Hematoxylin
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| General | |||||||||||||||||||
| Surname | Hematoxylin | ||||||||||||||||||
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| Molecular formula | C 16 H 14 O 6 • xH 2 O | ||||||||||||||||||
| Brief description |
light-sensitive yellow-brown solid |
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| properties | |||||||||||||||||||
| Molar mass | 302.29 g mol −1 | ||||||||||||||||||
| Physical state |
firmly |
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| Melting point |
~ 200 ° C (decomposition) |
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| solubility |
slightly soluble in water |
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| As far as possible and customary, SI units are used. Unless otherwise noted, the data given apply to standard conditions . | |||||||||||||||||||
Hematoxylin is a natural chemical compound from the group of indenochromes and polyphenols . It is obtained from the heartwood of the logwood tree ( Haematoxylum campechianum ).
history
The blue wood extracts already used in America were first brought to Europe by the Spanish conquistador Hernán Cortés in 1520 . It was first mentioned in writing as a dye by Robert Hooke in 1665 . Quekett ( 1848 ) and Waldeyer ( 1863 ) introduced this dye into histochemistry. However, the results were not satisfactory as both used this dye directly. The combined application of hematoxylin and a metal salt still used today goes back to Böhmer ( 1865 ).
Extraction
Hematoxylin is obtained by extraction from the heartwood of the logwood tree . For this purpose, an aqueous extract is primarily produced, which is concentrated after extraction with ether. Alternatively, hematoxylin can also be obtained from the aqueous extract by precipitation with urea . A synthetic preparation of hematoxylin is also described.
properties
In its pure form, hematoxylin is colorless to beige. Oxidation by means of atmospheric oxygen (slowly) or a suitable oxidizing agent (e.g. sodium iodate , potassium permanganate , hydrogen peroxide or iodine ) results in the ocher-brown colored hematein in an aqueous or alcoholic solution from hematoxylin . This process is also called (natural or artificial) "maturation". Hematein is a slightly acidic dye. The desired coloring effect is only achieved by adding polyvalent metal cations as a stain . In the presence of polyvalent cations , such as. B. Iron (Fe 3+ ) and aluminum (Al 3+ ) form dye complexes through chelation in the stoichiometry 1: 1. Complexes of hematein and alums are referred to as hemaunas . Haemas show the typical dark blue-violet color and are also known as hematoxylin varnish (actually hematein varnish).
There are over 100 different recipes for the preparation of hematoxylin-containing color solutions. The use of hematoxylin according to Mayer, Harris, Gill (hemaune) and Weigert (iron hematoxylin) is widespread. These color solutions contain the basic color lacquer in an acidic environment (pH 3–4). The binding of the colored varnish to the substrate takes place in turn by chelation with polyvalent cations with anionic structures (e.g. phosphate groups of the nucleic acids of the cell nucleus). This combination of covalent and coordinative bonding is stable enough to survive subsequent rinsing steps in water and alcohol. At the low pH value of the staining solution, the colored structures appear reddish-brown; the typical blue-violet coloration only occurs when the pH value is increased by rinsing in tap water (pH 6.8–7.0).
By varying the pH of a can selectivity be achieved for different structures. While numerous cell structures are stained at a pH value of> 4.5, the staining of cell nuclei dominates at a pH value of 2-3. Compared to the dye hematoxylin, Brazilin lacks a hydroxyl group.
use
It is used particularly in histology for staining cell and tissue structures, such as B. cell nuclei , mitochondria , myelin , elastin and collagen fibers are used. Hematoxylin was also used in dyeing and ink production.
histology
In histology, hematoxylin is used to stain cell and tissue structures, such as B. cell nuclei, mitochondria, myelin, elastin and collagen fibers are used. Numerous variants of the hematoxylin dye solutions have been developed, which differ in particular in the hematoxylin concentration, the choice of oxidizing agent and the choice of the cation (stain) and thus in the coloring behavior. Examples of this are the color solutions according to Harris, Ehrlich , Mayer, Weigert, Heidenhain and Verhoeff.
In combination with hematoxylin staining (cell nucleus staining), counterstaining with a high-contrast cytoplasmic dye is usually carried out. Classically, counterstaining with eosin is carried out ( hematoxylin-eosin staining ), in which cationic / eosinophilic structures (e.g. proteins) are stained. Cell nuclei are usually colored dark blue, while the other cell components appear light pink to pink. Haema are used for HE staining. In connective tissue staining (e.g. Van Gieson staining , Masson trichrome staining ), the iron hematoxylin solution according to Weigert is usually found in combination with other acidic dyes (e.g. picrofuchsin, aniline blue, orange G, acid fuchsin, and many others). In immunohistochemistry , nuclear staining with hemalum is used as a counterstaining. The coloring result shows positive reactions in reddish-brown (DAB) with pale blue seeds.
Individual evidence
- ↑ a b c d data sheet Hematoxylin hydrate, 96% (dry wt.), Water ca 6% from AlfaAesar, accessed on June 19, 2013 ( PDF )(JavaScript required) .
- ↑ a b Entry on haematoxylin in the GESTIS substance database of the IFA , accessed on January 10, 2017(JavaScript required) .
- ↑ Godwin Avwioro (2011): Histochemical Uses Of Haematoxylin - A Review. In: Journal of Pharmacy and Clinical Sciences. Vol. 1, pp. 24-34. (PDF; 303 kB)