High content screening applied to large-scale cell biology

Abraham, Vivek C.

doi:10.1016/j.tibtech.2003.10.012

Cited by 275 publications

(196 citation statements)

References 31 publications

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“…In principle, this technique can also be scaled to smaller array spots, as liquid handler systems are routinely able to pipette ≤0.1µl. Although the manual analysis of viable cell number performed in this manuscript is not high-throughput, there are several high-content image analysis techniques and software tools available [17,48] that could be readily used in conjunction with this array processing approach to enhance analysis throughput.…”

Section: Discussionmentioning

confidence: 99%

An adaptable hydrogel array format for 3-dimensional cell culture and analysis

et al. 2008

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Hydrogels have been commonly used as model systems for 3-dimensional (3-D) cell biology, as they have material properties that resemble natural extracellular matrices (ECMs), and their cellinteractive properties can be readily adapted in order to address a particular hypothesis. Natural and synthetic hydrogels have been used to gain fundamental insights into virtually all aspects of cell behavior, including cell adhesion, migration, and differentiated function. However, cell responses to complex 3-D environments are difficult to adequately explore due to the large number of variables that must be controlled simultaneously. Here we describe an adaptable, automated approach for 3-D cell culture within hydrogel arrays. Our initial results demonstrate that the hydrogel network chemistry (both natural and synthetic), cell type, cell density, cell adhesion ligand density, and degradability within each array spot can be systematically varied to screen for environments that promote cell viability in a 3-D context. In a testbed application we then demonstrate that a hydrogel array format can be used to identify environments that promote viability of HL-1 cardiomyocytes, a cell line that has not been cultured previously in 3-D hydrogel matrices. Results demonstrate that the fibronectin-derived cell adhesion ligand RGDSP improves HL-1 viability in a dose-dependent manner, and that the effect of RGDSP is particularly pronounced in degrading hydrogel arrays. Importantly, in the presence of 70µM RGDSP, HL-1 cardiomyocyte viability does not decrease even after 7 days of culture in PEG hydrogels. Taken together, our results indicate that the adaptable, array-based format developed in this study may be useful as an enhanced throughput platform for 3-D culture of a variety of cell types.

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Section: Discussionmentioning

confidence: 99%

An adaptable hydrogel array format for 3-dimensional cell culture and analysis

et al. 2008

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“…On the other hand microscopic analysis can also be performed. This is an important tool for evaluating morphological features like cell spreading but will also increasingly be applied to evaluate cellular processes in the context of high content screening 90,91 . The use of such stainings in combination with microscopy however mostly generates qualitative or semi-quantitative data.…”

Section: A Routine In Vitro Methodsmentioning

confidence: 99%

Assessing nanoparticle toxicity in cell-based assays: influence of cell culture parameters and optimized models for bridging the in vitro–in vivo gap

et al. 2013

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“…1B). We coexpressed the mCherry-tagged proteins KLF6, HMGA1, CFLAR, and CRMP1 together with a YFP-tagged HTT exon-1 fragment (YFP-HTTex1Q79) in SH-EP neuroblastoma cells and quantified the formation of YFP-HTTex1Q79 aggregates through high-content fluorescence imaging (Abraham et al 2004). We observed that three of the four selected proteins (HMGA1, CFLAR, and CRMP1) efficiently suppressed YFP-HTTex1Q79 aggregation in SH-EP cells (Fig.…”

Section: Interaction Network Filtering Enables the Identification Of mentioning

confidence: 99%

Systematic interaction network filtering identifies CRMP1 as a novel suppressor of huntingtin misfolding and neurotoxicity

et al. 2015

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Assemblies of huntingtin (HTT) fragments with expanded polyglutamine (polyQ) tracts are a pathological hallmark of Huntington's disease (HD). The molecular mechanisms by which these structures are formed and cause neuronal dysfunction and toxicity are poorly understood. Here, we utilized available gene expression data sets of selected brain regions of HD patients and controls for systematic interaction network filtering in order to predict disease-relevant, brain region-specific HTT interaction partners. Starting from a large protein-protein interaction (PPI) data set, a step-by-step computational filtering strategy facilitated the generation of a focused PPI network that directly or indirectly connects 13 proteins potentially dysregulated in HD with the disease protein HTT. This network enabled the discovery of the neuron-specific protein CRMP1 that targets aggregation-prone, N-terminal HTT fragments and suppresses their spontaneous self-assembly into proteotoxic structures in various models of HD. Experimental validation indicates that our network filtering procedure provides a simple but powerful strategy to identify disease-relevant proteins that influence misfolding and aggregation of polyQ disease proteins.

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High content screening applied to large-scale cell biology

Cited by 275 publications

References 31 publications

An adaptable hydrogel array format for 3-dimensional cell culture and analysis

An adaptable hydrogel array format for 3-dimensional cell culture and analysis

Assessing nanoparticle toxicity in cell-based assays: influence of cell culture parameters and optimized models for bridging the in vitro–in vivo gap

Systematic interaction network filtering identifies CRMP1 as a novel suppressor of huntingtin misfolding and neurotoxicity

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