Metachromasia

Metachromasia, from meta, a change in the kind of, and chroma, color, refers to the qualitative change which occurs in the color of certain dyes when they interact with other substances. The term was originally applied by Paul Ehrlich to the phenomenon of color change when a dye is adsorbed onto a substrate. Thus, cartilege and other mucopolysaccharide-containing tissues are stained red by toluidine blue. Similarly, Hartley observed that when bromophenol blue solution is added to cetrimide solution the color changes from purplish blue to clear blue. 

The phenomenon has been studied with respect to histochemical staining. It has also been applied to adsorption indicators in titration and to the detection of micellar aggregations. 

Metachromasia is of potential value in model systems for pharmacological phenomena because the strengths of the forces involved are probably in the same range as those involved in certain biological molecular processes.The metachromatic properties of heparin have received considerable attention in this context. 

Metachromasia has been attributed to the formation of ion pairs of micelles.Lewis and co-workershave shown that 

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Nerves stain yellowish-brown with cresyl violet (metachromasia)... 

Methylene Blue is used to detect pectin and phenolic compounds. It is applied to the specimen as a 0.01-0.15% (w/v) aqueous solution, incubated for 30 min. at room temperature, or for 5 min. at 60°C, and then removed by washing in distilled water. Methylene Blue is a blue dye, but it produces a red coloration as a result of the reaction with the phenolic or pectic substances. This phenomenon is referred to as metachromasia, and the cell or tissue components that exhibit it are called metachromatic. 

Toluidine Blue O is dissolved in a buffer with a pH between 4 and 8, or in 70% ethanol or in a 0.05% (w/v) borax solution and kept at 4°C. The specimen is stained for 1-10 min. washed for 1-2 min. in distilled water, or in the buffer used to dissolve the Toluidine Blue O, or in ethanol until all excess stain is removed. When the specimen is viewed under the microscope, different cell components produce colors different colors (metachromasia) DNA is bluish-green, RNA is violet, the middle lamella is red, non-lignified cell walls are red-violet or blue-violet, and polymerized phenolics such as lignin are green or blusih-green. This stain is therefore of general use. 

Figure 2. Chondrocye-like cells in developing mouse mandibular body bone showing metachromasia to TB (a), which immunohistochemically react positively to type 2 collagen (b) and type 1 collagen (c).

The content of long-chain PolyPs may be estimated by measuring the metachromatic effect in the absorption spectrum of toluidine blue (Chernysheva et al., 1971 Leitao et al., 1995 Lorenz and Schroder, 1999). Toluidine blue in an aqueous solution exhibits a concentration-dependent absorption spectrum due to a monomer (A.max, 632 nm)-dimer O.max, 590 nm) equilibrium. The PolyP induced the maximal shift of the absorption spectrum to 545 nm. Nucleic acids also induce metachromasia, but with a shift of about 570 nm with DNA and 590 nm with RNA. Figure 2.3 demonstrates the typical absorption spectra of toluidine blue and toluidine blue with different preparations of PolyPs (Chernysheva etal., 1971). 

In contrast, studies on the effect of ascorbic acid deficiency on the sequence of wound repair in guinea pig (D21) showed an initial decrease in amount of hexosamine and absence of metachromasia. Fibroblasts were poorly oriented and immature, and no collagen formation took place. By the twelfth day, the absence of coUagen and the presence of immature fibroblasts were established, and hexosamine content was found to be high instead of low as in normal repair. 

For possible mutational studies with these loci, it would be technically difficult to detect a reverse mutation accompanied by a loss of metachromasia. On the other hand, by using normal male or female cells, any forward recessive mutations either in one X chromosome or one autosome may be detectable if the expression of metachromasia is unequivocal. However, it may be that metachromasia is not as specific a cellular phenotype as was originally hoped (Nadler et aLy 1969 Taysi et al.y 1969 Milunsky and... 

Because of the possibility of ion association and charge transfer the use of conductometric titration is suggested. Preliminary unpublished conductometric studies by the authors on the reaction between bromophenol blue and quaternary ammonium and arsonium salts failed, within the sensitivity of the instrument used, to detect any conductivity changes. The reaction is, however, accompanied by metachromasia and can be followed spectrophotometrically, showing again the complementary nature of the physical methods available. 

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