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Direction of Innovation
CONFOCAL MICROSCOPY
CORE UNIT
Biotechnology Programme | Confocal Microscopy Core Unit
Diego Megías
Core Unit Head
RESEARCH HIGHLIGHTS
Technicians
Manuel Pérez, Joaquim Soriano
The Confocal Microscopy Core Unit is equipped with 3 laser
scanning confocal systems ( Leica SP2 and SP5 ) that incorporate
UV and multiphoton excitation, as well as a white light laser
and a Hybrid Detector. We also have 2 wide field systems,
namely, a Deltavision 4D deconvolution station and a Leica
DMRI6000 system, equipped with microinjection. All the
microscopes are automated and equipped with incubators
for live-cell imaging.
In addition, the Unit has applied high throughput ( HT )
technologies to confocal microscopy using 2 different systems :
ɗɗ An Opera ( Perkin Elmer ) High-Content Screening ( HCS )
system, which allows running HCS experiments on fixed and
live cells in multi-well plates, and enables the monitoring of
cell dynamics ( translocation, cell division, etc.) by means of
fluorescence markers.
ɗɗ A Matrix Screening Application integrated into the SP5
confocal systems, allowing HT feeding of the instrument,
not only in multi-well plates but also in tissue sections.
Figure Microfluidic experiment under different excitation wavelengths
dedicated to a FRET assay ( right ). The Figure shows Fret activity before and
after induction of glucose stimuli.
have incorporated new flow control devices for more accurate
dynamic live cell-based assays in perfusion chambers.
These technological advances increase the level of information
obtained from a sample and allow for the automated HT screening
of cell behaviour in response to different treatments.
We are very proud to have co-organised 2 international
conferences :
ɗɗ The 2nd Spanish National Advanced Microscopy Network
Meeting, which took place at the CNIO, brought together a
large number of international speakers and attendees. This
event explored new advances and techniques in microscopy.
ɗɗ The III Core Management Workshop, focused on the general
management, funding and stability of core units for all
disciplines. s
During 2014, the Confocal Microscopy Unit significantly
refurbished its computational capacity for image analysis by
upgrading its HTS Opera microscope and the Unit’s servers ; we
have developed new programmed routines that have now made
it possible for us to manage, stitch and analyse a huge amount of
images ; something that was previously impossible.
Additionally, we have upgraded our laser scanning confocals with
a new time gating application that has now made it possible for
us to run acquisitions, not only based on intensity, but also taking
fluorophore life time into account, thus allowing us to control
contamination from light reflection and autofluorescence. We
OVERVIEW
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The Confocal Microscopy Unit provides the CNIO Research
Groups with all the standard methodologies as well as the latest
advances in microscopy. The Unit offers access to state-of-the-art
equipment and software packages related to confocal microscopy,
including technical and scientific advice and support to the
CNIO scientists. The Unit is also actively involved in developing,
testing and implementing new microscopy technologies, tools
and imaging applications that could be of interest to Research
Groups at the CNIO. Training activities are also an essential
component of our mission.
SCIENTIFIC REPORT 2014
“ The Confocal Microscopy
Unit is fully committed to the
implementation of advanced
microscopy methodologies in cancer
research, with the aim of creating a
benefit for society by increasing our
understanding of the biology and
disorders of cells that cause cancer.“
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PUBLICATIONS
Miranda-Lorenzo I, Dorado J, Lonardo
E, Alcala S, Serrano AG, Clausell-Tormos
J, Cioffi M, Megias D, Zagorac S, Balic A,
Hidalgo M, Erkan M, Kleeff J, Scarpa A,
Sainz B Jr, Heeschen C ( 2014 ). Intracellular autofluorescence : a biomarker for
epithelial cancer stem cells. Nat Methods
11, 1161-1169.
Alonso-Curbelo D, Riveiro-Falkenbach
E, Perez-Guijarro E, Cifdaloz M, karras
P, Osterloh L, Megías D, Cañón E, Calvo
TG, Olmeda D, Gómez-Lopez G, Graña O,
Sánchez-Arévalo VJ, Pisano DG, Wang
∞∞
H-W, Ortiz-Romero P, Tormo D, Hoeck K,
Rodríguez-Peralto JL, Joyce JA, Soengas
MS ( 2014 ). RAB7 controls melanoma progression by exploiting a lineage-specific
wiring of the endolysosomal pathway.
Cancer Cell 26, 61-76.
Piazzolla D, Palla AR, Pantoja C, Cañamero
M, de Castro IP, Ortega S, Gómez-López
G, Domínguez O, Megías D, Roncador
G, Luque-Garcia JL, Fernandez-Tresguerres B, Fernandez AF, Fraga MF,
Rodriguez-Justo M, Manzanares M,
Sánchez-Carbayo M, García-Pedrero
JM, Rodrigo JP, Malumbres M, Serrano M
( 2014 ). Lineage-restricted function of the
SPANISH NATIONAL CANCER RESEARCH CENTRE, CNIO
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pluripotency factor NANOG in stratified
epithelia. Nat Commun 5, 4226.
Epifano C, Megias D, Perez-Moreno M
( 2014 ). p120-catenin differentially regulates cell migration by Rho-dependent
intracellular and secreted signals. EMBO
Rep 15, 592-600.
Morgado-Palacin L, Llanos S, Urbano M,
Blanco-Aparicio C, Megias D, Pastor J,
Serrano M ( 2014 ). Non-genotoxic activation of p53 through the RPL11-dependent
ribosomal stress pathway. Carcinogenesis
35, 2822-2830.
Alvarez R, Musteanu M, Garcia-Garcia
E, Lopez-Casas PP, Megias D, Guerra
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C, Muñoz M, Quijano Y, Cubillo A, Rodriguez-Pascual J, Plaza C, de Vicente
E, Prados S, Tabernero S, Barbacid M,
Lopez-Rios F, Hidalgo M ( 2014 ). Reply :
‘ Comments on Stromal disrupting effects
of nab-paclitaxel in pancreatic cancer ’. Br
J Cancer 111, 1677-1678.
Hergueta-Redondo M, Sarrió D, Molina-Crespo Á, Megias D, Mota A, Rojo-Sebastián A, García-Sanz P, Morales S, Abril
S, Cano A, Peinado H, Moreno-Bueno G
( 2014 ). Gasdermin-B promotes invasion
and metastasis in breast cancer cells. PLoS
One 9, e90099.
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