Updated on April 29, 2011
Apotome Fluorescence Microscope
- ZEISS Axio Imager Z1 (2009)
Axio Imager Z1 (motorized version with high-precision Z drive) with 6
objective lens-10x, 20x, 40x, 40x Oil, 63x Oil and 100x Oil. DIC and DF.
- LED modules (Colibri): 365, 470 and 590nm;
- UV light source Illuminator HXP120.
- Filter sets
(Colibri): 49 (DAPI), 38HE (GFP), 43HE (DsRed), 50 (Cy5), 46HE (YFP), 47 (CFP),
62HE (BFP/GFP) and Analy DIC TransLight, plus other options.
- Digital CCD camera: AxioCam MRm (black/white, 1.4MP)
- Imaging software: AxioVision 4.8
- ApoTome slider and control box
- Heating stage (adjustable RT-60oC)
- Application modules: Extended Focus; AutoMeasure etc.
is an innovative slider module (structured illumination) for fluorescence
microscopy. With a flexible choice of LEDs and filters, the microscope can produce high quality
2D and 3D digital images for fixed or living cells or
Before you start, please note:
scope was designed for fluorescence microscopy with a wide range of
choice of emitters and reflectors. With the additions of ApoTome
apparatus and powerful AxioVision software, flexible
imaging for 2D, 3D (Z-stacks), time-series and multi-channels
- Most components of the microscope are
motorized and can be operated manually (knobs,
buttons), by small touch screen (TFT display) or by
Axiovision (computer screen). However,
computer-driven operations are strongly recommended.
- You are
supposed to be familiar with the ordinary microscope operations
including mounting and focusing your specimen, adjusting lens. Special
care should be taken when you use oil
objective lens (40X, 63X and 100X).
your experiment, you can use one
or a combination of the following
light sources: transmitted light (HAL100 halogen illuminator) or
lights (including single wavelength LEDs, and a metal halide
white light Illuminator HXP120). Remember: transmitted light
and LEDs can be
on/off at your will, but HXP120
needs 20 minutes'
interval. For comparisons among LED, HXP and HBO light
- Software practice is the key to enhance your
- Turn on the power switches in the following sequence:
if you need), Colibri
unit, ApoTome (only if you need), microscope power supply, microscope toggle, and computer. Shutdown
sequence will be the opposite.
- Enter the Windows by logging on your user name and
then click the "Axiovision Rel. 4.8" icon to open the software.
- Load your specimen slide onto the stage, select appropriate
objective lens and focus the point of interest.
- Change lens can be controlled by touch
screen, Axiovision or manually.
switch from oil lens to dry lens. If it happens, remebmer to clean the
sure the transmitted light shutter is "Open" by checking
microscope, touch screen or
To obtain single bright
field (BF), DIC or PH image (using transmitted light).
"Workarea" to open the microscope/camera/processing window. Under the
tab "Microscope", click the tab "Transmitted Light" and make sure the
light is "On" with appropriate voltage.
appropriate condenser turret manually or on touch screen. Focus the
specimen, and adjust slider if required, via eyepieces to get
best imaging effect.
- Click "Cam side port" and select the 100% light to the
- Click "Live", a window will pop up with the rough image
- Click "Property" to open a histograph, and adjust the
quality of the image by playing with "Min/Max", "BestFit" and "Gamma"
- Click "Snap" to capture the image.
- Click "Save" to save your image. Always save as ".zvi"
- To edit the images, click "Scale bar" etc to
label, measure or change colors etc.
fluorescent image (using LED modules).
fluorescence image (using HXP120).
- Select the appropriate
LED module and filter
set, based on the excitation/emission property of your
are 3 LED modules available now: 365, 470 and 590nm; and 7 filter sets:
38HE (GFP), 43HE (DsRed), 50 (Cy5), 46HE (YFP), 47 (CFP), 62HE
(BFP/GFP), plus an Analy DIC TransLight.
- Under the tab
"Microscope", click the tab "Reflected Light".
- Under the "LED" list,
click "desired LED "On" or press the Colibri control panel
- Under the "Reflector"
list to select reflector (filter set).
- Make sure the FL
(reflected light) shutter is "On" and TL "Off".
- Following above
procedures to acquire and save images.
- Turn on the light by
switching on the power and pushing the Shutter button IN on
the HXP120 unit. If this light source has just been turned off, please
wait at least 20 minutes before you start it again!
- Click "Open" External
Shutter under the
"Colibri", or press "Ext"/"Shutter" on Colibri control panel.
- Choose appropriate
filter under the "Reflector" list.
- Then following same
procedures as for LEDs to acquire and save images.
images or multi-channel images (transmitted light and/or fluorescences) in the format of 2D, Z-stack or time-lapse.
or mutiple images with Apotome.
- Click "6D Acquisition"
botton; or under "Workarea", click
- Click "Experiment" tab,
"Load" factory default or the experiment you have saved
- Click "C" tab, set the
channels you want to use.
- There are five default
channels (Green, Blue, Red, BF and DIC). Right-clicking the number icon
to remove, left-clicking to add.
- You can easily arrange
the order of the channel by click-draging the number icon.
- If you want to add a
new channel to the channel pool, click
"Extended parameters", "Channel pool", and then "Add to
channel". However the maximum capacity of multiaquisition is 5
channels. Remember to correctly select the channel settings for your
new channel (e.g. exposure mode at "fixed"). If some setting s are not available for some dyes, ask Guosheng to help.
- Then click "Measure" to
optimize the exposure time for EACH individual channel. "OK" to accept
- Click "Start" at the
bottom to acquire multiple images.
- Following common steps
to save and edit the images.
- You can set "Z-stack" or
"Time Series" for each channel at your will by clicking the tab and following the on-screen instructions.
obtain image with Extended Focus module.
ApoTome power supply must be on when you start the microscope.
- First, following the same
procedures as describrd above to acquire ordinary single image.
- Then, carefully push the Apotome slider into the microscope.
- Click "Apotome" icon on the left-hand menu. You can check if the grid (structured
in ApoTome) was calibrated with your lens and dye used.
- Click "Live" to acquire the
image and snap/process as normal operations.
- You will see the dark raster on the live image.
- The light intensity usually decreases at ApoTome mode. Adjust as required.
- You should use ApoTome slider under reflected light sources and at any acquisition mode such as 2-D, Z-stack or
- The grid should be calibrated in each lens/filter combination. If
not yet, please calibrate the grid focus (NOT Phase) now according to the instructions.
- The ApoTome slider should be off after use. To switch off ApoTome, just
pull out the slider one step (hear a click-sound).
"AutoMeasure" module for image analysis after acquisition either from confocal or apotome microscopes. Please see tutorial at YouTube: http://www.youtube.com/watch?v=hr73-1-bavs.
normal Z-stack images under multidimentional mode and save. Select
"Extended focus" under tab "Processing" to open the saved file. It will
convert the Z-stack images to a single sharp image.
acquire the extended focus image by "Extended Focus" module in
"Workarea". Obtain the "live" image, manually adjust different focal
plane and meantime click the "start" button. The images will
automaticall fuse to single extendedfocus format.
the equipment administrator (Guosheng Liu @ 966-4428).
- Point the cursor to the position and press "F1"
key on the keyboard.
- Tel: 1-800-509-3905
- Email: firstname.lastname@example.org