Histochemical staining

The polyene macrolide filipin was isolated in 1955 from the cell culture filtrates of Sterptomyces filipinensis, and was later shown to be a mixture of four components [36]. Although too toxic for therapeutic use, the filipin complex has found widespread use as a histochemical stain for cholesterol and has even been used to quantitate cholesterol in cell membranes [37]. The flat structure of filipin III, the major component of the filipin complex, was assigned from a series of degradation studies [38]. Rychnovsky completed the structure determination by elucidating the relative and absolute stereochemistry [39]. The total synthesis plan for filipin III relied heavily on the cyanohydrin acetonide methodology discussed above. 

A 1,5-hour inhalation exposure of mixed breed rabbits to airborne concentrations of 72 ppm of hydrogen sulfide resulted in ventricular repolarization, while a 5-day, 0.5-hour/day exposure to this concentration resulted in cardiac arrhythmia (Kosmider et al. 1967). Histochemical staining of the myocardial cells revealed a reduction in adenosine tri-phosphate (ATP) phosphohydrolase and NADPH2 oxidoreductase (Kosmider et al. 1967). Cardiac arrhythmia, suggestive of a stimulus transmission disorder, was... 

Table 2.1. Filarial nematodes infected with intracellular bacteria. The method of detection is shown. Neither electron microscopy nor the immuno-histochemical staining techniques used are to be regarded as Wolbachia specific (see note). Positive identification of intracellular bacteria as Wolbachia is shown only where PCR amplified products of rRNA or ftsZ genes have been sequenced. ...

Immuno-histochemical staining of intracellular bacteria in filarial nematodes has been obtained using antibodies against GroELand catalase (Henkle-Duhrsen etal., 1998 Hoerauf etal., 1999) the specificity of these antibodies is unknown, but it is expected to be low because both GroEL and catalase show high level of amino acid conservation throughout the proteobacteria. nd = not done. 

Grizzle WE, Stockard CR, Billings PE. The effects of tissue processing variables other than fixation on histochemical staining and immunohistochemical detection of antigens. J. Histotechnol. 2001 24 213-219. 

Furuya T, Ikemoto K, Kawauchi S, et al. A novel technology allowing immuno-histochemical staining of a tissue section with 50 different antibodies in a single experiment. /. Histochem. Cytochem. 2004 52 205-210. 

Fig. 10 Histochemical staining of Hippeastrum hybridum pollen ( A), Epiphyllium hybridum pollen (B) and part of pistil of Hippeastrum hybridum (C) by red analogue of Ellman reagent. Blue color means the presence of cholinesterase.

Observation Depending on the purpose, the fresh plant samples on slices may be observed immediately or after the histochemical staining with following analysis. 

HRP substrates (Sred) can be selected to give products that can be monitored easily by colorimetric, fiuorometric or chemiluminescent methods. A popular choice in colorimetric assays is 3,3, 5,5 -tetrameth-ylbenzidine (TMB), a colorless substance that gives a blue product (Sox) on oxidation. Important considerations when choosing suitable substrates are cost, safety, sensitivity, solubility, and stability. It is sometimes necessary to use a substrate that gives an insoluble colored product, for example, in histochemical staining or membrane-bound immunoassays. 

The in vitro invasive assay consisted of grafting rat tracheas by implanting into severe combined immxmodeficiency disease (SCID) mice. The tracheas were first injected with the adenocarcinoma cells to promote tumor growth. Invasiveness was determined by histochemical staining of the iso-... 

Ogawa et al. (1997) investigated the formation of H202 and the superoxide radical (02 ) in spinach (Spinacia oleracea) hypocotyls with the use of histochemical stains. Nitroblue tetrazolium (NBT) is used to detect 02 radicals. The colored reaction product formazan was only detected in the vascular tissue of developing spinach hypocotyls if CuZn-superoxide dismutase (CuZn-SOD E.C. 1.15.1.1) was inhibited by DDC... 

VISUALIZATION OF PHENOLIC COMPOUNDS IN PLANTA USING HISTOCHEMICAL STAINS... 

A number of histochemical stains are available to visualize phenolic compounds in the plant, either in thin sections, or applied to whole tissues. De Neergaard (1997) published a series of detailed protocols that provides an excellent source of information. Below follows a summary of the available reagents and the specific compounds they detect. 

Visualizing plant-pathogen interactions involving phenolics with histochemical stains... 

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