Quantitative Analysis of Digital Microscope Images

Wolf, David E.; Samarasekera, Champika; Swedlow, Jason R.

doi:10.1016/s0091-679x(06)81017-4

Cited by 54 publications

(16 citation statements)

References 2 publications

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“…Quantitative analysis of fluorophore distribution from microscope images assumes that the emitted and detected fluorescence intensities are proportional to the concentration of the probe (Wolf, 2007;Wolf et al, 2007). This assumption, however, needs to be verified before conclusions on probe distribution between intracellular membranes can be made with certainty.…”

Section: Steady-state Distribution Of Fluorescent Sterols In Living Cmentioning

confidence: 99%

Quantitative assessment of sterol traffic in living cells by dual labeling with dehydroergosterol and BODIPY-cholesterol

Wüstner

Solanko

Sokol

et al. 2011

Chemistry and Physics of Lipids

105

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Section: Steady-state Distribution Of Fluorescent Sterols In Living Cmentioning

confidence: 99%

Quantitative assessment of sterol traffic in living cells by dual labeling with dehydroergosterol and BODIPY-cholesterol

Wüstner

Solanko

Sokol

et al. 2011

Chemistry and Physics of Lipids

105

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“…35 Figure 5 shows the various images used for image processing. These include flat-field images which use a single fluorescent solution at all inputs of the device and dark-field images which use nonfluorescent buffer at all inputs of the device.…”

Section: Confocal Microscopymentioning

confidence: 99%

Analysis of a laminar-flow diffusional mixer for directed self-assembly of liposomes

et al. 2012

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The present work describes the operation and simulation of a microfluidic laminarflow mixer. Diffusive mixing takes place between a core solution containing lipids in ethanol and a sheath solution containing aqueous buffer, leading to self assembly of liposomes. Present device architecture hydrodynamically focuses the lipid solution into a cylindrical core positioned at the center of a microfluidic channel of 125 Â 125-lm 2 cross-section. Use of the device produces liposomes in the size range of 100-300 nm, with larger liposomes forming at greater ionic strength in the sheath solution and at lower lipid concentration in the core solution. Finite element simulations compute the concentration distributions of solutes at axial distances of greater than 100 channel widths. These simulations reduce computation time and enable computation at long axial distances by utilizing long hexahedral elements in the axial flow region and fine tetrahedral elements in the hydrodynamic focusing region. Present meshing technique is generally useful for simulation of long microfluidic channels and is fully implementable using COMSOL Multiphysics. Confocal microscopy provides experimental validation of the simulations using fluorescent solutions containing fluorescein or enhanced green fluorescent protein.

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“…For example, plate-to-plate variation in marker intensity may mask the biological effects of interest. The impact of such artifacts could be reduced by appropriate corrections 7,8 such as plate to plate normalization and background subtraction. The profiles generated by PhenoRipper could also be influenced by variations in cell density.…”

Section: Discussionmentioning

confidence: 99%

Rapid Analysis and Exploration of Fluorescence Microscopy Images

Pavie

Rajaram

Ouyang

et al. 2014

JoVE

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Despite rapid advances in high-throughput microscopy, quantitative image-based assays still pose significant challenges. While a variety of specialized image analysis tools are available, most traditional image-analysis-based workflows have steep learning curves (for fine tuning of analysis parameters) and result in long turnaround times between imaging and analysis. In particular, cell segmentation, the process of identifying individual cells in an image, is a major bottleneck in this regard. Here we present an alternate, cell-segmentation-free workflow based on PhenoRipper, an open-source software platform designed for the rapid analysis and exploration of microscopy images. The pipeline presented here is optimized for immunofluorescence microscopy images of cell cultures and requires minimal user intervention. Within half an hour, PhenoRipper can analyze data from a typical 96-well experiment and generate image profiles. Users can then visually explore their data, perform quality control on their experiment, ensure response to perturbations and check reproducibility of replicates. This facilitates a rapid feedback cycle between analysis and experiment, which is crucial during assay optimization. This protocol is useful not just as a first pass analysis for quality control, but also may be used as an end-to-end solution, especially for screening. The workflow described here scales to large data sets such as those generated by high-throughput screens, and has been shown to group experimental conditions by phenotype accurately over a wide range of biological systems. The PhenoBrowser interface provides an intuitive framework to explore the phenotypic space and relate image properties to biological annotations. Taken together, the protocol described here will lower the barriers to adopting quantitative analysis of image based screens.

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Quantitative Analysis of Digital Microscope Images

Cited by 54 publications

References 2 publications

Quantitative assessment of sterol traffic in living cells by dual labeling with dehydroergosterol and BODIPY-cholesterol

Quantitative assessment of sterol traffic in living cells by dual labeling with dehydroergosterol and BODIPY-cholesterol

Analysis of a laminar-flow diffusional mixer for directed self-assembly of liposomes

Rapid Analysis and Exploration of Fluorescence Microscopy Images

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