updated 7/14/11
Facility Guidelines
Nikon Laser-scanning confocal microscope
1. All users must undergo basic training on the instrument to be approved for operation.
2. If you would like to use the microscope, schedule training and/or usage time with Lacy
Nelson. The laser units are operated with keys which will need to be checked-out.
3. All users must log-in at every use and record usage times.
4. All image files should be saved in a folder named as your lab PI’s name. Files that are
not clearly identified or not stored in the correct folder(s) are subject to deletion. It is
suggested that you immediately copy data and take it with you. The computer is not
currently networked.
5. All users are responsible for keeping the microscope in good working condition and
keeping the work area clean. Users are responsible for supplying their own slides, cover
slips, and any other supplies. Any samples, slides or supplies left in the working area
are subject to disposal. Ensure that everything is properly powered-down and the scope
is covered before you leave.
6. User costs will be $25 per hour, and will be charged in increments of one-half hours (i.e.,
any usage between 61-90 min. will be charges as 1.5 hr, and so on). There is a one-hour
minimum charge per use. A University cost center number should be provided upon
reserving time on the instrument.
7. Please complete as much sample preparation outside the facility as possible. Samples
(such as whole plants) in soil or potting medium should not be brought into the facility.
8. The purchase costs of the microscope were provided by an NSF-EPSCoR grant. If you
publish images captured with the scope, please acknowledge support of the grant in
your work. If you provide Dr. Korth with a citation from published work where the grant
was acknowledged, your lab will receive one free hour of beam time.
Suggested text for the acknowledgement is: “Confocal images were obtained on an
instrument provided by the P3 Center, funded through the Arkansas ASSET Initiative II
(EPS-1003970) by the National Science Foundation and the Arkansas Science and
Technology Authority”.
The microscope is located in the Rosen Alternative Pest Control Center, room 215.
Primary contacts:
Lacy Nelson ldnelson@uark.edu
Phone: 575-5378 or 575-7081
Ken Korth kkorth@uark.edu
Phone: 575-5191
updated 7/14/11
Instrument highlights:
Nikon 90i upright scanning laser confocal microscope
10X, 20X, 40X, and 60X objective lenses
Laser scanning capabilities, including:
-scanning lasers at 405/488/561/640 nm
-scan head and controller
-detector unit with three high-sensitivity, low-noise photomultiplier tubes with access
for up to two sets of interchangeable dichroic mirrors and emission filters
-detector board for two additional channels for fluorescence and/or transmitted light
-laser-safety interlock for use with digital imaging head
Fluorescence components, including:
-Motorized digital imaging head
-illumination system with manual light attenuator
-excitation/emission filter sets for detection of DAPI/GFP/TRITC
DIC/transmitted light detection, including:
-Motorized scanning transmission detector for brightfield, phase contrast, and DIC
-required DIC modules and sliders for 10X/20X/40X/60X
-rotatable polarizer
Image acquisition software with easy, one-button switching between confocal and
regular microscope modes. All settings and procedures required for live image capture
available in a single window.
Epi- and diascopic aperatures with automatic change to correct position for each
objective
High quality confocal image capture, up to 2048x2048 pixel resolution and 12-bit
grayscale
The three PMTs allow for detection of three fluorescent channels simultaneously
updated 7/14/11
Basic instructions for use of the Nikon Confocal Microscope
Take care not to bend or kink any of the cables. Some contain optic fibers and they
will break if mishandled. Also, ensure that the fluorescence light source is off when
finished – the bulb has a finite lifespan and is expensive to replace. The minimum on-time
for the bulb is five minutes, so never turn it off unless it has been left on for the minimum
time.
1. Start-up (IMPORTANT: please perform all steps in the order listed)
1. Lasers—turn four keys (labeled ‘1’) to the “on” position [turn on only the laser(s)
that you will be using].
2. Controller (located below the lasers, labeled ‘2’, when the ‘ready’ light comes on,
it is okay to start the software)
3. Start Nikon microscope power supply (in the center, under air table, labeled ‘3’)
4. If needed, start the fluorescence power supply (on left side, under air table,
labeled ‘4’)
2. Open “EZ-C1 3.91” program on the desktop; you will also use NIS-Elements AR 3.2
software to view and manage images.
3. Use digital Imaging Head (DIH) button on front of scope to set viewer. This must be on
“Bino” position to view the sample via eyepieces. Set to FP (front port) for laser scanning.
4. Within the software, the icon at the top that looks like the scanner head puts the
microscope in confocal mode.
-You can set-up your own start-up mode with your own default settings
-Start your scanning with minimal laser power, start “Gain” at half-strength
5. Shut-down (IMPORTANT: please perform all steps in the order listed)
1. Turn off laser controller (labeled ‘2’).
2. Turn off each of the lasers (labeled ‘1’).
3. Turn off microscope power supply (labeled ‘3’).
4. Turn off fluorescence power supply (labeled ‘4’).
Cover scope and remove all supplies and samples from the area.