The Preparation & Mounting of Microscopic Objects

by microscopist Thomas Davies

From the stomachs of common fish—as the cod, sole, haddock, &c.—many specimens of Diatomaceæ may be obtained, but especially from the crab, oyster, mussel, and other shell-fish. Professor Smith states that from these curious receptacles he has taken some with which he has not elsewhere met. To remove them from any of the small shell-fish, it is necessary to take the fish or stomach from the shell, and immerse it in strong hot acid (nitric is the best) until the animal matter is dissolved, when the residue must be washed and treated as the ordinary Diatomaceæ hereinafter described.

Many diatoms are seen best when mounted in a dry state, the minute markings becoming much more indistinct if immersed in liquid or balsam; and for this reason those which are used as test objects are usually mounted dry. Many kinds also are now prepared in this way, as opaque objects to be examined with the lieberkuhn, and are exquisitely beautiful. Others, however, are almost invariably mounted in balsam; but as these will be again referred to in [Chapter III.], and require the same treatment to fit them for the slide, it will not be out of place to describe the cleaning and preparation of them here. As before stated, there is much matter surrounding them which must be got rid of before the “siliceous covering” can be shown perfectly. As, however, we may first wish to become acquainted in some degree with what we have to do, it is well to take a small piece of talc, and place a few of the diatoms upon it. This may be held over the flame of the spirit-lamp until all the surrounding matter is burnt away, and a tolerable idea may be thus obtained as to the quality of our treasure.

In some cases it is well to use this burning operation alone in mounting specimens of diatoms, when they may be placed in their natural state upon the thin glass, burnt for awhile upon the platinum plate, hereafter described, and mounted dry or in balsam.

In the preparation and cleaning of Diatomaceæ, there is little satisfaction unless these operations have been successfully performed, as a very small portion of foreign matter seriously interferes with the object. The mode of preparing them varies even amongst the most experienced. It will be found, therefore, most satisfactory to examine the principal of these separately, although it may be at the risk of some little repetition.

The method which is the most frequently made use of is the following:—Place the “gathering” containing the Diatomaceæ in a small glass or porcelain vessel, add strong nitric acid, and, by the aid of Bunsen’s burner or spirit-lamp, boil for some minutes. From time to time a drop of the mixture may be put upon a slide, and examined under the microscope to see if all foreign matter be got rid of. When the valves are clean, the vessel containing them must be filled with water, and the whole left for an hour or two, so that all the diatoms may settle perfectly. The liquid must then be poured off carefully, or drained away by the aid of a syphon, so that none of the diatoms are removed with it. Indeed, it is well to examine the liquid drained off each time with the microscope, as the finer forms are frequently lost in the washings. The vessel must then be refilled with pure water, allowed to settle, and drained as before. This washing must be repeated until a drop being placed upon a slide and evaporated leaves no crystals. When it is desirable to preserve the diatoms in this state before mounting (which process will be described in another place), they may be placed in a small phial with a little distilled water.

There are many cases in which the above method will not effect a perfect cleansing, as certain substances with which the diatoms are frequently mixed are not soluble in nitric acid. For this reason the following method is resorted to:—Take a quantity of the matter containing the Diatomaceæ and wash first with pure water, to get rid of all the impurities possible. Allow this to settle perfectly and decant the water. Add hydrochloric acid gradually, and when all effervescence has subsided, boil for some minutes by aid of the lamp. When cool and the particles have subsided, decant the hydrochloric and add nitric acid. The boiling must then be repeated until a drop of the liquid when placed under the microscope shows the valves or “frustules” clean. After allowing the diatoms to settle, the acid must be decanted, and pure water substituted. The washing must be repeated as in the former process until all the remains of crystals or acid are removed, when the specimens may be preserved in small phials as stated above.

Such are the usual modes of treating the Diatomaceæ, but there are certain cases in which particular methods are required to give anything like perfect results. Persons of great experience combine a variety of treatments, and thus obtain better and more uniform specimens. Perhaps it will be advantageous to give the young student the process adopted by one of the most successful preparers of these objects; but I will first state the different methods of mounting the cleaned diatoms dry: how to employ Canada balsam and fluid in their preservation will be elsewhere described.

It was before stated that the diatoms when cleansed might be preserved in small phials of distilled water. When required for mounting, shake the phial, and with a thin glass tube or rod take up a drop of the fluid and spread it upon the surface of the slide in the desired position. This must then be allowed to dry gradually, or by the aid of the lamp if necessary, without being shaken or interfered with, otherwise uniformity of dispersion will be prevented. When thoroughly dry, a thin ring of one of the adhesive varnishes—gold-size will be found as good as any—may be drawn round the diatoms, and allowed to dry in a slight degree. The slide and thin glass cover should then be warmed and the latter gently pressed upon the ring of varnish until the adhesion all round is complete.

As some of the diatoms require object-glasses of extremely high power, and, consequently, short focus, to show them, they must be as close to the outer surface of the cover as possible. For this reason they are sometimes placed upon the under side of the thin glass, as follows. Clean the surfaces of the slide and cover, and with the rod or pipe place the liquid containing the diatoms upon the thin glass, and dry as before. Trace the ring to receive the cover upon the slide, and when almost dry, warm both and proceed as above. Whichever of these methods is employed, the outer ring of coloured varnish may be applied as elsewhere described and the slide finished.

The diatoms are also sometimes mounted betwixt two thin glasses, as described in [Chapter I.], so that the light by which they are examined may receive as little interference as possible, and that an achromatic condenser may be brought into focus under the slide.