is a new generation confocal microplate imager providing solutions for fully
automated simultaneous high speed and high resolution screening. The system has
environmental control and a fluorescence lifetime imaging (FLIM) capability.
can be used to perform selections based on cell morphology,
protein-protein interactions, protein-localization etc. The range of
morphological alterations and biochemical processes we will be able to
interrogate will be extremely broad due to the open architecture of the
instrumentation and the use of novel biophysical approaches to imaging,
including fluorescence lifetime imaging.
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High resolution screening
The water-immersion objectives of the Opera allow high resolution
high-content screening. This is combined with confocal imaging, where the image
degrading out of focus blur is blocked, meaning that the Opera has can achieve
spatial resolution sufficient to perform screening of morphological changes in
Yeast expressing targeted forms of eGFP and mRFP
A. Chameleon Ti:Saffire pulsed IR laser; B: frequency doubling unit; C: fast
gated ICCD; D: picosecond delay unit.
to the MBF Opera includes the addition of a pulsed violet-laser (IR-laser plus
frequency doubler); fast gated intensified camera and integrated electronics.
Protein-protein interactions can be screened by exploiting fluorescence
resonance energy transfer - FRET between donor and acceptor fluorophores
conjugated to protein pairs on interest.
FLIM is the
best way to quantify FRET and intermolecular FRET between two proteins on
interest needs to be correlated with the acceptor concentration (i.e.
intensity). The PE FLIM-Opera is the only truly integrated commercial system
available that can perform high-resolution confocal steady state imaging as
concurrent confocal FLIM.
The inherent optical sectioning of a spinning disk confocal allows us to acquire
fast z-series of the objects to be analysed. The Opera at MBF is upgraded with
temperature, CO2 and humidity control in the imaging chamber. This allows
acquisition of multi-dimensional times series with high temporal resolution in a
single 384 well plate.
Lower temporal resolution with
more plates can be achieved with a robotic incubator which is integrated to the
Opera with a Twister robot. This allows multiple plates to be removed from the
incubator, screened and placed back to continue their growth prior to