However, a great many plant cells can fluoresce throughout the visible spectrum, and the intensity of this fluorescence is higher than in animal cells due to a great variety of pigments. For instance, the bright red fluorescence of green photosynthetic cells is caused by the presence of chlorophyll pigments. But chlorophyll-free cells of mature pollen and secretory cells (hairs, glandules) fluoresce green or yellow-green because of the presence of phenols, terpenes and other compounds.

Studies that we have conducted in our laboratory show: the fluorescence of intact plant cells makes it possible to diagnose them according to chemical composition, assess changes in the envelope and protoplasm that could occur in the process of growth or under the effect of adverse factors. New vistas are thus open for studying the biochemistry of cells in vivo; up to now studies into metabolic changes of cells were carried out by destructive methods whereby the structure of objects was destroyed. Besides the visual observations, it is now possible to register fluorescence spectra with the aid of a microspectrofluorimeter developed by Dr. Karnaukhov and coworkers for a fluorescence microscope. This new device was patented in many countries in the 1980s. The microspectrofluorimeter allows to register fluorescence spectra of individual cells of plants, animals and microorganisms-even spectra of cell walls. Depending on the specialization of a plant cell, its growth phase and the effect of exogenous factors, such spectra exhibit, on and off, maxima reflecting the conversions of definite substances.

What is the balance of our work? We have found above all that the most valuable information is obtained by analyzing the secretory cells of plant hairs and glandules. One can hardly make them out with the use of an ordinary microscope against the predominant mass of opaque, green or yellow-green tissues. But it is quite different when the plant surface is exposed to UV light in investigations with a fluorescence microscope. Now Drs. Rosh-china and Melnikova have made a microspectrofluorimetric study of intact secretory cells of more than 50 plant species containing fluorescent metabolic products like phenols and terpenoids, alkaloids and polyacetylenes. They were in sharp contrast with the nonfluorescent sites of tissue or with those of a different fluorescent color.

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Also fluorescent was the secretory surface of pollen cells (male gametophyte).

One more example is the specific tissue of the insectivorous vesicular water plant bladderwort ( Utricularia). Its cleft leaves look like a thread with numerous small bladderlike parts (traps) covered with mucus-secreting glandules. Observed with a fluorescence microscope, the surface of a trap looks red due to the green pigment of chlorophyll. Clearly distinct against this background are mucus-containing glandules that luminesce yellow. Their fluorescence spectra (solid line) and the secret (dash line) have an identical maximum (545 nm). Yet this maximum is absent from the fluorescence spectra of nonsecretory cells (dotted line)- instead, there is a 680 nm maximum corresponding to the red luminescence proper to chlorophyll. The yellow fluorescence of mucus glandules is apparently due to the presence of alkaloids.

The luminescence of different type secretory cells is found to depend on their chemical composition. The blue or blue-green luminescence is characteristic of terpenoid-containing hairs of mint ( Mentha); the same is true of comfrey (Symphytum) whose cells accumulate phenols and alkaloids, and of phenol-rich birch buds too. Yellow or orange luminescence is also observed in the secretory cells of marigold (Tagetes) roots abundant in polyacetylenes and alkaloids, while the bulk of their nonsecretory cells fluoresces blue. The secreting surface of pollen (male gametophyte) from the selfsame plants, depending on the chemical composition of the secretae and surface pigments (phenols, carotenoids, azulene, etc.), fluoresces in the blue, yellow, green and red regions of the spectrum.

So, in many plants secretory cells can be identified with a fluorescence microscope. This is important for botanists and experts involved with secretae (secrets), for these substances are a source of a great variety of biologically active products, including those attracting or repelling insects, and also compounds protecting against pests and infection (medicinal plants are a case in point). Some agents keep predators away and thus help preserve useful products.

The fluorescence of the pistil (female gametophyte) and pollen likewise furnishes important information-above all on their physiological condition and fertilization capability, and on their sensitivity to pollution. Here's what we have seen in our laboratory: with pollen germinating in an artificial nutrient medium the fluorescence of viable pollen grains attenuates, while the nonviable grains fluo-resce bright and do not germinate. So we can tell apart both even a few minutes after their pollen has been wetted. Besides, fluorescence analysis enables us to distinguish fairly well the pollen of plants capable of self- pollination (self-compatible) from that of plants incapable of self-pollination (self-incompatible). Say, petunia, which is incapable of self-pollination, has no spectral maxima characteristic of carotenoids and azulenes or else shows low maxima.

In fact, pollen is sensitive to various unfavorable factors. Under their effect or during long storage many pigments on its surface are destroyed with the resulting loss of viability-something that fluorescence analysis allows to

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diagnose right away. For instance, high concentrations of ozone and hydrogen peroxide as well as exposure to UV light destroy carotenoids and, as a consequence, the characteristic fluorescence in the yellow-orange region disappears. In a nutshell, our techniques make it possible to assess the viability of pollen and its fertilization capacity, and that without a long period of germination and sprouting, and without biochemical control. Needless to say, geneticists and plant-breeders would benefit much in their practical work.

But it is not only the practical side of the pistil-and-pollen fluorescence that matters. It's fantastic, but such fluorescence characterizes the ability to identify both the native and the foreign pollen on the pistil's surface. Back in 1994 we first obtained spectra to this effect when for several seconds immediately after the contact of the pistil and pollen of an intact plant we could see in them new maxima substituting the old ones. That occurred only in response to the contact of cells belonging to a native, not a foreign, species. Despite various manipulations under a fluorescence microscope, the native pollen on the pistil germinated normally and, upon fertilization, gave birth to a sound embryo that bore fruit and produced viable seed.

Yet the phenomenon of plant cell fluorescence has not been studied to the full. Further studies will bring us new discoveries and surprises in the kingdom of living organisms.

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