Technical assistance in the acquisition of contocal microscopy ¡mages using microscopy equipment Leica TCSP2 and TCSP5.
In 2013 this Facility has been granted with the ISO9001 certificate in according to the AENOR code ER-0286/2009.
- Confocal Laser Scanning Microscope (CLSM) LEICA TCS SP5 fitted with eight excitation lines (405, 457, 477, 488, 496, 514, 543, 633 nm) from UV, Ar and HeNe lasers and AOTF (Acousto-Optical Tunable Filter) – AOBS (Acousto-Optical Beam Splitter) System. Tandem-scanning system SP5 II standard and resonant scanner which reduces the phototoxicity disminuye land therefore the damage of the biological material being very convenient for in vivo samples.
- Confocal Laser Scanning Microscope (CLSM) LEICA TCS SP2 fitted with nine excitation lines (351, 364, 457, 477, 488, 496, 514, 543, 633 nm) from UV, Ar and HeNe lasers and AOTF (Acousto-Optical Tunable Filter) – AOBS (Acousto-Optical Beam Splitter) System. Visualization ability for UV and various GFP’s.
- Multidimensional Microscopy System Leica AF6000 LX for in vivo cell imaging. It is equipped with an incubation camera with CO2 and temperature controlled. High resolution Hamamatsu C9100-02 CCD camera (until 30 imáges per second), controlled by hardware and software for multiple adquisitión , deconvolution análysis, time-lapse, FRET, etc.
- CCD (Charge Coupled Device) Digital Camera Leica DFC 350 FX coupled with a light microscopy Zeiss Axioplan with transmitted and incident light fluorescence microscope.
- Stereo Microscope Leica MZ16F with color camera Leica DFC490.
Fluorochromes: Furthermore, there are various conjugated fluorescent antibodies available for the users at reduced price (100 μl aliquots).
Clients belonging to the Centro de Investigaciones Biológicas (CIB) have to book in the intranet calendar according to standards that are exposed on the reservations page.
External clients to CIB can contact by e-mail (firstname.lastname@example.org ) or phone ( 034 918 373 112 ext. 4401). The person from the facility who gets the message, will make booking at the intranet calendar. The Client interested on use any equipment of the facility must fill out a “Service request”, which should bring to or send by postal mail, signed by the Head Researcher, who requesting the use of the service
Contact with the Department by phone or e-mail for booking. Timetable: from 8 AM to 17 PM (Monday to Wednesday) and from 8 AM to 15 PM (Thursday & Friday).
CLSM LEICA TCS – SP5 – AOBS Model Leica DMI 6000 CS Inverted microscope with fully motorized objective nosepiece and fluorescence filter cube change,incorporating the contrasting techniques of brightfield and DIC (transmitted light) and incident light fluorescence. Hg 120 w mercury lamp and filter cubes A, GFP, I3, N2.1
AVAILABLE OBJECTIVES Magnification/A.N. Working distance
HCX PL APO CS 10x/0.40 2,1mm HCX PL APO CS 20X/0.7 0,59mm HCX PL APO CS 40x/1.25-0.75 oil 0,1mm HCX PL APO CS 63x/ 1.4-0.60 oil 0,22mm
EXCITATION LINES Diodo Laser 50 mW 405 nm Argon 100 mW with five lines at 458, 476, 488, 514 and 496nm: DiodoP solid state 10 mW at 561 nm. Helio-Neon 10 mW at 633 nm.
CLSM LEICA TCS – SP2 – AOBS Model DMIRE2:
Inverted microscope with fully motorized objective nosepiece and fluorescence filter cube change, incorporating the contrasting techniques of brightfield and DIC (transmitted light) and incident light fluorescence. Hg 50 w mercury lamp and filter cubes for Alexa 350, DAPI, Hoechst, Alexa 488, Sitox, Alexa 546, Alexa 568, Texas Red, GFP, CFP, YFP and CY5.
AVAILABLE OBJECTIVES Magnification/A.N. Working distance
HC PL FLUOTAR 5x/0.15 12 mm HC PL APO CS 10x/0.40 2,2 mm HC PL APO CS 20x/0.70 0,17mm oil y glyc-0,26 water HCX PL APO CS 40x/1.25-0.75oil 0,1mm HCX APO 40X/0.8 W 0,28mm HCX PL APO 63x/1.4-1.60oil 0,1mm HCX PL APO 63x/ 1.20 W 0,22mm HCXPLAPO CS 100x/1.4-0.70oil 0,09 mm
The laser beams in use are:
UV with two excitation lines at 351 and 364 nm. 20 mw Ar with five “ “ “457-477-496: 5 mw and 488, 514nm: 20 mw DiodeP Solid State excitation line at 561 10 mw HeNe with one “ “ “ 633 nm. 10 mw
Acousto-Optical Tunable Filter (AOTF) with eight channels and Acousto-Optical Beam Splitter (AOBS).
Both confocal have control temperature system by thermostatic stage and axis Z control by galvanometric stage.
Spectral Confocal Module based on prism with high sensitivity multiband detection system. Three confocal detection channels for reflection and fluorescence with tunable band-width and central wavelenght in continuous and independent form for each channel. One detection channel for transmitted light. Maximum scanning resolution of 4096 x 4096 pixels (SP2) and 8192 x 8192 pixels (SP5). Maximum scanning speed of 2000 lines per second (SP2) and 16000 lines per second (SP5).
ADQUISITION AND VISUALIZATION
Control panel with seven digital potentiometers for parameter setting of direct exploration. Working station: Software for exploration control, multidimensional series adquisition (x, y, z, t), visualization, bidimensional processing and image quantification. Software for image tridimensional processing. Software for 2D and 3D colocalization analysis. Software for performance of physiological measures. Software for FRET. Software for FRAP. Software of spectral separation.
MULTIDIMENSIONAL MICROSCOPY SYSTEM LEICA AF6000 LX FOR IN VIVO CELL IMAGING
Inverted microscope Leica DMI6000B fully motorized. Contrasting techniques of bright field, phase contrast, and DIC (transmitted light) and incident light fluorescence with various filter cubes: BGR, A4, L5, N3, Y5, CFP/YFP.
AVAILABLE OBJECTIVES working distance
PL FLUOTAR 2,5X/0.07 9 mm HCX PL FLUOTAR 5x/0.15 12 mm HCX PL FLUOTAR 10x/0.30 11 mm. HCX PLAN APO 20x/0.70 0,590 mm HCX PLAN APO 40x/0.75 0,280 mm HCX PLAN APO 63x/1.3 glyc 0,280 mm HCX PLAN APO 100x/1.4oil 0,09 mm
INCUBATION CAMERA FOR LIVING CELLS
The Microscope is equipped with an environmental control system: 1. - Control temperature system (thermostatic stage) and heating unit, can vary the temperature from 3º C above room temperature up to about 60º C 2. - CO2 control above 7,5% 3. - Humidity control Everything is protected by a Climate chamber made of acrylic transparent glass.
ADQUISITION AND PROCESSING RAW DATA
Electron Multiplier CCD Camera Hamamatsu C9100-02 and external filter wheels of excitation, emission and intensity control, offered high speed to obtain images with real time.
Application solutions for FRET by “sensitized emission” method, being could be able to acquire above 10 image sets per second.
For processing: Image optimization, data reduction, deconvolution with the blind method, 3D projection.
EPIFLUORESCENCE MICROSCOPE EQUIPMENT WITH CCD CAMERA AND IMAGE ANALYSIS SYSTEM
Light transmitted and epifluorescence Axioplan Universal Microscope Zeiss with contrasting techniques of bright field and phase contrast. CCD. Digital Camera Leica DFC 350 FX Power Mac G4 Software for image adquisition and Adobe Photoshop 6.0 for image processing.
It allows “in situ” detection of fluorescent antibody conjugates with UV, green and orange-red light. The image sensitivity reception provides a high intensity fluorescence signal and avoids the fading effect after examination. Furthermore, it has a digital image processing system that allows further treatment and quickly and convenient data storage.
Stereo Microscope Leica MZ16 FA
Objective 1× AN 0,14 /oculars 10×. Zoom capability (16:1):7.1× – 115×. Resolution 420Lp/mm. Field of view about 29,6mm – 1,8mm (according to zoom system). 8 Mpx Camera in color Leica DFC490.
APPLICATIONS It allows the observation of samples and the capture of images both, with visible light (diascopy base with Rottermann contrast, bright field, dark field and cold light source), as with fluorescence light (UV, GFP, TXRed). Acquisition of images in different resolutions and formats.