Calibration
The system is calibrated by ITG staff, and
new users get these initial calibration
settings at first login and startup.
The calibration is done using transmit-
ted brightfield method.
Calibrate only the objectives you plan to
use in your experiment.
Recalibrate only if you notice drastic
errors in software contour placement
between objectives.
Oil is not needed to Grid tune lenses.
Setup
1. Log in to the computer.
2. Load the Stereo Investigator or Neu-
rolucida software. (Shortcut on desktop
after training)
3. Load the calibration grid slide. Select
the 10x objective on the microscope
and in the software.
4. Push in the push/pull rod to deflect
light to the binocular tube.
5. Move the small 25
m
m grid into center
view and focus.
6. Mouse click on the right top corner
in the software image window to mark
a the reference point. This will help find
the small grid when you are ready to
calibrate higher power objectives; just
select move to reference point and the
software will drive stage to the location
of the small grid. Another option is to
count 8 squares in from top or bottom
of large 250
m
m grid, and then move left
or right until small grid is found.
7. Move to the large 250
m
m grid and se-
lect the lowest power objective you plan
to use in both the microscope and in the
software. Align for Koehler Illumination.
Grid Tune Calibration
8. Select Tools > Grid Tune Current Lens
9. Tell the software the box size:
Large grid =250
m
m
Small grid =25
m
m
10. Drop the anchor at intersection so
vertical dashed line is to the right of the
vertical grid line. The horizontal dashed
line should be just above the horizontal
grid line. The horizontal and vertical
Parcentric Parfocal Calibration
Move to reference point or find the up-
per right corner of small grid.
Do not move the stage during parcentric
parfocal calibration.
Start with highest power objective focus
on corner of small grid.
1. Select Tools > Parcentric Parfocal
Calibration.
dashed lines should lay on the edge of
highest contrast of the actual grid im-
age.
Check the layout of the dashed grid
lines along one vertical column and one
horizontal row; ensure they mirror the
placement of the dashed lines passing
through the origin or anchor point. Ad-
just dashed lines if necessary. Right click
and select finished gridtuning lens.
11. Repeat on all other lenses to be
calibrated. Note the default config does
not have the 100x oil or 50x oil on the
microscope though they have basic
calibrations set in the software.
2. Discard all unnecessary lenses. NL
and SI both use video lenses. Select OK.
3. Follow software prompts. Do not
touch stage X-Y controls or move joystick
X-Y during Parcentric Parfocal steps.
Once completed a dialogue will appear,
ensure all boxes are checked (default).
