Core Facilities - ImageStream®X Mark II Imaging Flow Cytometer
ImageStream®X Mark
II charges 2017: to be discussed with Dr
Joana Cabral ( joana.cabral@nuigalway.ie).
Booking and Usage: The
ImageStream®X Mark II is to be operated solely by Joana Cabral. Users should
contact Dr Joana Cabral to discuss and plan their experiments with at least 2
weeks in advance. Further information on the instrument and sample preparation will be provided upon request.
Hours of booking: The ImageStream®X
Mark II is available for sample analysis during normal working hours, from
Monday to Friday, 11am to 5.30pm. The set-up and acquisition is performed by Dr
Joana Cabral only. Out of hours use of the ImageStream®X
Mark II is not permitted.
The revolutionary Amnis ImageStream®X Mark II Imaging Flow Cytometer
(Merck Millipore) combines the speed, sensitivity, and phenotyping abilities of
flow cytometry with the detailed imagery and functional insights of microscopy.
This unique combination provides the tools for numerous applications that
cannot be pursued using either technique alone.
Flow cytometry is a technology that allows quantitative and qualitative
analysis of cell populations at a single cell level, providing multi-parametric
data based on measurements of scattered light and fluorescent signals produced
by cells as they pass through a laser light source. The addition of the image
acquisition feature of Imaging Flow Cytometry greatly increases the amount and
value of the information obtained from each experiment when compared to either
of the techniques alone (Flow Cytometry and Microscopy). This instrument produces
multiple high-resolution images of every cell as it flows (at a rate of up to
5000 cells per second), including brightfield and darkfield (SSC), and up to 10
fluorescent markers with sensitivity exceeding conventional flow cytometers.
When compared to conventional Flow Cytometry, researchers can obtain
numerous additional parameters on their samples and quantify the intensity,
specific location, and distribution of signals within tens of thousands of
cells per sample. These include cell morphology, nuclear shape and subcellular
localization and distribution of target molecules which enable for multiple
features to be analysed in great detail. The majority of the dyes used in
conventional Flow Cytometry can also be detected by the instrument with compensation
being also applied to the images, rendering the users with the ability to use
many more dyes than they could ever use when performing microscopic analysis.
Furthermore, it allows for the visual confirmation that rare events are real
cells and not just artefacts.
objectives (20x, 40x and 60x) and the extended depth
of field option (EDF). EDF keeps the depth of cell in focus without loss
of fluorescence sensitivity and can be of great value when imaging FISH spots, γ-H2AX
foci or other sub-cellular features. The data analysis software, IDEAS®, offers
powerful tools for graphical representation and quantification of more than 85
parameters per cell. Coupled with the short acquisition time, this technology
allows the analysis of statistically relevant numbers of images, which is not
feasible using classical microscopy.
Taken together, the capabilities of the ImageStream®X Mark II system
make it equivalent or superior to traditional flow applications for
multi-parameter cell/particle analysis while also integrating the scope of flow
cytometry and microscopy.
Typical applications include the study of:
· Autophagy
· Apoptosis
· Phagocytosis
· Protein
internalization, nuclear translocation and subcellular localization
· DNA
damage response
· Nuclear
architecture
· Cell
signalling
· Cell-cell
interactions and immune synapse formation
· Cell
cycle
· Nanoparticle
uptake