RU2708241C1 - Oral fluid examination method - Google Patents

Abstract

FIELD: monitoring and measuring equipment.

SUBSTANCE: invention refers to oral fluid examination. Oral fluid examination method involves collecting and centrifuging, preparation of the preparation and its further study. At that, except saliva glands secretion microflora and products of their vital activity are selected, content of periodontal pockets, gingival fluid, desquamated epithelium, decay products of leukocytes migrating into oral cavity, for which purpose in the process of extraction interdental gaps are washed, periodontal pockets by means of an oral irrigation agent using corresponding nozzles for cleaning and washing of oral cavity objects, all produced fluid from one patient is discharged into a separate sterile container, centrifuged for 30 seconds at speeds 600 rpm, sediment is separated and placed in even thin layer on slide and immediately studied by phase-contrast microscopy.

EFFECT: invention provides the possibility of studying a histologic preparation with an enriched cytological picture.

1 cl, 12 dwg

Description

The invention relates to the field of dentistry, and more specifically to methods for examining oral fluid and can be used to make a histological preparation from it and its subsequent study with a picture.

A known method for examining oral fluid, including collecting it using collectors containing a cotton swab from dental (surgical) cotton. A cotton swab is placed under the tongue for 10 minutes without stimulation of salivation. After the swab is soaked with saliva, it is placed in a collector closed by a hermetically sealed plastic stopper, the collector is sealed and sent with accompanying documentation to the laboratory in a cooler bag (see Order of the Ministry of Health and Social Development of the Russian Federation of January 27, 2006 No. 40 “On the Organization of Chemical and Toxicological research in the analytical diagnosis of the presence in the human body of alcohol, drugs, psychotropic and other toxic substances ”).

The disadvantage of this technical solution is that the amount of fluid is limited, single cells are identified in the smears, so the method is not very informative, and does not give an idea of the state of the oral cavity.

There is also known a method for studying oral fluid, including its collection and centrifugation, preparation of the drug and its subsequent study (see RU No. 2432620, IPC G09B 23/28, 2010). Fluid intake is carried out in the morning, on an empty stomach, before brushing your teeth. Centrifuged for 10 minutes at a speed of 3000 rpm. To determine the physicochemical parameters of the oral fluid, we used methods adapted in dentistry to study the oral fluid (Pitaeva A.N., Korshunov A.P., Distel V.A. et al. “Physicochemical methods for the study of mixed saliva in clinical and experimental dentistry ”(study guide) - Omsk, 2001).

As a result, in the preparation, the cells in the field of view are single, i.e. drugs are uninformative. Thus, the disadvantage of this method is that it is not possible to form a histological preparation from the oral fluid with an enriched cytological picture.

The problem to which the invention is directed is expressed in providing the opportunity to study a histological preparation with an enriched cytological picture formed from oral fluid.

The technical result, which is manifested in the solution of the problem, is expressed in providing the possibility of studying a histological preparation with an enriched cytological picture formed from oral fluid.

To solve this problem, a method for studying oral fluid, including its collection and centrifugation, preparation of the drug and its subsequent study, differs in that in addition to the secretion of the salivary glands, microflora and products of their vital activity, the contents of periodontal pockets, gingival fluid, desquamated epithelium, and decay products of migrants are selected into the oral cavity of leukocytes, for which, during the selection process, interdental spaces, periodontal pockets are washed with an oral irrigator using the appropriate nozzles for cleaning and rinsing the objects of the oral cavity, while all the liquid obtained from one patient is taken into a separate sterile container, then centrifuged for 30 seconds at a speed of 600 rpm, the precipitate is separated and placed with a smooth thin layer on a glass slide, and immediately examined using phase contrast microscopy.

In addition, if there are orthodontic structures and / or artificial crowns and / or bridges and / or implants, appropriate nozzles are used to clean the oral fluid during the selection process.

A comparative analysis of the features of the claimed solution with the signs of the prototype and analogues indicates the conformity of the claimed solution to the criterion of "novelty."

The combination of features of the claims provides the possibility of forming a histological preparation with an enriched cytological picture from the oral fluid, while the features of the characterizing part of the claims solve the following functional tasks.

The signs "... in addition to the secret of the salivary glands, the microflora and their metabolic products, the contents of periodontal pockets, gingival fluid, desquamated epithelium, and the breakdown products of leukocytes migrating into the oral cavity are selected ..." provide the enrichment of the oral fluid with additional components that, if necessary, expand the set of diagnostic signs, not only regarding the oral cavity, but also other body systems.

The signs "... during the selection process, they wash interdental spaces, periodontal pockets by means of an oral irrigator using appropriate nozzles for cleaning and washing objects of the oral cavity ..." provide washing with a thin stream of water under pressure and cleaning material from the teeth and other objects of the oral cavity into the oral fluid and thus enrichment of the oral fluid.

The sign "... all the fluid received from one patient is taken to a separate sterile container ..." provides the "binding" of the selected material of the oral cavity to a specific patient, which allows us to study the dynamics of changes in the composition of the oral fluid.

A sign indicating that the material of the oral fluid is “centrifuged for 30 seconds at a speed of 600 rpm” ensures the preservation of the cell structure in the test material. The sediment under such centrifugation conditions is represented only by living cells, which are observed in large quantities in the smear. The morphology of cells corresponds to their real intravital structure in the oral cavity.

Signs indicating that "the precipitate is separated and placed in an even thin layer on a glass slide", provide preparation for the study.

Signs indicating that the prepared preparation is “immediately studied using phase contrast microscopy” provides the possibility of using the obtained material to study the state of the oral cavity and analyze the dynamics of the reaction of the oral mucosa to the installation of braces, implants, dentures of various chemical composition or treatment, which expands the range of diagnostic possibilities and provides timely prevention of rejection reactions or allergic manifestations.

Signs indicating that "in the presence of orthodontic structures and / or artificial crowns and / or bridges and / or implants, appropriate nozzles are used to clean the oral fluid during the selection of the oral fluid," they increase the efficiency of cleaning the oral cavity and the concentration of the material from the surface of the oral cavities in the oral fluid.

In Fig.1 - Fig.4 shows the steps of collecting oral fluid, while in Fig.1 the installation procedure of the retaining holder; figure 2 shows the washing and cleaning of the oral cavity; figure 3 shows the selection of oral material from the oral cavity; figure 4 shows the placement of the material in a test tube, for subsequent centrifugation; figure 5 - figure 6 shows a scraping from the mucous membrane of the mouth of the patient of the study group with post-traumatic defects of the jaws, while figure 5 shows the drug without fixation; 6 shows phase contrast after fixation and centrifugation; in FIG. 7 - Fig. 12 shows a scraping from the oral mucosa, while Fig. 7 shows mitosis of the epithelial cell; on Fig and 9 shows dendritic cells; 10 and 11 show macrophages; 12 shows an epithelial cell.

To implement the method, known means are used: an oral irrigator (an apparatus forming a thin stream of water, under pressure), nozzles for cleaning the tongue, for washing periodontal pockets, nozzles for cleaning orthodontic structures (braces), nozzles for cleaning artificial crowns and bridges, nozzles for cleaning implants. In addition, other well-known products are used: spatula, saliva ejector, scraper, sterile tubes, jaw retainer. In addition, a research centrifuge, for example, K1015, is used for soft centrifugation. A microscope, for example, an Olympus Bx51 with a DP25 camera, is used to study drugs and make illustrations.

Stage 1 - collection of oral fluid. The technique for collecting material for research from the oral cavity involves:

- installation of a retaining holder, ensuring not closing the jaws (figure 1);

- washing the oral cavity and objects in it by means of an oral irrigator, as well as cleaning objects in the oral cavity using the corresponding, aforementioned nozzles (figure 2);

- collection of material from the oral cavity by means of saliva ejector (figure 3), also, for the collection of material, we use a spatula and a scraper;

- placing the material in a test tube (figure 4).

The material is taken from the oral cavity using an oral irrigator, forming a thin stream of water, which under pressure rinses the interdental spaces from plaque, as well as periodontal pockets, tongue, orthodontic structures (braces), artificial crowns, bridges are cleaned (if any). implants.

The material is centrifuged at low speeds, without destroying the cells - according to our data, the cell structure is preserved in the studied material at a speed of 600 rpm for 30 seconds.

All of the liquid obtained is centrifuged under the mentioned conditions and, thus, an enriched concentrate consisting of epithelial cells, leukocytes and microorganisms is obtained in the sediment. The sediment under such centrifugation conditions is represented only by living cells, which are observed in large quantities in the smear. The morphology of the cells corresponds to their real intravital structure in the oral cavity, since the integrity of the membrane and organelles in the cytoplasm, which form subcellular fractions at high speeds, is maintained.

The obtained enriched precipitate, consisting of epithelial cells and leukocytes, as well as microorganisms, is applied to a glass slide and a spatula, a cover glass, or a glass slide, the material is distributed on a glass slide and immediately examined using phase contrast microscopy. The number of cells in the field of view exceeds that of conventional smear production. For making illustrations, use an Olympus Bx51 microscope with a DP25 camera. The microscopic technique involves the analysis of the drug without a coverslip with preliminary drying and application of immersion oil to work with the highest magnification under the immersion lens. In the objects under consideration, nuclei, mitosis figures, cell processes, cytoplasm contents, including vesicles and contaminated organisms, are identified.

Comments on photographs (microphoto. UV. X400.1): Fig. 5 shows a living cell, microorganisms are identified in the cytoplasm; Fig.6 in the field of view there is a greater number of cells, identified nuclear epithelial cells with contamination by microorganisms; Fig.7 shows mitosis of epithelial cell; on Fig and 9 shows dendritic cells; 10 and 11 show macrophages; 12 shows an epithelial cell contaminated with microorganisms. The presence of contaminated microorganisms makes it possible to evaluate the infectious process, the level of microbial seeding, the barrier properties of the epithelial plates of the oral mucosa and the cellular composition. The number of desquamated epithelial cells with microbes gives an idea of the protective properties of the epithelium. Migrant cells and their number can give an idea of the level of protective reactions and, in large quantities, indicate the possibility of implant rejection.

The advantages of the method:

- has a low cost, is available to any clinic with any budget;

- informative, gives a complete picture of the state of the oral cavity;

- based on the data of the sediment content, it is possible to predict the outcome of the pathological process and timely influence key pathogenesis events and prevent complications;

- from the moment of sampling the material for diagnosis to the results of the analysis, time is spent no more than 10 minutes;

- ease of implementation does not require special training and acquisition of skills of diagnostic manipulations;

- enables multiple sampling of diagnostic material;

- non-invasiveness and injury safety for both the patient and the doctor (excludes infection through the blood of HIV and hepatitis C), as well as other socially significant infections.

Claims (2)

1. A method for examining the oral fluid, including its collection and centrifugation, preparation of the drug and its subsequent study, characterized in that in addition to the secretion of the salivary glands, microflora and products of their vital activity, the contents of periodontal pockets, gingival fluid, desquamated epithelium, and decay products migrating into the cavity are selected leukocyte mouth, for which, during the selection process, interdental spaces, periodontal pockets are washed with an oral irrigator using appropriate nozzles for cleaning and rinsing objects of the oral cavity, while all the liquid obtained from one patient is taken into a separate sterile container, then centrifuged for 30 seconds at a speed of 600 rpm, the precipitate is separated and placed with an even thin layer on a glass slide and immediately examined using phase contrast microscopy. 2. The method according to claim 1, characterized in that in the presence of orthodontic structures and / or artificial crowns and / or bridges and / or implants in the process of selecting oral fluid, appropriate nozzles are used to clean them.

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