Motic Instruments Inc. Canada
October 06, 2006
32
Selecting fluorescence filters requires a thorough understanding of filter technology.
This will enable the user to utilize stain and illumination selection to improve the image
quality of the desired fluorescence signal.
Selection of filter combinations also requires knowledge of the excitation and emission
spectra of the stain.
4.4
Using an oil immersion objective
Oil immersion objectives are labelled with the additional engraving “Oil” and are to be
immersed in oil between the specimen and the front of the objective.
The immersion oil supplied by Motic is synthetic, non-fluorescing and non-resining oil,
with a refractive index of 1.515
Normally, cover glass must be used with oil immersion objectives with a few exceptions.
Deviations from thickness are not important as a layer of immersion oil acts as
compensation above the cover glass.
The small bottle of oil supplied with every immersion objective facilitates application of
the oil to the cover slip.
Remove any air bubbles in the nozzle of the oil container before use.
Immersion oil must be used sparingly.
Freedom from air bubbles must be ensured. To check for air bubbles, remove one
eyepiece, open field diaphragm as far as possible and look at the exit pupil of the
objective within the eyepiece tube (The exit pupil will appear as a bright circle). If it is
difficult to see if there are any bubbles, use phase centering telescope and rotate the
eyepiece part of the centering telescope to focus on the exit pupil of the objective. Air
bubbles in the oil will deteriorate the specimen image. To purge bubbles, swing the
immersed objective forward and backwards by rotating the revolving nosepiece or add
more oil or else wipe off the oil and apply new oil.
Turn the specimen upside down with the cover glass facing the objective.
Slip on a rubber “O” ring between the outer collar
and retractable lens assembly of the oil immersion
objective.
Place a single drop of immersion oil on the lens.
Make contact with cover glass, focus.
View and wipe clean the objective with a lens cleaning tissue.
Note: The rubber “O” ring should be discarded after each use.
Any residual film of oil on immersion type or where it has spread to the surface of dry
type objective will have a discernible, negative effect on the image.
