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CMI | SOP003_rev1.0 

 

 

8

 

Calibration 

8.1

 

Use the silicon wafer for calibration (accepted Raman peak at 521 wavenumbers) 

8.2

 

Choose objective to use and switch to brightfield mode (with edge filter and dichroic mirror off); 
beam splitter should be in Bright Field (BF) mode (Pos 1) 

 

 

 
8.3

 

Set the detector knob, at the side of binocular, to the eyepiece mode (and not the camera mode) 

8.4

 

Set the brightness to 100% in the software (‘Illumination’). Make sure to see some intensity 
changes in the video image; check it by opening and closing the aperture stop (A) at the 
microscope. Typically keep this just slightly open, unless for dim samples/dic imaging. 

Laser coupler 

Slide rightwards(follow 
arrow direction) 

Options are block:  

Open:  

Either open , block for 
633nm and 785 nm 
can be chosen. All 
other positions have 
no filter , used for 
brightfield.  

Filter turret 

Position 1: Brightfield 
imaging  

Position 2: DIC for 
water dipping lens 

Position 3: DIC for all 
other lenses 

Position 4: Raman 
imaging, spectra 

Summary of Contents for SOP003

Page 1: ...hor Changes 1 0 12 5 14 P Owens Initial draft 1 Purpose 1 1 This document specifies the work instructions for the CMI Witec Confocal Raman microscope located in the ground floor Physiology lab Quadran...

Page 2: ...ation in a Laser safety course is strongly recommended for operating this system 3 2 Witec Alpha 500 manual hardcopy in the lab soft copy on the instrument PC and training notes 3 3 The CMI access pol...

Page 3: ...quire a Raman spectrum of most substances in seconds via Charge Coupled Device detectors CCDs that have a wide dynamic range for users to select the appropriate exposure time for their sample Dependin...

Page 4: ...with the system folders located on the shelf above the equipment 7 Start Up Procedure 7 1 Book equipment on CMI web booking system 7 2 Turn on lasers or individual laser if only one is to be used For...

Page 5: ...o be warmed up for at least 30 mins before acquisition while warming up alignment of the microscope can be done 7 7 Click on the Witec Project Control to start the raman software interface 633nm laser...

Page 6: ...tness to 100 in the software Illumination Make sure to see some intensity changes in the video image check it by opening and closing the aperture stop A at the microscope Typically keep this just slig...

Page 7: ...t directed to eyepiece camera Eyepiece camera Aperture stop Field stop Up down toggle between z control xy stage control X Y Stage controls Z Stage controls Z stage moves away from objective Z stage m...

Page 8: ...tching between wavelengths but can be possible as a quick check just to swap in the fibre cables Note Take extreme care when handling the fibre cables as they can be damaged very easily Ensure that th...

Page 9: ...or cells vibrational bands of C H stretching around 700 and 1000 wavenumbers can be used for mapping 9 4 After selecting the large area scan images with better contrast between cells and nanoparticles...

Page 10: ...should be 60 o C 11 Shut Down Procedure 11 1 You can save the experiment parameters for your own scanning recipe by clicking Save Project in File 11 2 Close the laser shutter switch off the laser powe...

Page 11: ...mple on the microscope Demonstrate finding the sample optimizing acquisition and taking spectra Demonstrate changing the fibre cable during the measurement session Demonstrate finishing and shutting d...

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