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Chapter 4 System Setup 35
Note: C-mount cameras are shipped with a dust cover lens installed (identifiable by its
red rim). Although this lens is capable of providing images, its throughput is low and the
image quality is not as good as can be obtained with a high quality camera lens. You
should replace the dust cover lens with your own high quality laboratory lens before
making measurements.
To mount an F-mount lens on the camera, locate the large indicator dot on the side of the
lens. There is a corresponding dot on the front side of the cam
era lens mount. Line up the
dots and slide the lens into the mount. Then turn the lens counterclockwise until a click is
heard. The click means that the lens is now locked in place.
Removing either type lens is equally simple. In the case of a C-m
ount lens, simply rotate
the lens counterclockwise until it is free of the mount. In the case of an F-mount lens,
press the locking lever on the mount while rotating the lens clockwise until it comes free
and can be pulled straight out.
Both types of lenses typically have provision for focusing and aperture adjustment, with
the details vary
ing according the make and model of the lens. In addition, in the case of
the F-mount, there is provision for adjusting the focus of the lens mount itself, if
necessary, to bring the focus within range of the lens focus. See the discussion on
page
51 for more detailed information.
Mounting procedures are more complex when mounting to a microscope and vary
according to the m
ake and model of the microscope as discussed in Mounting to a
Microscope, which follows.
Mounting to a Microscope
This section discusses the setup and optimization of your digital imaging system as
applied to microscopy. Since scientific grade cooled CCD imaging systems are usually
employed for low light level microscopy, the major goal is to maximize the light
throughput to the camera. In order to do this, the highest Numerical Aperture (NA)
objectives of the desired magnification should be used. In addition, you should carefully
consider the transmission efficiency of the objective for the excitation and emission
wavelengths of the fluorescent probe employed. Another way to maximize the
transmission of light is to choose the camera port that uses the fewest optical surfaces in
the pathway, since each surface results in a small loss in light throughput. Often the
trinocular mount on the upright microscope and the bottom port on the inverted
microscope provide the highest light throughput. Check with the manufacturer of your
microscope to determine the optimal path for your experiment type.
A rule of thumb employed in live cell fluorescence microscopy is “if you can see the
fluorescence by
eye, then the illumination intensity is too high”. While this may not be
universally applicable, it is a reasonable goal to aim for. In doing this, the properties of
the CCD in your camera should also be considered in the design of your experiments. For
instance, if you have flexibility in choosing fluorescent probes, then you should take
advantage of the higher Quantum Efficiency (QE) of the CCD at longer wavelengths
(contact factory for current CCD specifications). Another feature to exploit is the high
resolution offered by cameras with exceptionally small pixel sizes (6.7 µm for
MicroMAX:1300Y, 1300YHS, and 1300YHS-DIF or 8.3µm for MicroMAX:782Y and
782YHS). Given that sufficient detail is preserved, you can use 2x2 binning (or higher) to
increase the light collected at each “super-pixel” by a factor of 4 (or higher). This will