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Biomedical subjects

R F Chen

Publications and source records attributed to R F Chen.

At least 55 records · Page 3Linked to original sources
PubMed1980

Electrolytic degradation of DNA fluorochromes during flow cytometric measurement of electronic cell volume.

Changes in flow cytometric measurement of DNA content can result from electrolytic chemical degradation of mithramycin, ethidium bromide, and propidium iodide during simultaneous measurement of electronic cell volume. Bench electrolysis also degrades these fluorochromes without changing the quantum yields, even when they are complexed to DNA. In the flow cytometer, electrolytic production of chlorine at the anode is the probable cause of this degradation, since exposure of these fluorochromes to chlorine gas produces the same effect. It is therefore advisable to measure the DNA content distribution alone before simultaneously measuring the DNA content and the electronic cell volume. If unavoidable effects on the DNA distribution are present, narrow forward-angle light scatter should be used as the cell size indicator during dual parameter measurements. Modifying instrument design by reversing electrode polarity might eliminate this problem.

Cytological Techniques↗
PubMed1979

Fluorescence properties of o-phthaldialdehyde derivatives of amino acids.

The fluorescence properties of the products formed by reaction of o-phthaldialdehyde with amino acids and their derivatives, in the presence of thiol compounds, have been studied. The emission spectra, quantum yields, and lifetimes depend on the primary amine and thiol compound used; the observations confirm the report (Simsons, S.S., Jr. and Johnson, D.F. (1978) J. Org. Chem 43, 2886--2891) that the product incorporates molecules of all three types of compounds. The fluorescence quantum yields of o-phthaldialdehyde derivatives of the naturally occuring amino acids ranged from 0.33 to 0.47, using 2-mercaptoethanol as the thiol compound. The fluorescence lifetimes were about 18--20 ns. Lower quantum yields were obtained when mercaptoethanol was replaced by dithiothreitol or ethnethiol. Derivatives of amino acid amides and peptides had quantum yeilds as low as 0.03, due to quenching by the carboxamide group. The intramolecular quenching was relieved by the detergent, sodium dodecyl sulfate, and by dimethylsulfoxide. Monosubstituted lysine exhibited a normal fluorescence, but the di-substituted product was largely quenched, presumably due to interaction between the two isoindole fluorophors. Fluorscence stopped-flow experiments showed that the alpha- and epsilon-amino groups reacted at different rates, with the epsilon-amion group reacting 10 times faster, with a t 1/2 of about 6 s under pseudo first order conditions at pH 9.0 with 10(-3) M o-phthaldialdehyde. The amount of instability shown by the o-phthaldialdehyde derivatives depended on the thiol compound used, the primary amine involved, and the solvent. Cysteine and o-phthaldialdehyde reacted to give an unstable, weakly fluorescent product; but cysteine could be assayed normally if its sulfhydryl was blocked. The o-phthaldialdehyde reagent was discussed in relation to fluorescamine, another reagent for primary amines.

Amino Acids↗
PubMed1976

Immunohistochemical localization of alpha-amylase in barley aleurone cells.

Alpha-Amylase was localized in aleurone cells of barley using immunohistochemical methods. Anti-alpha-amylase antibody was produced by rabbits immunized with enzyme purified from malt diastase and Himalaya variety barley seeds. Immunoelectrophoresis showed that the antibodies to both antigens were immunologically similar, therefore, they were used interchangeably in the localization of alpha-amylase. Fluorescence of 8-10 mum sections of freeze-substituted and paraffin embedded, gibberellic acid (GA)-treated aleurone tissue incubated with rabbit anti-alpha-amylase IgG and rhodamine-conjugated goat-anti-rabbit IgG is localized in the cytoplasm, the nuclear region and the innermost portion of the cell wall. Cytoplasmic immunofluorescence is not associated with a specific organelle but rather is diffusely distributed. The fluorescence of the nuclear region, however, is intense and in thinner (4-5 mum) sections is associated not with the nucleoplasm but with the nuclear envelope and perinuclear region of the cytoplasm. Fluorescence of the cell wall is confined to the inner boundary of the wall corresponding to the resistant wall layer. The immunofluorescent properties of non-GA-treated cells are quantitatively different; fluorescence of these sections is low and diffuse and is particularly reduced in the nuclear region. Electron microscopy shows that GA-treatment results in the proliferation of endoplasmic reticulum (ER) in the perinuclear region of the cell. We suggest that the alpha-amylase localized by immunofluorescence in the perinuclear region of the cell is localized in this ER produced in response to GA treatment. Immunohistochemical localization of alpha-amylase in cells zonated by centrifugation also suggests that the enzyme is intimately associated with the perinuclear area.

Amylases↗