The color purple: from royalty to laboratory, with apologies to Malachowski. Biotech Histochem

Department of Anatomy, Microbiology and Pathology, University of Minnesota, School of Medicine, Duluth, Minnesota, USA.
Biotechnic & Histochemistry (Impact Factor: 1.44). 02/2011; 86(1):7-35. DOI: 10.3109/10520295.2010.515490
Source: PubMed


The components of the blood stain, eosin and methylene blue, were introduced by Baeyer and Caro, respectively. Methylene blue was used primarily for detecting Mycobacterium tuberculosis until Ehrlich in 1880 mixed methylene blue with acid fuchsin to produce what he termed a "neutral stain," which allowed differentiation of blood cells. Eight years later, Chęciń ski changed the acidic component of the dye to eosin. Plehn subsequently altered the proportions of eosin and methylene blue to produce a greater range of red and blue hues. In 1891, Malachowski and Romanowsky independently developed stains composed of eosin and "ripened" methylene blue that not only differentiated blood cells, but also demonstrated the nuclei of malarial parasites. A number of "ripening" or "polychroming" techniques were investigated by different groups, but the aqueous dye solutions produced were unstable and precipitated rapidly. Subsequently, methanol was introduced as a solvent for the dye precipitate and techniques were developed that utilized the fixative properties of the methanolic solution prior to aqueous dilution for staining. This avoided the troublesome process of heat fixation of blood films. Giemsa further improved these techniques by using more controlled methods of methylene blue demethylation. In addition, he used measured amounts of known dyes and increased dye stability by adding glycerol to the methanol solvent. With the outbreak of World War I, it became difficult to obtain German dyes outside of Germany; during the World War II, it became impossible. In their effort to improve the inferior American versions of Giemsa's stain, Lillie, Roe, and Wilcox discovered that the best staining results were obtained using pure methylene blue, one of its breakdown products (azure B) and eosin. These three substituents remain the major components of the stain to this day.

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    • "Plasmodium falciparum infections during pregnancy in Africa rarely result in fever and therefore remain undetected and untreated (Hassan et al., 2011). Increased risk of maternal and foetal complications which include maternal anaemia and low birth weight are some of the complications associated with malaria infection in pregnancy (Sutherland and Hallett, 2009; Krafts et al., 2011). Such severe symptoms call for improved, rapid and accurate diagnostic methods. "
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    ABSTRACT: Malaria in pregnancy remains a significant threat in sub-Saharan Africa as it is associated with suboptimal pregnancy outcomes. Due to the fact that the current standard diagnostic method is cumbersome, we assessed the diagnostic performance of an alternate method-fluorescent microscopy, compared to optical microscopy after Giemsa staining- the current standard.Venous blood samples were collected from 407 consenting pregnant women and tested by optical microscopy (OM) after Giemsa staining and by fluorescent microscopy (FM) using the Partec-Cyscope (Partec GmbH, Munster, Germany). All participants were asymptomatic at the time of enrollment. Of the 407 samples tested by OM, 255 (62.5%) were plasmodiumpositive. Of the 255 OM positive samples, 207 were also plasmodium-positive by FM, thus a sensitivity of 81.1% (95% confidence interval (CI): 75.8, 85.8%). Of the 152 samples negative by OM, 75 were also negative by FM, thus a specificity of 49.3% (95%CI: 41.1, 57.6%). The positive and negative predictive values of FM were respectively 72.9% (95%CI: 67.3, 78.0%) and 61.0% (95%CI: 51.8, 69.6%). The percentageagreement between both methods was 69.3% (Kappa=0.32, p-value<0.01). There was moderate agreement between FM and OM results. The low specificity and negative predictive value of FM suggest a high likelihood of false negative results if FM is used in place of OM. OM after Giemsa staining remains the gold standard.
    01/2015; 8(4):1604. DOI:10.4314/ijbcs.v8i4.22
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    • "Malaria is considered to be completely endemic where the enlargement of the spleen in children of 2 to 9 years of age always exceeds 50%, but is lower in adults due to their relative immunity to this disease. The main diagnosis of malaria is through microscopic blood tests [20]. Although blood is the sample often used for diagnosis, both saliva and urine are also used as substitutes for blood [21]. "

    The 12th International Congress of Immunology and Allergy of Iran, Iran; 01/2014
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    • "Unfortunately, RG stains are somewhat unstable and tinctorial results can also vary markedly owing to a number of technical factors. Consequently , the mechanism of the RG stain (Marshall 1978, Wittekind 1983, Wittekind and Kretschmer 1987, Horobin 2011) was not understood until nearly a century after the original reports by Dimitri Romanowsky, Carl Gustav Giemsa, and others who attempted to improve these dye mixtures (Krafts et al. 2011). Actually, details of the mechanism remain a matter of debate. "
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    ABSTRACT: We describe a detailed protocol for using Romanowsky-Giemsa (RG) counterstaining on formalin fixed, paraffin embedded tissue sections that are stained immunohistochemically (IHC) after antigen retrieval using hot acidic citrate buffer. RG staining is easy to perform and provides consistent results that are similar to hematoxylin and eosin (HE) staining. The counterstaining was applied after a variety of antibodies that used the DAB chromogen and the intensity of IHC stained structures was preserved. Moreover, RG counterstaining provided finer cell detail than HE, methyl green or nuclear fast red. A detailed troubleshooting guide is provided for the RG staining protocol.
    Biotechnic & Histochemistry 05/2013; 88(6). DOI:10.3109/10520295.2013.785595 · 1.44 Impact Factor
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