Cain, Robert J.; Vanhaesebroeck, Bart; Ridley, Anne J. (2021). CIL:30549, Homo sapiens, blood vessel endothelial cell, monocytic leukemia cell. In Cell Image Library. UC San Diego Library Digital Collections. Dataset. https://doi.org/10.6075/J03B5XS6

TNF-alpha stimulated p110alpha or control siRNA transfected HUVECs previously labelled with celltracker orange dye were grown on collagen matrices in transwell chambers. Celltracker green-labelled THP-1 cells were added to HUVECs, and TEM allowed to proceed toward an MCP-1 gradient in the lower chamber, for either 10 or 60 minutes before fixation. Fixed samples were stained with AlexaFluor 633-conjugated phalloidin to visualize F-actin (grey) and z-stacks collected. Three dimensional reconstructions were produced using Volocity software. A small gamma correction was applied to the red channel in order to visualise weak cell tracker staining at the edges of cell, which are difficult to distinguish upon 3D rendering. Transwells were fixed by immersion in 3.7% (wt/vol) paraformaldehyde before being processed for immunofluorescence microscopy. Images were acquired at room temperature using a confocal microscope (LSM 510 META; Carl Zeiss, Inc.) with three single photomultiplier tube confocal detectors mounted on an inverted microscope (AxioObserver Z1; Carl Zeiss, Inc.) at a magnification of 40 with an oil immersion objective (EC Plan-Neofluar 40× NA 1.30 oil differential interference contrast M27). z sections were acquired at 200-nm intervals by confocal microscopy. Corresponds to Figure 5 from J Cell Biol. 2010 Mar 22;188(6):863-76.

• 2021

• Cain, Robert J.
• Ridley, Anne J.
• Vanhaesebroeck, Bart

Preparation: formaldehyde fixed tissue
Relation to intact cell: dispersed cells in vitro
Item type: recorded image
Imaging mode: single-spot confocal microscopy
Parameter imaged: fluorescence emission
Source of contrast: distribution of a specific protein
Visualization methods: intracellular filling; phalloidin; Alexa Fluor 633; CellTracker Green; CellTracker Orange
Processing history: 3D volume rendering; gamma correction

• Cell Image Library Group ID: 8124

• Homo sapiens

• Actin filament
• Blood vessel endothelial cell
• Cell
• Monocytic leukemia cell

• HUVEC
• THP-1
• Trans-endothelial migration

• data
• image

• No linguistic content; Not applicable

Samplenumber: 30549

Doi: https://doi.org/10.6075/J03B5XS6

Source data

• Source Record in the Cell Image Library: https://doi.org/10.7295/W9CIL30549

Reference

• J Cell Biol. 2010 Mar 22;188(6):863-76. https://www.ncbi.nlm.nih.gov/pubmed/?term=20308428

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2022-08-12