Gene Expression Analysis of a Single or Few Cells

Zohlnhöfer, Dietlind; Petat-Dutter, Karina; Wendler, Nicole

doi:10.1002/0471142905.hg1108s44

Cited by 3 publications

(2 citation statements)

References 11 publications

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“…Very stringent selection criteria were applied to ensure the best possible separation of the two populations. Owing to the limited number of SP cells available, microarray analysis was performed on a platform optimized for small cell numbers (Klein et al 2005). Although the expression profile of SP vs non-SP cells clearly differed, subtle expression changes could be found for sets of genes only, but not for individual genes, a mandatory prerequisite for the positive identification of suitable adrenal progenitor markers.…”

Section: Discussionmentioning

confidence: 99%

“…To ensure optimal cell quality and purity, FACS sort gates were set restrictively at the most representative areas of SP and non-SP populations. Taken into account the limited number of SP cells resulting from such stringent selection criteria, gene array analysis was performed on a platform, designed for very small cell numbers (Klein et al 2005).…”

Section: Microarray Analysismentioning

confidence: 99%

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The side population phenomenon enriches for designated adrenocortical progenitor cells in mice

Lichtenauer

Shapiro

Sackmann

et al. 2012

Journal of Endocrinology

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Somatic adrenal stem cells are believed to reside in the periphery of the adrenal cortex throughout life for organ maintenance. Herein, we used the side population (SP) phenomenon to enrich for these progenitors, which made up to 0 . 01-0 . 64% of the total cell count. Microarray analysis revealed an expression profile of SP cells, which clearly differed from that of non-SP cells. However, a promising adrenal specific stem cell marker could not be identified. In vitro, SP cells could be maintained in long-term culture, whereas non-SP cells did not proliferate. After 4 weeks of culturing, immunohistochemistry revealed the expression of steroidogenic enzymes such as 3b-HSD, StAR, and P450SCC, suggesting spontaneous differentiation. Interestingly, the quantity of SP cells was significantly diminished in

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Section: Discussionmentioning

confidence: 99%

Section: Microarray Analysismentioning

confidence: 99%

The side population phenomenon enriches for designated adrenocortical progenitor cells in mice

Lichtenauer

Shapiro

Sackmann

et al. 2012

Journal of Endocrinology

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Stochastic profiling of transcriptional regulatory heterogeneities in tissues, tumors and cultured cells

Wang

Janes

2013

Nat Protoc

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Single-cell variations in gene and protein expression are important during development and disease. Cell-to-cell heterogeneities can be directly inspected one cell at a time, but global methods are usually not sensitive enough to work with such a small amount of starting material. Here, we provide a detailed protocol for stochastic profiling, a method that infers single-cell regulatory heterogeneities by repeatedly sampling small collections of cells at random. Repeated stochastic sampling is performed by laser capture microdissection or limiting dilution, followed by careful exponential cDNA amplification, hybridization to microarrays, and statistical analysis. Stochastic profiling surveys the transcriptome for programs that are heterogeneously regulated among cellular subpopulations in their native tissue context. The protocol is readily optimized for specific biological applications and takes about one week to complete.

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Heterogeneity of Astrocytes: From Development to Injury – Single Cell Gene Expression

et al. 2013

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Astrocytes perform control and regulatory functions in the central nervous system; heterogeneity among them is still a matter of debate due to limited knowledge of their gene expression profiles and functional diversity. To unravel astrocyte heterogeneity during postnatal development and after focal cerebral ischemia, we employed single-cell gene expression profiling in acutely isolated cortical GFAP/EGFP-positive cells. Using a microfluidic qPCR platform, we profiled 47 genes encoding glial markers and ion channels/transporters/receptors participating in maintaining K+ and glutamate homeostasis per cell. Self-organizing maps and principal component analyses revealed three subpopulations within 10–50 days of postnatal development (P10–P50). The first subpopulation, mainly immature glia from P10, was characterized by high transcriptional activity of all studied genes, including polydendrocytic markers. The second subpopulation (mostly from P20) was characterized by low gene transcript levels, while the third subpopulation encompassed mature astrocytes (mainly from P30, P50). Within 14 days after ischemia (D3, D7, D14), additional astrocytic subpopulations were identified: resting glia (mostly from P50 and D3), transcriptionally active early reactive glia (mainly from D7) and permanent reactive glia (solely from D14). Following focal cerebral ischemia, reactive astrocytes underwent pronounced changes in the expression of aquaporins, nonspecific cationic and potassium channels, glutamate receptors and reactive astrocyte markers.

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Gene Expression Analysis of a Single or Few Cells

Cited by 3 publications

References 11 publications

The side population phenomenon enriches for designated adrenocortical progenitor cells in mice

The side population phenomenon enriches for designated adrenocortical progenitor cells in mice

Stochastic profiling of transcriptional regulatory heterogeneities in tissues, tumors and cultured cells

Heterogeneity of Astrocytes: From Development to Injury – Single Cell Gene Expression

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