Live-cell imaging with fluorescence microscopy

Viewing chambers were constructed as previously described (38 (link)). Images were recorded on a Nikon Eclipse Ni light microscope fitted with a Hamamatsu C11440 digital camera and Nikon N Plan Apo λ 63×/1.45NA oil immersion objective. For time-lapse video microscopy, differential inference contrast images were taken at 10-s intervals over 30 min, whereas fluorescence (GFP, mTagBFP2, and mCherry) images were taken every 2 min to prevent bleaching. Time-lapse videos were analysed and annotated using Fiji (83 (link)).

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Koussis K., Withers-Martinez C., Baker D.A, & Blackman M.J. (2020). Simultaneous multiple allelic replacement in the malaria parasite enables dissection of PKG function. Life Science Alliance, 3(4), e201900626.

Publication 2020

Eclipse Ni light microscope C11440 N Plan Apo λ 63×/1.45NA

Fluorescence Immersion Microscope light Video microscopy

Corresponding Organization : London School of Hygiene & Tropical Medicine

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Variable analysis

independent variables

Time-lapse microscopy settings (10-s intervals over 30 min for differential interference contrast, and 2-min intervals for fluorescence)

dependent variables

Microscopy images (differential interference contrast and fluorescence)

control variables

Viewing chambers constructed as previously described (38 (link))
Nikon Eclipse Ni light microscope with Hamamatsu C11440 digital camera and Nikon N Plan Apo λ 63×/1.45NA oil immersion objective
Fiji software used for time-lapse video analysis and annotation (83 (link))

controls

No positive or negative controls were explicitly mentioned.

Annotations

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