Phase contrast: a taster episode!

We do love those old-fashioned Hertel&Reuss microscopes from the 1970s. Solid build, bargain price. But there is a caveat: those instruments were primarily designed for scholarly education. Only very few of them made their way into scientific institutions. As a consequence, the phase contrast upgrade is a rarity, seldom found for sale. Moreover, a phase contrast system is prone to damages. So a used system might turn out to be completely useless.
But, when one of those phase contrast systems came up on Ebay recently, we decided to take the risk and buy it. And yes, we were lucky, it is in perfect working order!

Fig. 1: One of our Hertel&Reuss "CN-hF" microscopes, fitted with the phase contrast upgrade, both dating back to the 1970s. The phase contrast condenser (under the stage) can be recognized by its big disk shape above the "strict green filter" holder.
The dedicated, "Ph" marked objectives are fitted with internal phase contrast rings. These rings can be seen when looking at their rear side. Each phase contrast objective is intended to be used in combination with a dedicated circle ring shaped diaphragm situated in the condenser disk.

After the installation of the phase contrast the upgrade objective phase rings and the condenser diaphragms should be adjusted as shown in the following image:

Fig. 2: View through the microscope tube with removed eye-piece on the rear side of one of the phase contrast objectives. The objective phase contrast rings and the diaphragm rings must overlap as shown here.

It was relatively easy to find a tardigrade permanent slide with poor contrast, perfectly suited as a test specimen:

Fig. 3: A very bad permanent slide, containing a Macrobiotus tardigrade egg (diameter 98 µm - with the egg protrusions included). Even with fully closed condenser diaphragm this slide is delivering a very poor visual impression in normal brightfield illumination!

But when switching to phase contrast the details come out much better:

Fig. 4: The same egg as shown in fig. 3 but this time in phase contrast. The green color is due to the green filter which should deliver a slightly better resolution - at least in theory.

When converting the green image to black and white a typical phase contrast image with bright contrast circumference lines will be the result:

Fig. 5: Same as fig. 4, but converted to black and white

Resume: Phase contrast can be very helpful for finding poor contrast specimens on your slide. Tiny details might come out more clearly as well.
But one shouldn't investigate EVERYTHING by means of phase contrast. Normal bright field might deliver better resolution in some situations, in particular when using classical bright field illumination enhancements like a decenterable condenser diaphragm. Furthermore, a phase contrast will not reveal the true color of your object, instead present a green image (with the green filter in place) or some brownish muddle (without the green filter), similar to an old black and white worn-out photo print on paper. In this respect phase contrast light microscopy and scanning electron microscopy are kind of siblings: both are definitely unable to reveal the true color of your objects!

The risks of phase contrast upgrading

We think that the upgrade of an existing vintage microscope to a phase contrast system is not a trivial task. It must be kept in mind that a phase contrast system is made up of many parts, all of which must fit together and be well preserved - kind of orchestra. Here are some more detailed informations, most of them based on our Hertel&Reuss microscopes but applicable to many other, similar systems by other brands:

(1) Brand and type compatibility, illumination quality
The components of a phase contrast system are highly specific and not interchangeable among different brands. E.g. a 40x phase contrast objective normally will not work flawlessly in combination with a 40x phaco condenser iris provided by a different company. Furthermore, any upgrade system must fit mechanically in the substage of your microscope: as the phase contrast condenser will replace your classical condenser it must be behave identically with respect to position, centering and height adjustment.
And be warned that the phase contrast will need a strong light source. When working with weak light in combination with the green filter it will be difficult to properly focus on your object.

(2) Completeness
A phaco set is consisting of a bundle of components. In the case of our Hertel&Reuss microscope these were:
- the phase condenser (as already mentioned :-)
- a full set of dedicated phase contrast (Phaco) objectives
- an adjustment eye-piece (for inspection of the objectives' back sides)
- two dedicated, tiny adjustment screwdrivers
- a transparent "strict green" color filter, fitting in the condenser underside
- Adapters: in the case of Hertel&Reuss two threaded adapters will be necessary to properly mount the condensor to the microscope stand.

(3) State of preservation
A phase contrast systems is consisting of many individual parts - all of which should be well-preserved. Each condenser diaphragm might be damaged, the objectives delaminated or scratched. Moreover, former users might have performed problematic "repairs" not visible from the outside.

(4) Adjustment
Take your time, look out for a manual and try to understand the effects of the adjustment screws. Normally they are intended to work against springs. Those screws can fall off, get lost or overtightend. Brute force might ruin everything!

Literature (and more solid information, both in German language)
Dieter Gerlach: Das Lichtmikroskop. p. 127-149 and p. 290-294. Stuttgart 1976.
Gerhard Göke: Moderne Methoden der Lichtmikroskopie. p. 122-138. Stuttgart 1988.

© Text, images and video clips by  Martin Mach  (webmaster@baertierchen.de).
The Water Bear web base is a licensed and revised version of the German language monthly magazine  Bärtierchen-Journal . Style and grammar amendments by native speakers are warmly welcomed.

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