Setting Of Epi-Fluorescence - Zeiss Axioskop 2 FS plus Manual

Carl Zeiss
3.3.5

Setting of epi-fluorescence

(1) General principle
The epi-fluorescence technique permits high contrast images of fluorescent substances in typical
fluorescence colors. In the epi-fluorescence microscope, light generated by a high-performance
illuminator reaches the excitation filter (band-pass) via a heat protection filter. The filtered, short-wave
excitation emission is reflected by a dichroic beam splitter and focused on the specimen via the objective.
The specimen absorbs the short-wave emission and then emits the long-wave fluorescence (Stoke's law),
which is now gathered by the objective and transmitted by the dichroic beam splitter. Finally, the rays
pass a barrier filter (long-pass/band-pass), which only allows the long-wave emission from the specimen
to be transmitted.
Excitation and barrier filters, which are both positioned in the FL P&C reflector module together with the
relevant dichroic beam splitter, must be perfectly matched.
(2) Axioskop 2 FS plus and Axioskop 2 FS
− FL P&C reflector module and shutter plate in reflector turret
− HBO 103 or HBO 50 mercury vapor short-arc lamp for incident-light illumination
− HAL 100 halogen illuminator for transmitted-light illumination
Before the epi-fluorescence technique is applied, it is absolutely necessary to adjust the
mercury vapor short-arc lamp in accordance with section 2.1.14 by using the adjusting aid. If
required, re-adjustment must be performed depending on the operation time.
(3) Setting of epi-fluorescence on the Axioskop 2 FS plus and Axioskop 2 FS
Initial epi-fluorescence setting is made much easier if a strongly fluorescent specimen is used first.
Demonstration specimens can also be used first.
• Switch on the HAL100 halogen illuminator.
• Swing in the objective.
• Move condenser turret to position H, transmitted-light brightfield (or also phase contrast), and then
move to the specimen area to be examined.
• Keep light path in the incident-light illuminator blocked at first using the shutter plate on the reflector
turret (3-23/1) or the barrier position of the incident-light filter slider (3-23/5).
3-30
OPERATION
Illumination and contrasting techniques
configuration
MOT
Axioskop 2 FS plus
Axioskop 2 FS
MOT
MOT
B 40-076 e 02/01