Timelapse instructions March 2014
Contents
1. the Incremental base name box is ticked this will automatically number your images under the base name you selected Choose the number of wavelengths you need 24 March 2014 Page 3 J v wl M Sjen A Sjn A m m Arse EE E Acoure cose Multi Dimensional Acquisition ulti Dimensions Experiment M 040456 Multi Dimensional Acquisition 9 Select Light to camera from toolbar on LHS 10 Click Show Live icon live image will appear on screen you may need to adjust the focus 11 Adjust Gain and Exposure for each wavelength To change the assigned wavelengths select an illumination from the dropdown menu If you need to use Auto Scale click on the icon on the LHS of the Live Image Window and select it from the dropdown menu Within the parameters of the gain and exposure that have been set you can alter the image displayed by sliding the triangles on the side bar NB If the uppermost tail of the histogram is not viable then some areas of your image will be overexposed 24 March 2014 Page 4 ter Light to Camera jnl 2 Multi Dimensional Acquisition Main oe o FL GFP FITC Saving Wavelengths WI FL GFPZFI Display Auto Expose No Auto Expose 3000 Alignment Cropping 2 p fo Set Alignment A Frevious Next wh a E E aj FLGFP FITC E E Acquire Close 2 Multi Dimensional Acquisition Illumination FLDAFI Gain avelengths2. your experiment If not delete your old data Make sure you empty the recycle bin afterwards 24 March 2014 Page 2 1 Open MetaMorph software ai NB The software takes a while to load mmapp Shortcut 2 Place slide on microscope stage and _ Objectives available select objective from the menu bar lire Devices Display Process ox TO DOO Ar Mag None 40x 63x oil aA ee Automated do not move by hand NB microscope is inverted so you will need to turn fixed specimens upside down __ ih Light to eyes jnl 3 Select Light to eyes from toolbar on LHS OS Light to Camera nl a DAPL nl 4 Select a channel a iy RFP jn il AF647 jnl Use these buttons to change between O BF jnl Open and close shutter using shutter 5 Check the shutter is open and find state button your sample a ae F g az Color Combine _v Multi Dimensional Acquisition 6 Select Multi Dimensional Acquisition a new window will appea r 7 Review Multi Dimensional Data El Save As CEE x Close All 7 Set the parameters of your experiment using the step by step tabs in the multi dimensional acquisition window 8 Once you ve set everything up you can save your acquisition settings using the Save State button and reload them using the Load State button Save your data on the E Drive In the Saving tab click Select Directory and choose your folder on the E Drive Type in a Base Name for your experiment Check
3. 24 March 2014 Page 1 OPERATING qq INSTRUCTIONS Timelapse epl inv inverted epi fluorescence microscope You must not operate this equipment without prior training from a BALM facility staff member To arrange training and for help please contact Facility Manager Dr Ann Wheeler ext 2406 a p wheeler qmul ac uk Microscopy Technologist Isma Ali ext 2407 i ali qmul ac uk Standard Operating Procedure Basic instructions How to turn the equipment on 1 Switch on the extension lead A 2 Computer and log in B 3 Camera using the button on the camera C 4 Ifyou want to do bright field bright field controller box D How to turn the equipment off 1 Bright field controller box D 2 Computer B 3 Extension lead switch A If you are using CO2 the instructions for use of the Gas cylinder must be followed See next to cylinder Rules of use This microscope should be treated with respect and care at all times This Microscope can only be used by Masters by Research or PhD students Postdocs and members of staff MBBS and BSc Students must be supervised by a member of research staff at all times The microscope lenses must be cleaned after every usage If you have any problems at all with the microscope no matter how trivial they may seem please see a member of BALM facility staff immediately REMEMBER You have 20GB of disk space on this microscope Check before you start if you have room for
4. WAI FL DAPI W2 FL GFP FIT T FL AFSBS F Exposure Display FL AF568 Rhodamine Digitizer FL GFR FITC Phase 10 20 Phase 40x Phase 60s Summary Auto Expose Alignment Cropping xf Y Set Al Live 50 v Best Fit Range 8 Bit Range 0 255 10 Bit Range 0 1023 12 Bit Range 0 4095 14 Bit Range 0 16383 16 Bit Range 0 65535 Scale Within Active Region Scale Image 24 March 2014 Page 5 Multi Dimensional Acquisition Illumination FL GFP FITC 7 Wavelengths Gain fi 12 Click Acquire T WAEFLGEPARI pigie EI a separate picture will be ee taken for each channel Auto Expose No Auto Expose Alignment Cropping xf 4 y 0 Set Alignment SS fl Previous Next wje 3 yen Sel el el rrr BB TB Acauire Close 13 Creating a Merged Image a p s Color Combine click Colour Combine a in toolbar on LHS N select 48 bit 7 Color Combine 7 F l in Colour Combine window pte oE Components Stack C Single Image assign the appropriate images gt Result Depth C gbi f 24 bit C sebi i FED component No Images d GREEN component No Images BLUE component No Images hi Destination C Color Combine Hue rendering for each colour component click Colour Combine N Save combined image to your T Enable hue rendering Eolo Combine folder in tiff format Cancel W
5. hen you have finished transfer all your data to the Z network drive PLEASE TIDY UP Clean lenses throw away used tissue lens tissue dispose of old slides in the yellow sharps bin
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