Some new observations and interpretations on the vital staining of leaf epidermal tissue by neutral red

Summary The lower leaf epidermis from 5 plant species was stained with neutral red at 2 pH's (7.1 and 5.6) in the light and dark when the stomata were open or closed. At pH 5.6 no globule (= droplet) formation was observed in the guard cells whether the stomata were open or closed and cell walls possessed a high affinity for the stain. At pH 7.1 globules appeared in guard cells of open stomata, but not closed stomata, within 15 minutes. Anaerobic conditions prevented this globule formation. InZea mays, globules also appeared in subsidiary cells when the stomata were closed and in certain epidermal cells. Where globule formation did not occur increased diffuse staining of certain epidermal cells was considered to be the indication of cell integrity. In old leaf material very large numbers of dark blue globules appeared in epidermal cells ofCommelina diffusa, C. communis andSenecio odoris and this was associated with cell senescence.The staining characteristics were discussed in terms of cellular K⁺, Cl^–, tannin and flavonoid content and vacuolar pH.     

Further observations on the chemistry of Pararosaniline-Feulgen staining

Summary Pararosaniline-Feulgen staining of cells in suspension produces nucleus- and chromatin-specific fluorescence as well as color. Experiments were designed to test postulated reaction mechanisms responsible for the fluorescent staining with the nonfluorescent pararosaniline. The reduction in fluorescent-staining intensity by pretreatment of cells with 2.2×10⁻²M K₂S₂O₅ tends to rule out the alkysulfonic acid pathway; conditions favoring the formation of this intermediate reduce staining intensity. The fluorescence enhancement, observed when cells stained in pararosaniline without K₂S₂O₅ are post-treated with K₂S₂O₅, suggests that there is an initial Schiff-base linkage between pararosaniline and an aldehyde of hydrolyzed DNA, and that this linkage is stabilized in the presence of K₂S₂O₅. Microspectrofluorometer measurements of cells stained at various pararosaniline concentrations in 2.2×10⁻²M K₂S₂O₅, show that the fluorescence emission maximum ranges from about 627 nm at 3.1×10⁻³M pararosaniline to about 604 nm at 3.1×10⁻⁵M. All of the employed staining protocols appear to produce the same fluorescent product, perhaps a heterocyclic pyronin analog formed from pararosaniline. Flow microfluorometric analysis of cells stained in suspension verified that the relative fluorescence intensity represents relative DNA content. Staining at reduced pararosaniline concentration (3.1×10⁻⁴M) reduces the coefficient of variation of the flow microfluorometric histograms, showing that maximum quantitation does not necessarily correlate with maximum staining intensity.     

Further observations on the chemistry of pararosaniline-Feulgen staining.

Pararosaniline-Feulgen staining of cells in suspension produces nucleus- and chromatin-specific fluorescence as well as color. Experiments were designed to test postulated reaction mechanisms responsible for the fluorescent staining with the nonfluorescent pararosaniline. The reduction in fluorescent-staining intensity by pretreatment of cells with 2.2 x 10-2M K2S2O5 tends to rule out the alkysulfonic acid pathway; conditions favoring the formation of this intermediate reduce staining intensity. The fluorescence enhancement, observed when cells stained in pararosaniline without K2S2O5 are post-treated with K2S2O5, suggests that there is an initial Schiff-base linkage between pararosaniline and an aldehyde of hydrolyzed DNA, and that this linkage is stabilized in the presence of K2S2O5. Microspectrofluorometer measurements of cells stained at various pararosaniline concentrations in 2.2x10-2M K2S2O5, show that the fluorescence emission maximum ranges from about 627 nm at 3.1x10-3 M pararosaniline to about 604 nm at 3.1x10-5M. All of the employed staining protocols appear to produce the same fluorescent product, perhaps a heterocyclic pyronin analog formed from pararosaniline. Flow microfluorometric analysis of cells stained in suspension verified that the relative fluorescence intensity represents relative DNA content. Staining at reduced pararosaniline concentration (3.1x10-4M) reduces the coefficient of variation of the flow microfluorometric histograms, showing that maximum quantitation does not necessarily correlate with maximum staining intensity.     

Some observations on the cultivation of kenaf

In the breeding work performed so far on this substitute for strategically important jute, several desirable features have been overlooked, the genetic factors for which may be hidden in the various wild strains of Africa.     

Some observations on the tunnelling of shipworms

The sea-water with which a shipworm inflates its mantle cavity is expelled posteriorly through its dorsal siphon or released anteriorly into its tunnel through the modified pedal opening. Radiographs of shipworms tunnelling through air-filled softwood confirm the presence of water in the blind ends of tunnels. Retracted shipworms can extend themselves and resume tunnelling where they left off, but they must get rid of tunnel water first. They do this by forcing it into the grain at the blind end. It is suggested that tunnelling is continually interrupted by small shrinking movements which ensure that the valves are lubricated. The tendency for shipworms to tunnel with the grain in softwood blocks is accounted for in terms of their need of lubrication. Tunnelling at an angle to the grain and turning round before the solid boundary is reached are accounted for in terms of a reaction to decreased hardness.     

Some observations on the biosynthesis of the plant sulpholipid by Euglena gracilis

1. dl-Cysteine decreases the uptake of ³⁵SO₄²⁻ by Euglena gracilis but does not decrease the relative incorporation of the isotope into sulpholipid; cysteic acid, on the other hand, does not affect the uptake of ³⁵SO₄²⁻ but does dilute out its incorporation into the sulpholipid. 2. Both l-[³⁵S]cysteic acid and dl-+meso-[3-¹⁴C]cysteic acid appear almost exclusively in 6-sulphoquinovose. 3. Molybdate inhibits the incorporation of ³⁵SO₄²⁻ into sulpholipid but not its uptake into the cells; this suggests that adenosine 3′-phosphate 5′-sulphatophosphate may be concerned with the biosynthesis of sulpholipid, and it was shown to be formed by chloroplast fragments. 4. An outline scheme for sulpholipid biosynthesis based on these observations is discussed.     

Some introductory comments on silver staining

Silver staining has become a versatile method for the visualization of specific cell structures and products. The similarity of the impregnation "nuclei" of reduced silver staining to the silver "specks" or "nuclei" of the latent image in photography is noted. "Physical" development (reduction of ionic silver in solution) in silver staining as compared to "chemical" development (reduction of ionic silver remaining in a silver halide crystal) in photographic procedures is briefly discussed.