Dye names are written into the image file metadata. However, fluorophores with broadly similar
excitation and emission spectra will use the same laser line and filters. For example, FITC, EGFP,
Alexa488 and Mitotracker Green will all use the 488 nm laser and a bandpass filter of 500 nm to 550 nm
2.1.1 Changing the Dye Selection
1. Remove a selected fluorophore by double-clicking on the dye name
2. To add a dye
a. double-click on the new label from the dye list
b. set the detectors for the new dye by clicking on “Apply”.
Q. My fluorescent stain is not on the list – what do I do? Refer to “Optimizing the Image”.
Table 2. Objective Lenses Available
*Special purpose, not usually kept on microscope.
*Correction collar allows correction for coverslip thickness and refractive index mis-match.
20X LWD is intended for use with plastic culture dishes.
60X/W lens is used for live tissue or fixed tissue mounted in low RI media such as PBS
2.2 Confocal acquisition controls.
1. Focus X2, 4.5 - Scans only the odd lines and replicates them, at 2 µs pixel dwell time.
a. rapidly scanning is very helpful when focusing and moving the stage.
b. may be used for recording rapid fluorescence changes, such as ion measurements.
2. Focus X4, 4.5 - Scans alternate odd lines, replicating the scanned lines to generate an image of
4X reduced resolution in the Y-axis, at 2 µs pixel dwell time for extremely fast scanning.
3. XY Repeat, 4.6 – Scans at full resolution, at the user’s chosen dwell time.
4. XY, 4.7 – Acquires one image, or defined set of images, using the current settings.
5. Stop, 4.8 – Stops any scan, retaining the current image in the Live Image Window.
6. Sequential Acquisition, 4.9 – collect each channel separately, scanning with one laser line at a
time.
Olympus Fluoview-1000 User’s Guide
V.M. Bloedel Hearing Research Center, Core for Communication Research
Center on Human Development and Disability, Digital Microscopy Center
May 11, 2011 12
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