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At least 253 records · Page 14Linked to original sources
PubMed2006

Meta-analysis of differentiating mouse embryonic stem cell gene expression kinetics reveals early change of a small gene set.

Stem cell differentiation involves critical changes in gene expression. Identification of these should provide endpoints useful for optimizing stem cell propagation as well as potential clues about mechanisms governing stem cell maintenance. Here we describe the results of a new meta-analysis methodology applied to multiple gene expression datasets from three mouse embryonic stem cell (ESC) lines obtained at specific time points during the course of their differentiation into various lineages. We developed methods to identify genes with expression changes that correlated with the altered frequency of functionally defined, undifferentiated ESC in culture. In each dataset, we computed a novel statistical confidence measure for every gene which captured the certainty that a particular gene exhibited an expression pattern of interest within that dataset. This permitted a joint analysis of the datasets, despite the different experimental designs. Using a ranking scheme that favored genes exhibiting patterns of interest, we focused on the top 88 genes whose expression was consistently changed when ESC were induced to differentiate. Seven of these (103728_at, 8430410A17Rik, Klf2, Nr0b1, Sox2, Tcl1, and Zfp42) showed a rapid decrease in expression concurrent with a decrease in frequency of undifferentiated cells and remained predictive when evaluated in additional maintenance and differentiating protocols. Through a novel meta-analysis, this study identifies a small set of genes whose expression is useful for identifying changes in stem cell frequencies in cultures of mouse ESC. The methods and findings have broader applicability to understanding the regulation of self-renewal of other stem cell types.

Adaptation, Physiological↗
PubMed2004

Pathways regulating lens induction in the mouse.

For more than a century, the lens has provided a relatively simple structure in which to study developmental mechanisms. Lens induction, where adjacent tissues signal the cell fate changes that result in lens formation, have been of particular interest. Embryological manipulations advancing our understanding have included the Spemann optic rudiment ablation experiments, optic vesicle transplantations as well as more contemporary work employing lineage tracers. All this has revealed that lens induction signaling is a multi-stage process involving multiple tissue interactions. More recently, molecular genetic techniques have been applied to an analysis of lens induction. This has led to the identification of signaling pathways required for lens induction and early lens development. These include the bone morphogenetic protein (Bmp) signaling pathways where Bmp4 and Bmp7 have been implicated. Though no fibroblast growth factor (Fgf) ligand has been implicated at present, the Fgf signaling pathway clearly has an important role. A series of transcription factors involved in early lens development have also been identified. These include Pax6, the Meis transcription factors, Six3, Mab21l1, FoxE3, Prox1 and Sox2. Importantly, analysis has indicated how these elements of the lens induction pathway are related and has defined genetic models to describe the process. It is a future challenge to test existing genetic models and to extend them to incorporate the tissue interactions mediated by the molecules involved. Given the complexity of this and many other developmental processes, a second century of analysis will be welcome.

Animals↗
PubMed2006

Mechanisms controlling embryonic stem cell self-renewal and differentiation.

Embryonic stem (ES) cells are pluripotent cells with indefinite replication potential and ability to differentiate into all types of cells. An understanding of the regulatory mechanisms responsible for pluripotency in ES cells is critical for realizing their potential in regenerative medicine and science. Cross-species studies on ES cells have identified pathways and networks that are either fundamental to or species-specific for self-renewal and differentiation. Although pluripotency as an essential function in multicellular organisms is conserved through evolution, mechanisms primed for differentiation contribute substantially to the differences among stem cells derived from different tissues or species. Transcriptome mapping analysis has determined the chromosomal domains of gene coexpression patterns specific to the ES state and demonstrated that regulation of ES cell development is operative at both the local chromosomal domain level and global level. Combinatorial signals from multiple pathways regulate the expression of key intrinsic factors critical for ES cell fate determination. The regulatory core formed by Oct4, Sox2, and Nanog, in particular, activates genes critical for self-renewal and represses genes initiating differentiation, controlling ES cell pluripotency. Here, we review recent findings on mechanisms controlling ES cell development. By integrating data from different sources, we present a global picture of how ES cells reach the decision of self-renewal or differentiation.

Animals↗
PubMed2005

Plasticity of cultured mesenchymal stem cells: switch from nestin-positive to excitable neuron-like phenotype.

Bone marrow mesenchymal stem cells (MSCs) can differentiate into several types of mesenchymal cells, including osteocytes, chondrocytes, and adipocytes, but, under appropriate experimental conditions, can also differentiate into nonmesenchymal cells--for instance, neural cells. These observations have raised interest in the possible use of MSCs in cell therapy strategies for various neurological disorders. In the study reported here, we addressed the question of in vitro differentiation of MSCs into functional neurons. First, we demonstrate that when they are co-cultured with cerebellar granule neurons, adult MSCs can express neuronal markers. Two factors are needed for the emergence of neuronal differentiation of the MSCs: the first one is nestin expression by MSCs (nestin is a marker for the responsive character of MSCs to extrinsic signals), and the second one is a direct cell-cell interaction between neural cells and MSCs that allows the integration of these extrinsic signals. Three different approaches suggest that neural phenotypes arise from MSCs by a differentiation rather than a cell fusion process, although this last phenomenon can also coexist. The expression of several genes--including sox, pax, notch, delta, frizzled, and erbB--was analyzed by quantitative reverse transcription polymerase chain reaction (RT-PCR) in order to further characterize the nestin-positive phenotype compared to the nestin-negative one. An overexpression of sox2, sox10, pax6, fzd, erbB2, and erbB4 is found in nestin-positive MSCs. Finally, electrophysiological analyses demonstrate that MSC-derived neuron-like cells can fire single-action potentials and respond to several neurotransmitters such as GABA, glycine, and glutamate. We conclude that nestin-positive MSCs can differentiate in vitro into excitable neuron-like cells.

Action Potentials↗
PubMed2005

Long-term proliferation of human embryonic stem cell-derived neuroepithelial cells using defined adherent culture conditions.

Research on the cell fate determination of embryonic stem cells is of enormous interest given the therapeutic potential in regenerative cell therapy. Human embryonic stem cells (hESCs) have the ability to renew themselves and differentiate into all three germ layers. The main focus of this study was to examine factors affecting derivation and further proliferation of multipotent neuroepithelial (NEP) cells from hESCs. hESCs cultured in serum-deprived defined medium developed distinct tube structures and could be isolated either by dissociation or adherently. Dissociated cells survived to form colonies of cells characterized as NEP when conditioned medium from human hepatocellular carcinoma HepG2 cell line (MEDII) was added. However, cells isolated adherently developed an enriched population of NEP cells independent of MEDII medium. Further characterization suggested that they were NEP cells because they had a similar phenotype profile to in vivo NEP cells and expression SOX1, SOX2, and SOX3 genes. They were positive for Nestin, a neural intermediate filament protein, and Musashi-1, a neural RNA-binding protein, but few cells expressed further differentiation markers, such as PSNCAM, A2B5, MAPII, GFAP, or O4, or other lineage markers, such as muscle actin, alpha fetoprotein, or the pluripotent marker Oct4. Further differentiation of these putative NEP cells gave rise to a mixed population of progenitors that included A2B5-positive and PSNCAM-positive cells and postmitotic neurons and astrocytes. To proliferate and culture these derived NEP cells, ideal conditions were obtained using neurobasal medium supplemented with B27 and basic fibroblast growth factor in 5% oxygen. NEP cells were continuously propagated for longer than 6 months without losing their multipotent cell characteristics and maintained a stable chromosome number.

Animals↗
PubMed2005

Correlation of murine embryonic stem cell gene expression profiles with functional measures of pluripotency.

Global gene expression profiling was performed on murine embryonic stem cells (ESCs) induced to differentiate by removal of leukemia inhibitory factor (LIF) to identify genes whose change in expression correlates with loss of pluripotency. To identify appropriate time points for the gene expression analysis, the dynamics of loss of pluripotency were investigated using three functional assays: chimeric mouse formation, embryoid body generation, and colony-forming ability. A rapid loss of pluripotency was detected within 24 hours, with very low residual activity in all assays by 72 hours. Gene expression profiles of undifferentiated ESCs and ESCs cultured for 18 and 72 hours in the absence of LIF were determined using the Affymetrix GeneChip U74v2. In total, 473 genes were identified as significantly differentially expressed, with approximately one third having unknown biological function. Among the 275 genes whose expression decreased with ESC differentiation were several factors previously identified as important for, or markers of, ESC pluripotency, including Stat3, Rex1, Sox2, Gbx2, and Bmp4. A significant number of the decreased genes also overlap with previously published mouse and human ESC data. Furthermore, several membrane proteins were among the 48 decreased genes correlating most closely with the functional assays, including the stem cell factor receptor c-Kit. Through identification of genes whose expression closely follows functional properties of ESCs during early differentiation, this study lays the foundation for further elucidating the molecular mechanisms regulating the maintenance of ESC pluripotency and facilitates the identification of more reliable molecular markers of the undifferentiated state.

Animals↗
PubMed2005

Expression of neurodevelopmental markers by cultured porcine neural precursor cells.

Despite the increasing importance of the pig as a large animal model, little is known about porcine neural precursor cells. To evaluate the markers expressed by these cells, brains were dissected from 60-day fetuses, enzymatically dissociated, and grown in the presence of epidermal growth factor, basic fibroblast growth factor, and platelet-derived growth factor. Porcine neural precursors could be grown as suspended spheres or adherent monolayers, depending on culture conditions. Expanded populations were banked or harvested for analysis using reverse transcription-polymerase chain reaction (RT-PCR), immunocytochemistry, microarrays, and flow cytometry, and results compared with data from analogous human forebrain progenitor cells. Cultured porcine neural precursors widely expressed neural cell adhesion molecule (NCAM), polysialic acid (PSA)-NCAM, vimentin, Ki-67, and Sox2. Minority subpopulations of cells expressed doublecortin, beta-III tubulin, synapsin I, glial fibrillary acidic protein (GFAP), and aquaporin 4 (AQP4) consistent with increased lineage restriction. A human microarray detected porcine transcripts for nogoA (RTN4) and stromal cell-derived factor 1 (SDF1), possibly cyclin D2 and Pbx1, but not CD133, Ki-67, nestin, or nucleostemin. Subsequent RT-PCR showed pig forebrain precursors to be positive for cyclin D2, nucleostemin, nogoA, Pbx1, vimentin, and a faint band for SDF1, whereas no signal was detected for CD133, fatty acid binding protein 7 (FABP7), or Ki-67. Human forebrain progenitor cells were positive for all the genes mentioned. This study shows that porcine neural precursors share many characteristics with their human counterparts and, thus, may be useful in porcine cell transplantation studies potentially leading to the application of this strategy in the setting of nervous system disease and injury.

Animals↗
PubMed2006

Transplanted embryonic stem cells successfully survive, proliferate, and migrate to damaged regions of the mouse brain.

An understanding of feasibility of implanting embryonic stem cells (ESCs), their behavior of migration in response to lesions induced in brain tissues, and the mechanism of their in vivo differentiation into neighboring neural cells is essential for developing and refining ESC transplantation strategies for repairing damages in the nervous system, as well as for understanding the molecular mechanism underlying neurogenesis. We hypothesized that damaged neural tissues offer a niche to which injected ESCs can migrate and differentiate into the neural cells. We inflicted damage in the murine (C57BL/6) brain by injecting phosphate-buffered saline into the left frontal and right caudal regions and confirmed neural damage by histochemistry. Enhanced yellow fluorescent protein-expressing ESCs were injected into the nondamaged left caudal portion of the brain. Using immunohistochemistry and fluorescent microscopy, we observed migration of ESCs from the injection site (left caudal) to the damaged site (right caudal and left frontal). Survival of the injected ESCs was confirmed by the real-time polymerase chain reaction analysis of stemness genes such as Oct4, Sox2, and FGF4. The portions of the damaged neural tissues containing ESCs demonstrated a fourfold increase in expression of these genes after 1 week of injection in comparison with the noninjected ESC murine brain, suggesting proliferation. An increased level of platelet-derived growth factor receptor demonstrated that ESCs responded to damaged neural tissues, migrated to the damaged site of the brain, and proliferated. These results demonstrate that undifferentiated ESCs migrate to the damaged regions of brain tissue, engraft, and proliferate. Thus, damaged brain tissue provides a niche that attracts ESCs to migrate and proliferate.

Animals↗
PubMed2006

Integrins are markers of human neural stem cells.

The identification of markers for the isolation of human neural stem cells (hNSCs) is essential for studies of their biology and therapeutic applications. This study investigated expression of the integrin receptor family by hNSCs as potential markers. Selection of alpha6(hi) or beta1(hi) cells by fluorescence-activated cell sorting led to an enrichment of human neural precursors, as shown by both neurosphere forming assays and increased expression of prominin-1, sox2, sox3, nestin, bmi1, and musashi1 in the beta1(hi) population. Cells expressing high levels of beta1 integrin also expressed prominin-1 (CD133), a marker previously used to isolate hNSCs, and selection using integrin beta1(hi) cells or prominin-1(hi) cells was found to be equally effective at enriching for hNSCs from neurospheres. Therefore, integrin subunits alpha6 and beta1 are highly expressed by human neural precursors and represent convenient markers for their prospective isolation.

AC133 Antigen↗
PubMed2006

Generation and characterization of rabbit embryonic stem cells.

We described the derivation of four stable pluripotent rabbit embryonic stem cell (ESC) lines, one (RF) from blastocysts fertilized in vivo and cultured in vitro and three (RP01, RP02, and RP03) from parthenogenetic blastocysts. These ESC lines have been cultivated for extended periods (RF >1 year, RP01 >8 months, RP02 >8 months, and RP03 >6 months) in vitro while maintaining expression of pluripotent ESC markers and a normal XY or XX karyotype. The ESCs from all lines expressed alkaline phosphatase, transcription factor Oct-4, stage-specific embryonic antigens (SSEA-1, SSEA-3, and SSEA-4), and the tumor-related antigens (TRA-1-60 and TRA-1-81). Similar to human and mouse ESCs, rabbit ESCs expressed pluripotency (Oct-4, Nanog, SOX2, and UTF-1) and signaling pathway genes (fibroblast growth factor, WNT, and transforming growth factor pathway). Morphologically, rabbit ESCs resembled primate ESCs, whereas their proliferation characteristics were more like those seen in mouse ESCs. Rabbit ESCs were induced to differentiate into many cell types in vitro and formed teratomas with derivatives of the three major germ layers in vivo when injected into severe combined immunodeficient mice. Our results showed that pluripotent, stable ESC lines could be derived from fertilized and parthenote-derived rabbit embryos.

Animals↗
PubMed2007

A meta-analysis of human embryonic stem cells transcriptome integrated into a web-based expression atlas.

Microarray technology provides a unique opportunity to examine gene expression patterns in human embryonic stem cells (hESCs). We performed a meta-analysis of 38 original studies reporting on the transcriptome of hESCs. We determined that 1,076 genes were found to be overexpressed in hESCs by at least three studies when compared to differentiated cell types, thus composing a "consensus hESC gene list." Only one gene was reported by all studies: the homeodomain transcription factor POU5F1/OCT3/4. The list comprised other genes critical for pluripotency such as the transcription factors NANOG and SOX2, and the growth factors TDGF1/CRIPTO and Galanin. We show that CD24 and SEMA6A, two cell surface protein-coding genes from the top of the consensus hESC gene list, display a strong and specific membrane protein expression on hESCs. Moreover, CD24 labeling permits the purification by flow cytometry of hESCs cocultured on human fibroblasts. The consensus hESC gene list also included the FZD7 WNT receptor, the G protein-coupled receptor GPR19, and the HELLS helicase, which could play an important role in hESCs biology. Conversely, we identified 783 genes downregulated in hESCs and reported in at least three studies. This "consensus differentiation gene list" included the IL6ST/GP130 LIF receptor. We created an online hESC expression atlas, http://amazonia.montp.inserm.fr, to provide an easy access to this public transcriptome dataset. Expression histograms comparing hESCs to a broad collection of fetal and adult tissues can be retrieved with this web tool for more than 15,000 genes.

Cell Differentiation↗
PubMed2007

Progenitor cells from the porcine neural retina express photoreceptor markers after transplantation to the subretinal space of allorecipients.

Work in rodents has shown that cultured retinal progenitor cells (RPCs) integrate into the degenerating retina, thus suggesting a potential strategy for treatment of similar degenerative conditions in humans. To demonstrate the relevance of the rodent work to large animals, we derived progenitor cells from the neural retina of the domestic pig and transplanted them to the laser-injured retina of allorecipients. Prior to grafting, immunocytochemical analysis showed that cultured porcine RPCs widely expressed neural cell adhesion molecule, as well as markers consistent with immature neural cells, including nestin, Sox2, and vimentin. Subpopulations expressed the neurodevelopmental markers CD-15, doublecortin, beta-III tubulin, and glial fibrillary acidic protein. Retina-specific markers expressed included the bipolar marker protein kinase Calpha and the photoreceptor-associated markers recoverin and rhodopsin. In addition, reverse transcription-polymerase chain reaction showed expression of the transcription factors Dach1, Hes1, Lhx2, Pax6, Six3, and Six6. Progenitor cells prelabeled with vital dyes survived as allografts in the subretinal space for up to 5 weeks (11 of 12 recipients) without exogenous immune suppression. Grafted cells expressed transducin, recoverin, and rhodopsin in the pig subretinal space, suggestive of differentiation into photoreceptors or, in a few cases, migrated into the neural retina and extended processes, the latter typically showing radial orientation. These results demonstrate that many of the findings seen with rodent RPCs can be duplicated in a large mammal. The pig offers a number of advantages over mice and rats, particularly in terms of functional testing and evaluation of the potential for clinical translation to human subjects. Disclosure of potential conflicts of interest is found at the end of this article.

Animals↗
PubMed2004

The transcriptome profile of human embryonic stem cells as defined by SAGE.

Human embryonic stem (ES) cell lines that have the ability to self-renew and differentiate into specific cell types have been established. The molecular mechanisms for self-renewal and differentiation, however, are poorly understood. We determined the transcriptome profiles for two proprietary human ES cell lines (HES3 and HES4, ES Cell International), and compared them with murine ES cells and other human tissues. Human and mouse ES cells appear to share a number of expressed gene products although there are numerous notable differences, including an inactive leukemia inhibitory factor pathway and the high preponderance of several important genes like POU5F1 and SOX2 in human ES cells. We have established a list of genes comprised of known ES-specific genes and new candidates that can serve as markers for human ES cells and may also contribute to the "stemness" phenotype. Of particular interest was the downregulation of DNMT3B and LIN28 mRNAs during ES cell differentiation. The overlapping similarities and differences in gene expression profiles of human and mouse ES cells provide a foundation for a detailed and concerted dissection of the molecular and cellular mechanisms governing their pluripotency, directed differentiation into specific cell types, and extended ability for self-renewal.

Animals↗
PubMed2006

In vitro propagation of spermatogonial stem cells from KM mice.

Spermatogonial stem cells (SSCs) are a unique type of stem cells in that they transmit genetic information to the next generation by producing sperms. Studies of SSC proliferation and differentiation have been hampered by the inability of reconstructing these processes in vitro, particularly in a serum-free culture system. Several groups have reported the long term culture of SSCs during which SSCs self-renew and restore spermatogenesis when transplanted back to recipient testes. However, different protocols and mice with particular genetic background have been used by different laboratories, and the techniques have not been adopted widely. In the present study, we first established a SSC isolation and culture system composed of differential adherence selection of SSCs, serum-free medium and mouse embryonic fibroblast (MEF) feeder cells. SSCs from KM pups could be cultured on MEF feeders in StemPro-34 SFM Medium supplemented with glial cell line-derived neurotrophic factor (GDNF), soluble GDNF family receptor alpha-1 (GFRa1) and basic fibroblast growth factor (bFGF) for 1 month. These SSCs were characterized morphologically and by examining the expression of marker genes. Expression of Oct4 and Sox2, which are crucial factors in embryonic stem cell (ESC) self-renewal, were detected in our cultured SSCs, suggesting that SSCs may share with ESCs some common mechanisms in self-renewal regulation. We also found that LIF had no effect on the proliferation of cultured SSCs derived from KM mice.

Animals↗
PubMed2026

Advanced and underlying therapeutic strategies in transformed small cell lung cancer.

Transformed small-cell lung cancer (T-SCLC) is a clinically important form of histologic transformation and a mechanism of acquired resistance in non-small-cell lung cancer (NSCLC). It is associated with poor prognosis, with a median overall survival of only about 9-13 months. This review summarizes recent advances in the mechanisms, diagnosis, monitoring, and treatment of T-SCLC. Repeat biopsy remains the gold standard for confirming histologic transformation, whereas molecular profiling and liquid biopsy may facilitate early detection and longitudinal disease monitoring. Platinum-etoposide remains the most commonly used clinical standard after transformation, but its benefit is typically transient and durable disease control remains uncommon. Continuation of EGFR tyrosine kinase inhibitors combined with chemotherapy may prolong progression-free survival in selected patients but has not consistently improved overall survival. Anti-angiogenic therapy, particularly anlotinib, and chemo-immunotherapy have shown encouraging activity in selected patients, while emerging strategies targeting DLL3, MYC, SOX2, and epigenetic regulators may broaden the therapeutic landscape. Prospective studies integrating repeat tissue sampling, comprehensive genomic profiling, biomarker-guided patient stratification, pharmacogenomics, functional drug-sensitivity testing where feasible, and integrated multi-omics approaches are needed to advance molecularly guided and individualized treatment for T-SCLC.

advanced therapy↗
PubMed2026

Emodin Induces AIF-Associated Apoptosis and Suppresses Wnt/β-Catenin Signaling in Colorectal Cancer Stem-Like Cells.

Colorectal cancer (CRC) remains a major cause of cancer-related mortality due to therapeutic resistance. Because colorectal cancer stem-like cells (CRCSCs) play a central role in tumor initiation and progression, therapeutic strategies addressing CSC-enriched populations are urgently needed. In this study, we investigated the anticancer effects of emodin, a natural anthraquinone, in CSC-enriched tumorsphere models. Emodin significantly suppressed the viability and self-renewal capacity of HCT116- and SW480-derived CSCs. It induced G0/G1 cell cycle arrest and markedly downregulated stemness-associated markers (CD44, CD133, ALDH1A1, SOX2, NANOG, and OCT4). Importantly, emodin-induced cell death was characterized by mitochondrial dysfunction, increased mitochondrial reactive oxygen species, loss of membrane potential, and nuclear translocation of apoptosis-inducing factor (AIF). This cytotoxicity was not rescued by the pan-caspase inhibitor Z-VAD-FMK, confirming caspase-independent apoptosis. Furthermore, network pharmacology and experimental validation identified GSK3β as a key target. Emodin reduced Wnt/β-catenin signaling by decreasing β-catenin stabilization and nuclear accumulation. Crucially, a rescue experiment utilizing LiCl confirmed that emodin's suppressive effects are mechanistically dependent on the GSK3β/Wnt/β-catenin axis. Collectively, emodin suppresses CRCSC characteristics in vitro by downregulating Wnt/β-catenin signaling and inducing AIF-associated caspase-independent apoptosis, highlighting its therapeutic potential against CRC.

Apoptosis-inducing factor (AIF)↗
PubMed1999

Experimental analysis of the emergence of left-right asymmetry of the body axis in early postimplantation mouse embryos.

The lateral asymmetry of the body axis of mouse embryo is revealed first by the asymmetric expression of genes such as nodal, lefty2 and Pitx2 in the lateral plate mesoderm of the neurulating embryo and subsequently by the looping of the heart tube, the rotation of the body axis and situ solitus of specific visceral organs. Analysis of gene expression in the early gastrula shows that there is a transient asymmetric localization of the transcripts of Cerrl, Fgf8, Hesx1 and Hnf3beta gene in the anterior visceral endoderm, Otx2 and Sox2 in the epiblast and Lim1 in the nascent mesoderm. However, the asymmetric expression is not consistent and varies among the embryos, which may be reflecting the lability of the mechanism for specifying the laterality of the body axis during gastrulation. The plasticity of the process that determines laterality is further manifested by the ability to randomise the expression of the Pitx2 gene in the lateral plate mesoderm in embryos that are grown in vitro. The expression of laterality requires the presence of the node and its axial mesodermal derivatives. In mutant embryos that lack the node and in node-ablated embryos, the loss of the axial notochord is associated with isomerism of the body axis, which is revealed either by the expression of the Pitx2 gene in the lateral plate mesoderm of both sides of the body or the complete absence of expression. Our results are therefore consistent with the concept that the specification of the laterality of the body axis goes through a dynamic phase during gastrulation and the activity of the node and its derivatives is instrumental in the conferment of left-right identity of the embryonic tissues.

Animals↗
PubMed2000

Micro-capillary tube in situ hybridisation: a novel method for processing small individual samples.

We have developed a strategy to individually analyse large numbers of small tissue samples by RNA in situ hybridisation. Samples of approximately 0.4 mm x 0.5 mm are processed in rectangular capillary tubes fitted with nylon mesh and glass beads using standard protocols. Eighteen samples can be assayed simultaneously without loss, and background is low. Specifically, mouse Sox2 RNA expression is examined in the chorion of extraembryonic tissue of 7.5 days post-coitum embryos. This technique works equally well for double RNA labelling and could potentially be used for antibody staining of proteins.

Animals↗