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Making mounts of pollen grains

Found in: Labwork

Permanent slides of pollen grains can be used as a reference for identifying unknown pollen samples. It is therefore important, that the pollen grains remain in an authentic, natural shape.

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The preparation and mounting of the pollen can introduce artifacts: the pollen may lose some of its pigment, start to shrink and shrivel or absorb water and swell. A careful preparation is therefore necessary.

There are several methods of preparing pollen grains, each one offers advantages and disadvantages. I can not give a general rule, it simply depends on the goal of the investigation and on the sample investigated. Pollen from wind-pollinated plants taken from a dry environment are probably best left in a dry condition, and not mixed with a water-based mountant, which may cause them to swell (depends on the osmotic potential of the medium, however). On the other hand, the obtained image quality and resolution may not be satisfactory in such a dry mount. It is a compromise, in which several factors have to be taken into consideration. A microscopy enthusiast, who does not need the slides for identification purposes, will again set different standards (such as avoidance of toxic solvents). People who want to publish their results, in turn, may have to rely on the preparatory technique which is customary in their field of research, for reasons of comparison. I recommend that the different methods are tried out.

Mounting techniques

Glycerol wet mount: Place a small drop of glycerol on a clean slide and tap the anthers of the plant so that the pollen falls into the glycerol. If necessary, carefully separate large chunks of pollen grains by stirring. Place a cover slip on top and seal the sides of the cover slip with nail polish. Use a very small amount of glycerol to make sure that the nail polish has enough area to stick the coverslip to the slide. Glycerol wet mounted slides can be stored for months if there is no leakage. The glycerol will withdraw water from the pollen. If the pollen is not dry, then there is a possibility of the pollen to shrink.

Air mounts (dry mounts): In this case, no liquid mounting medium is used. A cover slip is placed on top of the pollen grains and sealed on the side, either with nail polish or with tape. Nail polish may flow very quickly between cover slip and slide, so it may be best to use a nail polish of high viscosity (by letting some solvent evaporate first).

Glycerol jelly (according to Kisser): This is a very popular mounting medium for pollen. It is phenol-free (antiseptic additive) and therefore non-hazardous. It contains 10g of gelatin, 35ml distilled water and 30ml of glycerol (glycerin). After mounting, the sides of the cover slip need to be sealed. Due to the lack of an antiseptic, it is also necessary to work in a sterile manner, otherwise there is the risk of fungal growth in the medium. Maybe it is a good idea to treat the pollen grains first in alcohol to reduce the chance of fungal contamination by spores. Alternatively, one could experiment by increasing the concentration of glycerol.

Non-water-based mounting media: Euparal is a mounting medium which is not water based. Specimens which are present in alcohol can be directly transferred to Euparal. Place a pollen suspension on the slide and let the alcohol evaporate. Before mounting pollen in Euparal, I recommend that the pollen are first washed in alcohol and then compared to the original shape. Does washing in alcohol result in an unacceptable shrinking of the pollen or unacceptable loss of pigments? If not, then mounting the pollen in Euparal may be an alternative.

Reading materal

I found the following article: Marvels of pollen shown by your microscope (Popular Science, September 1939)
(The article recommends the use of organic solvents (such as xylol/xylene and others) to remove oil from the pollen. I do not recommend this due to health reasons, especially when preparing samples for educational purposes. Still, it gives a nice overview of the topic.)

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