On the nature of Romanowsky dyes and the Romanowsky Giemsa effect

ArticleinClinical & Laboratory Haematology 1(4):247-62 · February 1979with484 Reads
DOI: 10.1111/j.1365-2257.1979.tb01090.x · Source: PubMed
Abstract
This paper reviews the nature of Romanowsky staining and the relationship between Romanowsky dyes and the Romanowsky-Giemsa effect (RGE). On blood and bone marrow smears the RGE is characterized by a purple colouration of nuclei and neutrophil granules. The nuclear purple contrasts strongly with the blue cytoplasmic staining of cells rich in RNA. Requirement for the occurrence of RGE are: I A cationic dye: The best dye is azure B and, though azure A gives the nuclear purple colour, the cytoplasmic blue is inferior. No other cationic dye such as methylene blue is suitable. 2 An anionic dye: Most commonly eosin Y is used, but it can be replaced by the erythrosins. Full halogenation of the fluorescein (four atoms of bromine or iodine) is not necessary. Phloxine and rose bengal are unsuitable. 3 An appropriate substrate: These are proteins with acidic side groups or proteins bound to a polyanion. For the interaction with the dyes substrates must provide a suitable three-dimensional network which is why the RGE is not obtained in solutions. A tentative theory of RGE is advanced and briefly discussed.
    • "Tissues were collected for screening for zoonotic agents and the preparation of cell lines. Romanowsky-Giemsa staining was performed as described in detail by [26]. "
    [Show abstract] [Hide abstract] ABSTRACT: Approximately 60% of emerging viruses are of zoonotic origin, with three-fourths derived from wild animals. Many of these zoonotic diseases are transmitted by rodents with important information about their reservoir dynamics and pathogenesis missing. One main reason for the gap in our knowledge is the lack of adequate cell culture systems as models for the investigation of rodent-borne (robo) viruses in vitro. Therefore we established and characterized a new cell line, BVK168, using the kidney of a bank vole, Myodes glareolus, the most abundant member of the Arvicolinae trapped in Germany. BVK168 proved to be of epithelial morphology expressing tight junctions as well as adherence junction proteins. The BVK168 cells were analyzed for their infectability by several arbo- and robo-viruses: Vesicular stomatitis virus, vaccinia virus, cowpox virus, Sindbis virus, Pixuna virus, Usutu virus, Inkoo virus, Puumalavirus, and Borna disease virus (BDV). The cell line was susceptible for all tested viruses, and most interestingly also for the difficult to propagate BDV. In conclusion, the newly established cell line from wildlife rodents seems to be an excellent tool for the isolation and characterization of new rodent-associated viruses and may be used as in vitro-model to study properties and pathogenesis of these agents.
    Full-text · Article · Jul 2011
    • "In particular, the morphology can infl uence rate of formation of the color purple. This occurs fi rst in the thinner portions of the preparation (Wittekind 1979, Horobin et al. 1989). It is intriguing that similar rate effects also have been reported in cell-free DNA fi lms of varying thickness (Friedrich et al. 1990). "
    [Show abstract] [Hide abstract] ABSTRACT: An introduction to the nomenclature and concept of "Romanowsky stains" is followed by a brief account of the dyes involved and especially the crucial role of azure B and of the impurity of most commercial dye lots. Technical features of standardized and traditional Romanowsky stains are outlined, e.g., number and ratio of the acidic and basic dyes used, solvent effects, staining times, and fixation effects. The peculiar advantages of Romanowsky staining are noted, namely, the polychromasia achieved in a technically simple manner with the potential for stain intensification of "the color purple." Accounts are provided of a variety of physicochemically relevant topics, namely, acidic and basic dyeing, peculiarities of acidic and basic dye mixtures, consequences of differential staining rates of different cell and tissue components and of different dyes, the chemical significance of "the color purple," the substrate selectivity for purple color formation and its intensification in situ due to a template effect, effects of resin embedding and prior fixation. Based on these physicochemical phenomena, mechanisms for the various Romanowsky staining applications are outlined including for blood, marrow and cytological smears; G-bands of chromosomes; microorganisms and other single-cell entities; and paraffin and resin tissue sections. The common factors involved in these specific mechanisms are pulled together to generate a "universal" generic mechanism for these stains. Certain generic problems of Romanowsky stains are discussed including the instability of solutions of acidic dye-basic dye mixtures, the inherent heterogeneity of polychrome methylene blue, and the resulting problems of standardization. Finally, a rational trouble-shooting scheme is appended.
    Full-text · Article · Feb 2011
    • "In particular, the morphology can infl uence rate of formation of the color purple. This occurs fi rst in the thinner portions of the preparation (Wittekind 1979, Horobin et al. 1989). It is intriguing that similar rate effects also have been reported in cell-free DNA fi lms of varying thickness (Friedrich et al. 1990). "
    [Show abstract] [Hide abstract] ABSTRACT: Normal blood smears were stained by the standardised azure B-eosin Y Romanowsky procedure recently introduced by the ICSH, and the classical picture resulted. The effects of varying the times and temperature of staining, the composition of the solvent (buffer concentration, methanol content, & pH), the concentration of the dyes, and the mode of fixation were studied. The results are best understood in terms of the following staining mechanism. Initial colouration involves simple acid and basic dyeing. Eosin yields red erythrocytes and eosinophil granules. Azure B very rapidly gives rise to blue stained chromatin, neutrophil specific granules, platelets and ribosome-rich cytoplasms; also to violet basophil granules. Subsequently the azure B in certain structures combines with eosin to give purple azure B-eosin complexes, leaving other structures with their initial colours. The selectivity of complex formation is controlled by rate of entry of eosin into azure B stained structures. Only faster staining structures (i.e. chromatin, neutrophil specific granules, and platelets) permit formation of the purple complex in the standard method. This staining mechanism illuminates scientific problems (e.g. the nature of 'toxic' granules) and assists technical trouble-shooting (e.g. why nuclei sometimes stain blue, not purple).
    Article · Jan 1987
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