RU2624869C1 - Method for nervous structures identification in dentistry system - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: object is fixed for 3-5 days in a solution containing concentrated formic acid - 3.5 ml, chloral hydrate - 3.5 g, distilled water - 100 ml, the fixing solution i replaced 2-3 times a day. Further, the washed object under investigation, depending on its size, is placed into a mixture of alcohol and ammonia for 30-60 minutes with further washing in distilled water for 20-30 minutes. Impregnation is carried out in the thermostat for 5-7 days in 5% solution of silver nitrate until the object becomes light brown, rinsing it with distilled water 10 minutes before restoration and 10-15 minutes before laying, shading, cutting and embalming.

EFFECT: use of the method allows to accurate detection of unequal structures in the dentoalveolar system.

Description

The invention relates to medicine, in particular to dentistry, and can be used in neurostomatology to identify nerve structures in the dentofacial system at a neurohistological level.

At the disposal of neuromorphologists there is a wide selection of techniques for detecting nerve structures in tissues [Rasskazova E.I. Methods of impregnation of the neuroplasm of various peripheral nerve fibers and endings / E.I. Rasskazova // Questions of psychiatry. In the book. Scientific work of the Institute of Psychiatry. M., Medicine, - 1956. - S. 349-353].

However, the use of recommended prescriptions and conditions in practice often does not allow to obtain the desired result. The detection of nerve components in the bones of the maxillofacial region is no exception. Incomplete identification of the components of the nervous apparatus is partially explained by the biochemical identity of organs and tissues [Rasskazova E.I. Methods of impregnation of the neuroplasm of various peripheral nerve fibers and endings / E.I. Rasskazova // Questions of psychiatry. In the book. Scientific work of the Institute of Psychiatry. M., Medicine, - 1956. - S. 349-353].

One of the negative aspects of detecting nerve elements in bone tissue is decalcification during fixation of the studied objects and the fixation method itself ultimately negates the detection of nerve structures in the teeth and jaws due to their autolysis with acids [Zhabotinsky Yu.M. Normal and pathological morphology of a neuron. L .: Medicine, 1965. - 328 p.]. Decalcifications should be preceded by a good, thorough fixation. For these purposes, there are various mixtures based on formalin - Gels, Heidengain, Stive, Fleming, Delaunay liquids (1936), Schafer (1902), Ebner and several others, in which various acids are used as decalcification, the concentration range of which is very variable. Use: trichloroacetic, picric, nitric, sulfuric acids and their mixtures in different volume combinations. Moreover, many authors who proposed their methods are unanimous in the opinion that hydrochloric and nitric acids as such are not suitable for decalcification, since they completely dissolve the basic structural components of bone tissue [Rasskazova E.I. Methods of impregnation of the neuroplasm of various peripheral nerve fibers and endings / E.I. Rasskazova // Questions of psychiatry. In the book. Scientific work of the Institute of Psychiatry. M., Medicine, - 1956. - S. 349-353].

The best option for decalcification is formic acid, which is especially recommended for decalcification of teeth, since it weakly affects staining (Murayama, 1937; Richman, Gelfand and Hill. 1946) [Pierce E. Histochemistry. Theoretical and applied. M., 1962; Lloyd Z., Gossrau R., Schiebler T. Histochemistry of enzymes. Laboratory methods. M., "World" - 1982].

However, these methods give mainly positive results on the subject of staining of bone tissue structures with various general histological methods. Thus, the Schmorl method is the most optimal color for sections of decalcified teeth: the sections are stained with a thionine solution for 10-12 minutes, then rinsed with water and transferred to a concentrated aqueous solution of picric acid for 1.5-2 minutes. Rinse again in distilled water, then differentiate in 70% alcohol until the separation of the dye from the sections has ceased (usually 5-10 minutes), dehydrate in 96% alcohol, carbol-xylene and mount on a glass slide with a balsam. As a result, odontoblasts with processes and dentinal tubules are stained. [R.P. Samusev / Fundamentals of the clinical morphology of teeth // Samusev R.P., Dmitrienko S.V., Krajushkin A.I. - M .: Onix 21 Century Publishing House LLC: Mir and Education LLC, 2002. - 268 p. (S. 30)].

As a method for detecting nerve formations, this method of fixation is of little use because of the high concentration of acids that make up the fixing fluids, which dissolve the myelinated and non-myelinated components of nerve elements. Thus, according to the method of Mirayama (1937), it is necessary to use concentrated formic acid, diluting in equal amounts of 70 ° alcohol, and according to the method of Richman, Gelfand and Khil (1946), 100 ml of formic and 80 ml of hydrochloric acid per 1 liter of distilled water are used [Pierce E Histochemistry. Theoretical and applied. M., 1962; Lloyd Z., Gossrau R., Schiebler T. Histochemistry of enzymes. Laboratory methods. M., "World" - 1982].

These methods are suitable only for general histological staining, but not for the detection of nerve structures due to the side effects of hydrochloric acid.

According to the deCastro method, nitric acid is used as fixation and decalcification [K.G. Tayushev Guide to Laboratory Neurohistological Technique / Tayushev K.G. - St. Petersburg, 2005. - S. 68]. According to this technique, the following is given:

1. Fixation and simultaneous decalcification: absolute ethanol - 50 ml, chloral hydrate - 5.0 g, distilled water 50 ml, nitric acid - 2-4 ml (depending on bone density from 4 to 10 days). If the tissue is very dense, the amount of nitric acid can be increased from 4 to 7 ml, the end of fixation is determined by the color of the object: it becomes translucent, milky.

2. Rinsing in running water - 1 day.

3. 96% ethanol - 50 ml, ammonia - 1 day.

4. 1.5% silver nitrate solution (you can add pyridine 1 drop per 1 ml of solution) - 7-10 days. In the thermostat at a temperature of 37 ° C.

5. Recovery: pyrogallol - 1 ml, neutral formalin - 10 ml, distilled water - 90 ml per day.

6. Fast passage through alcohols, wiring and pouring into celloidin.

7. Preparation of sections and histological preparations.

However, this method is difficult to fully identify the nerve structures in the bone tissue of the jaw, especially in the teeth.

The technical result of the invention is to determine the optimal concentration of decalcifying-fixing mixture to more fully identify the structure of the nervous system in the dentofacial system of humans and animals.

The technical result is achieved by the composition and concentration of the fixing mixture based on formic acid and chloral hydrate, and the concentration of silver nitrate.

The proposed method is as follows.

An object of dental or jaw tissue is fixed for 3-5 days in a mixture of the following composition: concentrated formic acid - 3.5 ml, chloral hydrate - 3.5 g, 96% ethyl alcohol - 50 ml, distilled water - 100 ml. The mixture is changed during the day 2-3 times.

Next, the object is washed in tap water for 24 hours and placed in a mixture of the following composition: 50 ml of 96 ° alcohol with ammonia (50 ml of alcohol and 5-6 drops of ammonia alcohol) for 30-60 minutes, depending on the size of the object.

Then the object is washed in distilled water for 20-30 minutes and impregnated in a 5% solution of silver nitrate in a thermostat at a temperature of 37 ° for 5-7 days, until the pieces acquire a light brown tint.

After the thermostat, the object is washed in distilled water for 10 minutes, followed by recovery for 24 hours in a mixture of composition: pyrogallic acid - 1 g, formalin - 10 ml, distilled water - 90 ml.

Next, washing with distilled water for 10-15 minutes and traditional wiring through alcohols, xylene, pouring into celloidin, cutting on a microtome and embalming are carried out.

The use of formic acid as a decalcifying component in the composition of the fixing mixture against the background of chloral hydrate was a new solution that facilitates the simplification of the procedure for conducting the test object through neurochemical reagents, improving the quality of the resulting neurohistological preparations, certain segments of the dentofacial system at the light-optical level.

Claims (1)

A method for detecting nerve structures in the dentofacial system, including fixing the test object with washing for 24 hours in running water, placing it in a mixture of ethanol with ammonia, characterized in that the fixation of the object is carried out for 3-5 days in a solution containing concentrated formic acid - 3.5 ml , chloral hydrate - 3.5 g, distilled water - 100 ml, moreover, the fixing solution is replaced 2-3 times a day and the washed test object, depending on its size, is placed in a mixture of alcohol with ammonia for 30-60 minutes with further washing m in distilled water for 20-30 minutes, and the impregnation is carried out in a thermostat for 5-7 days in a 5% solution of silver nitrate until the object acquires a light brown color, washing it in distilled water 10 minutes before recovery and 10-15 minutes before wiring, pouring , sharp and embalming.