Live cell imaging of fluorescent parasites was either performed on a Zeiss Axiovert 200 with a magnification of 25x/63x or on a Nikon spinning disc microscope using a 100x objective. Nuclei were visualized with Hoechst.
Spinning disc microscope
Manufactured by Nikon
The Spinning Disc Microscope is a type of confocal microscope that uses a rapidly rotating optical disk to rapidly scan a sample. This allows for high-speed imaging of live cells and tissues with reduced phototoxicity and photobleaching.
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Lab products found in correlation
8 protocols using spinning disc microscope
1
Fluorescent Parasite Live Imaging
2
TIRF Imaging of Microtubule Dynamics
3
Quantifying Displaced Rod-to-RBC Synapses
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Image z-stacks of retina sections for rod-to-RBC synapse analysis were obtained from a Nikon spinning disc microscope at the University of Idaho Imaging Core. Image adjustments were made uniformly across whole images. For displaced synapse data collection, all synapses co-labelled with ribeye and dystroglycan antibodies were counted by hand, in a single in focus z-plane, including one image on either side of z-stack to accommodate for tissue not lying flat. Synapses were determined to be displaced if they were inset beyond one rod spherule into the outer nuclear layer. The number of displaced synapses was divided by the number of total synapses to determine the percent of total synapses that are displaced for each image and data presented are the means.
4
Quantifying Zinc Dynamics in Neuronal Cultures
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Measurements were taken on the Nikon spinning disc microscope using a GFP channel (488 nm excitation, 525/50 nm emission), acquiring images with a 40X (NA 0.95) air objective at 300 ms exposure, EM multiplier 300, 10 MHz camera readout speed, and 15% laser power.
Neuron cultures (DIV 10–14) were washed and incubated at room temperature in RNIM containing 5 µM FluoZin-3 AM for 30 minutes. Samples were washed in RNIM. Baseline measurements were obtained for 1 minute. Cells were then stimulated with a 10 second treatment of high K+ by mixing KNIM 1:1 with the RNIM already present. After 10 seconds, cultures were washed 3x with RNIM and measurements taken for 15 minutes. Calibrations were performed by adding 10 µM TPA for 2 minutes, then washing out with RNIM and adding 10 µM ZnCl2/0.5 µM pyrithione. Measurements were taken until several minutes after a maximum signal was observed.
Neuron cultures (DIV 10–14) were washed and incubated at room temperature in RNIM containing 5 µM FluoZin-3 AM for 30 minutes. Samples were washed in RNIM. Baseline measurements were obtained for 1 minute. Cells were then stimulated with a 10 second treatment of high K+ by mixing KNIM 1:1 with the RNIM already present. After 10 seconds, cultures were washed 3x with RNIM and measurements taken for 15 minutes. Calibrations were performed by adding 10 µM TPA for 2 minutes, then washing out with RNIM and adding 10 µM ZnCl2/0.5 µM pyrithione. Measurements were taken until several minutes after a maximum signal was observed.
5
Imaging of Live Parasites Protocol
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Imaging of live or fixed parasites was either performed on a Zeiss Axiovert 200 with a magnification of 25x/63x or on a Nikon spinning disc microscope using a 100x objective. Nuclei were visualised with Hoechst. For bloodstage parasites, one drop of infected tail blood was added to a microscope slide and covered with a cover glass. Image processing was done with ImageJ.
6
Quantifying Displaced Rod-to-RBC Synapses
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Image z-stacks of retina sections for rod-to-RBC synapse analysis were obtained from a Nikon spinning disc microscope at the University of Idaho Imaging Core. Image adjustments were made uniformly across whole images. For displaced synapse data collection, all synapses co-labelled with ribeye and dystroglycan antibodies were counted by hand, in a single in focus z-plane, including one image on either side of z-stack to accommodate for tissue not lying flat. Synapses were determined to be displaced if they were inset beyond one rod spherule into the outer nuclear layer. The number of displaced synapses was divided by the number of total synapses to determine the percent of total synapses that are displaced for each image and data presented are the means.
7
Quantifying Glucocorticoid Receptor Translocation
8
Whole Retina Imaging Protocol
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