience por la deshidratación de la obleas de silicio durante 30 minutos a 130 ° C. </li><li> Lugar de la oblea en un spin-aplicador, vierta SU8-2050 (Microchem) sobre la oblea, rampa de 300 rpm / s de 3000-4000 rpm, y el giro de 30 s para obtener la altura característica de 40-30 mm, respectivamente. </li><li> Después de girar, el lugar de la oblea en una placa 65 ° C, inmediatamente rampa encima de la temperatura a 95 ° C durante 5-6 minutos, y luego la rampa de bajada a 65 ° C. </li><li> Exponer las obleas a una dosis UV de 200 mJ / cm 2 en un alineador de contacto usando una máscara de campo oscuro. </li><li> Coloque las galletas en una placa de 65 ° C, inmediatamente rampa encima de la temperatura a 95 ° C durante 4-5 minutos, y luego la rampa de bajada a 65 ° C. </li><li> Desarrollar las obleas en acetato de PM y el isopropanol. </li><li> Silanizar las obleas durante 30 minutos utilizando hexametildisiloxano (Shin-Etsu MicroSi), o (tridecafluoro-1 ,1,2,2-tetrahydrooctyl)-1-triclorosilano (t2492-KG, Especialidades UCT, Bristol, PA), para evitar que a partir de PDMS la adhesión a la oblea de Si señor. </li></ol> Chips de moldeo <ol><li> Mezcla de PDMS (Dow Corning, Sylgard 184) en una proporción de 10:1, se vierte sobre la oblea maestro Si (~ 15 g por oblea), y se deja secar toda la noche a temperatura ambiente. </li><li> Pelar el curado PDMS de la oblea de silicio y corte cada chip con una cuchilla de afeitar. </li><li> Vínculo cada chip a un portaobjetos de microscopio 1x1 cm de corte para un fácil manejo. </li></ol> Preparación de la BFC para el uso <ol><li> Remoje la noche a la mañana BFC en PBS con el fin de prevenir la absorción de los medios de comunicación en el PDMS durante el experimento. </li><li> Seque los chips con un Kimwipe (KIMTECH Science). </li><li> Ponga una gota de 7,5% albúmina de suero bovino (BSA) (~ 200 ml) en la superficie de estampado de la BFC, y luego raspar con una punta de pipeta para dispersar a la BSA y eliminar las burbujas de los pozos. </li><li> Salir de la BSA sobre la superficie del BFC, por lo menos 0,5 horas. Lo ideal sería que agitan la BSA cada 15 minutos para evitar la formación de costras. </li><li> Para adaptarse a la BFC dentro de un plato de 35x10 mm TCPS (Falcon, 35-3.001), corte los bordes de una manera medio plato TCPS abajo para que el ¾ &quot;clips de la carpeta (Office Depot) caben sobre el borde plato para sujetar el chip y plato juntos. </li><li> Cortar una junta de separador (en forma de marco con el borde exterior de 20&#39;20 mm, borde interno 15&#39;15 mm y 250-500 mm de espesor) de una hoja de PDMS (Productos Especiales de silicona) o cualquier otra de silicona, y aplicarlo a la antena con unas pinzas. </li></ol> Células con un patrón de BFC <ol><li> Esterilizar el plato, la junta, chip recubierto de BSA, y dos clips de la carpeta a la luz ultravioleta durante 1 minuto. </li><li> Aspirar a la BSA y enjuague el BFC dos veces con 200 ml de PBS. </li><li> Añadir la gelatina al plato TCPS corte si es necesario para su línea celular. De lo contrario, humedecer la superficie TCPS con algunos medios de comunicación antes de aplicarlo en el chip (ver más abajo). Esto evita que se formen burbujas en la cámara. </li><li> Después de aspirar el PBS, una pipeta 200 ml de solución de células (~ 1 × 10 6 células / ml) sobre la superficie del BFC. Vamos a colocar las células durante 5-10 minutos. </li><li> La inclinación de la BFC a una esquina en un ángulo de ~ 15 ° y lentamente la pipeta la solución de células con una pipeta de 200 ml. Luego, coloque el piso BFC y añadir 100 ml de PBS o medio vacío en la esquina opuesta BFC. Enjuague el BFC de esta manera un adicional de 2.4 veces, según sea necesario, para eliminar todas las células de las regiones entre bien. </li><li> Pipeta 200 ml de los medios de comunicación en la superficie del chip. Aspirar la gelatina / media de los TCPS. Luego invierta el plato previamente mojado y empuje lentamente hasta la BFC, en el plato. </li><li> Aplicar una carpeta de clip de cada uno, a dos lados de la cámara para sellarlo. Quitar las púas de metal de los clips de carpeta para que el plato se mantiene el nivel cuando se volcó. </li><li> Por último, rápidamente la vuelta al la instalación a través, evitando cualquier movimiento innecesario, y lo coloca en la incubadora. </li></ol>

<ol>
	<li>Rosenthal, A., Macdonald, A., Voldman, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell+Patterning+Chip+for+Controlling+the+Stem+Cell+Microenvironment.">Cell Patterning Chip for Controlling the Stem Cell Microenvironment.</a> Biomaterials. 28, 3208-3216 (2007).</li></ol>

Aunque el protocolo y el video se muestra aquí se describe el uso de la BFC un patrón de células madre embrionarias de ratón (mESCs), BFC se puede utilizar para patrón de prácticamente cualquier tipo de célula de interés. El único cambio que uno puede necesitar hacer es cambiar el tamaño de las cúpulas. En nuestra experiencia, para utilizar el BFC a las células de un patrón único de otro tipo de células, un diámetro de micropocillos y altura igual a 10 micrones mayor que el diámetro de la celda libre que funciona...

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		<th>Material Name</th>
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		<th>Catalogue Number</th>
		<th>Comment</th>
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<tr>
<td>PDMS</td>
<td>&nbsp;</td>
<td>Dw Corning</td>
<td>Sylgard 184</td>
<td>&nbsp;</td>
<tr>
<td>Silicon master mould</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Tissue culture dishes</td>
<td>&nbsp;</td>
<td>Falcon</td>
<td>35-3001</td>
<td>35 mm</td>
</tr>
<tr>
<td>3/4" binder clips</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>PBS</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>hexamethyldisiloxane</td>
<td>&nbsp;</td>
<td>Shin-Etsu MicroSi</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
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<td>New Item</td>
<td>&nbsp;</td>
<td>Microchem</td>
<td>SU8-2050</td>
<td>&nbsp;</td>
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<td>PM acetate</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Isopropanol</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>BSA</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
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patterning of embryonic stem cells using the bio flip chip

Interacciones célula-célula que consiste en la señalización difusibles y contacto célula-célula (juxtacrine señalización) son importantes en numerosos procesos biológicos como el crecimiento del tumor, la diferenciación de células madre y células madre de la auto-renovación. Una serie de métodos existen actualmente para modular la señalización celular in vitro. Uno de los métodos de la modulación de la cantidad total de la señalización difusibles es variar la densidad de siembra de células durante el cultivo. Debido a la naturaleza aleatoria de la siembra de células, esto se traduce en una variación considerable en la actual célula-célula y la cantidad de espacio de contacto entre células, y no puede prescribir el medio ambiente local. Un enfoque más específico para la modulación de la señalización celular es el uso de inhibidores moleculares o métodos genéticos para derribar a proteínas específicas de señalización, pero ambos de estos métodos son más adecuados para la manipulación de un pequeño número de moléculas. En este caso, se demuestra un nuevo enfoque para la modulación de las células de señalización celular que modula el ambiente local de un grupo de células mediante la colocación de diferentes números de células a los lugares deseados sobre un sustrato. Este método proporciona una forma complementaria para controlar la señalización local difusible y juxtacrine entre las células. Nuestro método se basa en el chip de Bio Flip (BFC), un chip de silicona que contienen cientos microfabricated a miles de micropocillos cada tamaño a suscribir una sola celda o un pequeño número de células. Cargamos el chip con las células simplemente pipeteando ellos en la matriz de los pozos y el lavado de las células sin carga fuera de la matriz. El chip es luego volcó sobre un sustrato, por el cual las células se caen de los pozos y en el sustrato, el mantenimiento de su patrón. Después las células se han unido, el chip se puede quitar (o la izquierda en). Este enfoque de modelado celular es único, ya que: 1) no altera la química del sustrato, lo que permite a las células proliferan y migran, 2) permite a los patrones en cualquier soporte, incluidos los de poliestireno de cultivo de tejidos, vidrio, matrigel, y incluso alimentación capas de células, y 3) es compatible con la impresión de microcontacto tradicionales, permitiendo la creación de islas de la matriz extracelular con células coloca dentro de las islas. En este vídeo, se demuestra el patrón de las células madre embrionarias de ratón en poliestireno de cultivo de tejidos con la BFC.

Watch this Scientific Journal Video about Patterning of Embryonic Stem Cells Using the Bio Flip Chip at JoVE.com

Patterning of Embryonic Stem Cells Using the Bio Flip Chip

Demostramos un método sencillo para colocar las células en los puntos deseados en un substrato. Este método de los patrones de las células al lanzar un chip de silicona que contiene micropocillos llena de células en el sustrato. Este método ofrece una nueva forma de modular la señalización difusibles y juxtacrine entre las células.

Los patrones de las células madre embrionarias usando el Flip Chip Bio

Cell-cell interactions consisting of diffusible signaling and cell-cell contact (juxtacrine signaling) are important in numerous biological processes such ...

patterning-of-embryonic-stem-cells-using-the-bio-flip-chip

Research

JoVE Journal

Biology

9.4K vistas.  MIT - Massachusetts Institute of Technology. Demostramos un método sencillo para colocar las células en los puntos deseados en un substrato. Este método de los patrones de las células al lanzar un chip de silicona que contiene micropocillos llena de células en el sustrato. Este método ofrece una nueva forma de modular la señalización difusibles y juxtacrine entre las células.

Video: Patterning of Embryonic Stem Cells Using the Bio Flip Chip

Propagation of Human Embryonic Stem (ES) Cells

Propagation of Human Embryonic Stem ES Cells

Yeah, so I'm Lauren Deon. I'm in charge of the Human Amgen stem cell co facility at MGH. Yeah, so I'm Lauren, I'm in charge of the Human AM stem cell co facility at MGH.So I've been working with human Amgen stem cells for years now since I was a postdoc in George De Lab before, from four, four years. And so right now we have two cell lines, H one and H nine, which are both from Y cell. And so the way we are expanding or maintaining these cells in culture is by, at least the way we are splitting these cells is by using collagenous.So what I usually do is for the first few passage after throwing the cells, I use a technical, the picking colonies. So in this case, I can check under the microscope which colonies are good and differentiate and nice shape, and I will collect only these colonies and I will cut them in few pieces and put them back in, in a fresh flat. And the, the colonies that look kind of differentiate it are just left them alone.The other way to pass the cells is by using trypsin. So I don't use this technique here. I think collagenase is a little bit safer because human being stem cells don't like to be individualized or they don't like to be alone.So if you do trypsin, if you use trypsin for a little bit too long, if you use it for like three to four minutes, which is the way we use strips in for other type of cells, human stem cells will be individualized and most of them will die or just differentiate or will not attach to the myth. So the idea is you have to keep the cells as clumps. You don't want them to be alone in the in, in the flask.So using collagenase, the advantage is to keep cells as plants. Using chipsy, you just select for cells that will survive this very, very harsh process of g opsonization. So these cells are usually very highly proliferative.So you get, you can adapt your cells to this tripsin process. The problem is that you have cells that they behave differently, they go much faster. And one risk is that you can get some commercial abnormality to, to this, to this very high division process.So you say that ization can cause selection of cells in culture for most proliferative cells.Right. More ative and probably also higher survival rates than other cells. So after proliferation, basically culture after proliferation in tripsin, after several passages become a different culture, these different properties.So it's, it's hard to figure out if they are exactly identical. If you do the test, the usual test, let's say you check markers, regular markers of end differentiated cells like OC four, nano one 60 SC four C, C3, all this molecule might still be expressed. If you do TER formation, this cells might still differentiate and be formed like al like normal cells.If you look at the karyotype, you might have some abnormality, but you might still have a normal karyotype. The, the, the selective advantage might be due to just a mutation that you cannot see by doing a karyotype. So the fact that they grow much faster is by itself I think a little bit worser.And if we want to use these cells for transplantation at some point, I would definitely not use this type of cells.Okay. So would you like to tell me what are the main technical problems with using this technique for, for passages of cells? So using S, yes.It's just a little bit longer and also it's, since the cells are not going as fast, it's just much longer to get a high number of cells to do some experiment. So it's just less convenient I would say. Would you like to tell me about few words about the goals of your experiment?How do you use CS cells? Okay, so my main project here is to get endothelial cells from human AB stem cells. And the idea is to be able to make blood.And we are working a lot with tissue bioengineering labs and one of their goal is to make tissue in vitro. But one of the limitation to this is the lack of blood vessels going through this tissues. So if we can bring endothelial cells or put endothelial cells in this tissue, we would have better growth of this tissue.So the idea few sources of cells are known for endothelial cells like cold blood or pressure blood, but it would be interesting to have another source of cells like human years. And the advantage is since human years can be expanded and most infinitely, you could modify these cells, make some inducible system and be able to use it. Then in your final product, your material cells, in this case.

Propagación de la madre embrionarias humanas (ES) Células

Investigación

Biología

Derivation of Hematopoietic Stem Cells from Murine Embryonic Stem Cells

A stem cell is defined as a cell with the capacity to both self-renew and generate multiple differentiated progeny.  Embryonic stem cells (ESC) are derived from the blastocyst of the early embryo and are pluripotent in differentiative ability.  Their vast differentiative potential has made them the focus of much research centered on deducing how to coax them to generate clinically useful cell types.  The successful derivation of hematopoietic stem cells (HSC) from mouse ESC has recently been accomplished and can be visualized in this video protocol.  HSC, arguably the most clinically exploited cell population, are used to treat a myriad of hematopoietic malignancies and disorders.  However, many patients that might benefit from HSC therapy lack access to suitable donors.  ESC could provide an alternative source of HSC for these patients.  The following protocol establishes a baseline from which ESC-HSC can be studied and inform efforts to isolate HSC from human ESC.  In this protocol, ESC are differentiated as embryoid bodies (EBs) for 6 days in commercially available serum pre-screened for optimal hematopoietic differentiation.  EBs are then dissociated and infected with retroviral HoxB4.  Infected EB-derived cells are plated on OP9 stroma, a bone marrow stromal cell line derived from the calvaria of  M-CSF-/- mice, and co-cultured in the presence of hematopoiesis promoting cytokines for ten days.  During this co-culture, the infected cells expand greatly, resulting in the generation a heterogeneous pool of 100s of millions of cells.  These cells can then be used to rescue and reconstitute lethally irradiated mice.

This protocol details the derivation of transplantable hematopoietic stem cells from mouse embryonic stem cells (ESC) and their subsequent injection into lethally irradiated recipient mice. Briefly, ESC are differentiated as embryoid bodies, which are then infected with retroviral HoxB4 and co-cultured with OP9 stromal cells and hematopoietic cytokines.

Derivación de células madre hematopoyéticas de células madre embrionarias murinas

A stem cell is defined as a cell with the capacity to both self-renew and generate multiple differentiated progeny.  Embryonic stem cells (ESC) are ...

CD4+ T-Lymphocyte Capture Using a Disposable Microfluidic Chip for HIV

CD4 + T-linfocitos capturar usando un chip de microfluidos desechables para el VIH

RNA Extraction from Neuroprecursor Cells Using the Bio-Rad Total RNA Kit

La extracción de RNA a partir de células Neuroprecursor utilizando el kit de Bio-Rad ARN total

Derivation of Human Embryonic Stem Cells by Immunosurgery

The ability of human embryonic stem cells to self-renew and differentiate into all cell types of 
the body suggests that they hold great promise for both medical applications and as a research 
tool for addressing fundamental questions in development and disease. Here, we provide a 
concise, step-by-step protocol for the derivation of human embryonic stem cells from embryos 
by immunosurgical isolation of the inner cell mass. 


The ability of human embryonic stem cells to self-renew and differentiate into all cell types of 
the body suggests that they hold great promise for both medical applications and as a research 
tool for addressing fundamental questions in development and disease. Here, we provide a 
concise, step-by-step protocol for the derivation of human embryonic stem cells from embryos 
by immunosurgical isolation of the inner cell mass.

Derivación de células estaminales embrionarias humanas por Immunosurgery

The ability of human embryonic stem cells to self-renew and differentiate into all cell types of 
the body suggests that they hold great promise for both ...

Chromatin Immunoprecipitation from Human Embryonic Stem Cells

The functional and structural complexity of the myriad of cells in metazoan organisms arises from a small number of stem cells. Stem cells are characterized by two fundamental properties: self-renewal and multipotency that allows a stem cell to differentiate into virtually any cell type 1. The progression stem cell to differentiated cell is characterized by loss of multipotency, structural and morphological changes and the hierarchic activity of transcription factors and signaling molecules, whose activities establish and maintain cell-type specific gene expression patterns. At the molecular level, cell differentiation involves dynamic changes of the structure and composition of chromatin and the detection of those dynamic changes can provide valuable insights into the functional features of stem cells and the cell differentiation process 2,3. Chromatin is a highly compacted DNA-protein complex that forms when cells package chromosomal DNA with proteins, mainly histones 4. Stemcellness and cell differentiation has been correlated with the presence of specific arrays of regulatory proteins such as epigenetic factors, histone variants, and transcription factors 2,3,5.
 Chromatin immunoprecipitation (ChIP) provides a valuable method to monitor the presence of RNA, proteins, and protein modifications in chromatin 6,7. The comparison of chromatin from different cell types can elucidate dynamic changes in protein-chromatin associations that occur during cell differentiation.
Chromatin immunoprecipitation involves the purification of in vivo cross-linked chromatin. The isolated chromatin is reduced to smaller fragments by enzymatic digestion or mechanical force. Chromatin fragments are precipitated using specific antibodies to target proteins or protein and DNA modifications. The precipitated DNA or RNA is purified and used as a template for PCR or DNA microarray based assays. Prerequisites for a successful ChIP are high quality antibodies to the desired antigen and the availability of chromatin from control cells that do not express the target molecule. ChIP can correlate the presence of proteins, protein and RNA modifications, and RNA with specific target DNA, and depending on the choice of outread tool, detects the association of target molecules at specific target genes or in the context of an entire genome. The comparison of the distribution of proteins in the chromatin of differentiating cells can elucidate the dynamic changes of chromatin composition that coincide with the progression of cells along a cell lineage.

The differentiation of ESC coincides with cell-type specific changes in the structure and composition of chromatin. The detection of those changes provides valuable insights into the mechanisms that define stemcellness and cell differentiation. Chromatin immunoprecipitation (ChIP) represents a valuable method to dissect the molecular mechanisms underlying stem cell differentiation.

Inmunoprecipitación de la cromatina de células estaminales embrionarias humanas

The functional and structural complexity of the myriad of cells in metazoan organisms arises from a small number of stem cells. Stem cells are ...

In vitro Differentiation of Mouse Embryonic Stem (mES) Cells Using the Hanging Drop Method

Stem cells have the remarkable potential to develop into many different cell types. When a stem cell divides, each new cell has the potential to either remain a stem cell or become another type of cell with a more specialized function, This promising of science is leading scientists to investigate the possibility of cell-based therapies to treat disease.

When culture in suspension without antidifferentiation factors, embryonic stem cells spontaneously differentiate and form three-dimensional multicellular aggregates. These cell aggregates are called embryoid bodies(EB). Hanging drop culture is a widely used EB formation induction method. The rounded bottom of hanging drop allows the aggregation of ES cells which can provide mES cells a good environment for forming EBs. The number of ES cells aggregatied in a hanging drop can be controlled by varying the number of cells in the initial cell suspension to be hung as a drop from the lid of Petri dish. Using this method we can reproducibly form homogeneous EBs from a predetermined number of ES cells. 

In vitro Differentiation of Mouse Embryonic Stem mES Cells Using the Hanging Drop Method

This video demonstrates how to conduct in vitro differentiation of mouse embryonic stem cells to embryoid bodies using the hanging drop method.

En la diferenciación in vitro de la madre embrionarias de ratón (MES), las células con el método de gota

Stem cells have the remarkable potential to develop into many different cell types. When a stem cell divides, each new cell has the potential to either ...

Culture and Maintenance of Human Embryonic Stem Cells  

Human embryonic stem (hES) cells must be monitored and cared for in order to maintain healthy, undifferentiated cultures. At minimum, the cultures must be  fed  every day by performing a complete medium change to replenish lost nutrients and to keep the cultures free of unwanted differentiation factors. Although a small amount of differentiation is normal and expected in stem cell cultures, the culture should be routinely cleaned up by manually removing, or "picking" differentiated areas. Identifying and removing excess differentiation from hES cell cultures are essential techniques in the maintenance of a healthy population of cells.

Culture and Maintenance of Human Embryonic Stem Cells

This protocol demonstrates how to maintain healthy, undifferentiated human embryonic stem (ES) cells.

La cultura y el mantenimiento de células estaminales embrionarias humanas

Human embryonic stem (hES) cells must be monitored and cared for in order to maintain healthy, undifferentiated cultures. At minimum, the cultures must be ...

Freezing and Thawing Human Embryonic Stem Cells

Since James Thomson et al developed a technique in 1998 to isolate and grow hES in culture, freezing cells for later use and thawing and expanding cells from a frozen stock have become important procedures performed in routine hES cell culture. Since hES cells are very sensitive to the stresses of freezing and thawing, special care must taken. Here we demonstrate the proper technique for rapidly thawing hES cells from liquid nitrogen stocks, plating them on mouse embryonic feeder cells, and slowly freezing them for long-term storage.

Since James Thomson et al developed a technique in 1998 to isolate and grow hES in culture, freezing cells for later use and thawing and expanding cells from a frozen stock have become important procedures performed in routine hES cell culture. Since hES cells are very sensitive to the stresses of freezing and thawing, special care must taken. Here we demonstrate the proper technique for rapidly thawing hES cells from liquid nitrogen stocks, plating them on mouse embryonic feeder cells, and slowly freezing them for long-term storage.

Congelación y descongelación células estaminales embrionarias humanas

Since James Thomson et al developed a technique in 1998 to isolate and grow hES in culture, freezing cells for later use and thawing and expanding cells ...

Isolation and Derivation of Mouse Embryonic Germinal Cells

The ability of embryonic germinal cells (EG) to differentiate into primordial germinal cells (PGCs) and later into gametes during early developmental stages is a perfect model to address our hypothesis about cancer and infertility. This protocol shows how to isolate primordial germinal cells from developing gonads in 10.5-11.5 days post coitum (dpc) mouse embryos.  Developing gonadal ridges from mouse embryos (C57BL6J) were dissociated by mechanical disruption with collagenase, then plated in a mouse embryo fibroblast feeder layer (MEF-CF1) that was previously mitotically inactivated with mitomycin C in the presence of knockout media and supplemented with Leukemia Inhibitor Factor (LIF), basic Fibroblast Growth Factor (bFGF), and Stem Cell Factor (SCF).  Using these optimized methods for PCG identification, isolation, and establishment of culture conditions permits long term cultures of EG cells for more than 40 days.  The embryonic germinal cell lines showed embryonic phenotype and expression of common used markers of the pluripotent state.  Isolation and derivation of germinal cells in culture provide a tool to understand their development in vitro and offer the opportunity to monitor cumulative damage at genetic and epigenetic levels after exposure to oxidative stress.

The ability of embryonic germinal cells to differentiate into primordial germinal cells during early development stages is a perfect model to address our hypothesis about cancer and infertility. This protocol shows how to isolate primordial germinal cells from developing gonads in 10.5-11.5 days post coitum mouse embryos.

El aislamiento y la derivación de células embrionarias de ratón Germinal

The ability of embryonic germinal cells (EG) to differentiate into primordial germinal cells (PGCs) and later into gametes during early developmental ...