Confocal Microscopy Unit
Mission
Provide support to research studies requiring confocal fluorescence microscopy or video microscopy.
Description
It is a Research Support Service (SAI) equipped with a confocal microscope that enables the acquisition of fluorescence images of fixed or live samples. The confocal microscope obtains optical sections of the volume of a fluorescently labelled sample without the need to invade it, providing information on the expression and localisation of molecules in 3D (xyz) or 4D (3D+time). The most frequent applications are the three-dimensional reconstruction of subcellular structures, cells or tissues; colocalisation studies of markers; and the quantification of parameters of interest (fluorescence intensity, area, morphology, cell density, etc.).
The Confocal Microscopy Unit has its own research lines focused on the design and optimisation of experimental approaches, image processing and data analysis, with the aim of providing better advice to its users. In recent years, it has specialised in the in vivo phenotyping of individual cells in paraffin-embedded or frozen tissues. This service is intended for research groups at the Institute and other institutions that request it.
Services offered
- Spectral confocal microscopy of fixed samples (e.g. cells, tissues, cleared organs).
- Video microscopy (time-lapse) of live samples in a temperature- and CO2-controlled environment, using a digital camera or in confocal mode.
Rates available at www.iisgm.com (Research Support Services).
Technology
CONFOCAL FLUORESCENCE MICROSCOPE model LEICA-SPE (installed in 2015)
Spectral Confocal Module
- AOTF.
- One internal fluorescence detector (PMT) and one brightfield detector.
- Image digitisation: 2048 x 2048 pixels, 8 and 12 bits per channel.
- Scanning speed of 400, 600 and 800 Hz (optional bidirectional); K-scanner.
Laser Module
Four solid-state lasers: 405, 488, 561 and 635 nm.
Computer Equipment
- Computer running Windows 7.
- Software: LAS-X with quantification tools and an additional LAS-AF 3D module (3D reconstructions, projections and rendering).
DMi8 MOTORISED INVERTED MICROSCOPE
APO chromatic objectives:
- ACS APO 10x/numerical aperture 0.30 (dry).
- ACS APO 20x/0.60 immersion in water, glycerol and oil; corrected for 405nm.
- ACS APO 40x/1.30 oil immersion.
- ACS APO 63x/1.30 oil immersion.
- PL-APO 63x/1.3 immersion in water, glycerol and oil.
Illumination with a fluorescence lamp (EL6000, halide lamp) and white LED for transmitted light.
- Galvanometric stage.
- Four fluorescence blocks (type): “Dapi”, “Fitc”, “Rhodamine” and “Cy5”.
- Environmental control (temperature and CO2) for experiments with living cells or tissues.
Digital Camera
Orca Flash 4.0 LT sCMOS camera coupled to the DMi8 microscope.
Installation
The Confocal Microscopy SAI is located in the Experimental Medicine and Surgery Pavilion of the IiSGM. It has 19.2 m2 of space equipped with specific electrical grid and climate control specifications to meet the quality requirements of the services offered.
Contact
Dr. Rafa Samaniego
Head of Unit
E-mail: confocal(ELIMINAR)@hggm.es
Phone: 91 426 92 56
Experimental Medicine and Surgery Pavilion (1st floor)
Hospital General Universitario Gregorio Marañón
C/ Doctor Esquerdo, 46; Madrid 28007