RU1783435C - Method for diagnosis of pulmonary tuberculosis activity - Google Patents

Abstract

The invention relates to medicine, namely to TB and immunology. The purpose of the invention is to improve the accuracy of the method. Blood is taken from the patient, neutrophils are isolated, incubated in the presence of latex, followed by separation of the supernatant and incubation of donor neutrophils with it, the activity and intensity of phagocytosis are calculated, and if these values are below 1, the active form of tuberculosis is diagnosed. Compared to the prototype, accuracy is increased by 56%.

Description

The invention relates to medicine, in particular to phthisiology and immunology, and is intended to diagnose the activity of pulmonary tuberculosis.

A known method for the diagnosis of tuberculosis, including blood sampling, isolation of neutrophils, incubating them with latex, followed by determination of the activity (AF) and intensity (IF) of phagocytosis. Moreover, if the values of the obtained indicators are below normal, the active form of pulmonary tuberculosis III is diagnosed.

A disadvantage of the known method is its low accuracy.

The purpose of the invention is to improve the accuracy of the diagnostic method.

In order to achieve this goal, a pure population of patient's blood trophils is cultured in the presence of polystyrene monodisperse latex with a diameter of 1.7 μm, followed by isolation of the activated neutrophil supernatant. In addition, donor monocytes are isolated, to which a supernatant with a suspension of latex particles in a ratio of 1:20 is added, followed by determination of the activity (AF) and intensity of phagocytosis of monocytes in relation to latex particles and calculation of the phagocytosis intensity index by the formulas:

1L1LF IF of donor monocytes and supernatant of IF of donor monocytes in medium 199

MDF - AF of donor monocytes in the supernatant AF of donor monocytes in medium 199

and with values of these indices less than 1, an active form of pulmonary tuberculosis is diagnosed,

The following examples illustrate the implementation of the proposed method.

PRI me R 1. Stepanov V, P., 27 years. He was admitted to the hospital to clarify the activity of the tuberculosis process in the lungs.

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Pathologists in the lungs were detected for the first time during a professional examination. Complaints of minor weakness. Decreased appetite. With percussion and auscultation, no local changes were noted. Multiple microbiological examinations using various methods of mycobacterium tuberculosis were not detected. On the survey roentgenogram of the lungs, tomograms of the left apex of 6-7 cm in the upper lobe of the left lung - single large, medium-intensity foci with relatively clear contours, homogeneous structure. Conclusion: focal tuberculosis of the upper lobe of the left lung, phase of infiltration.

Blood test: hemoglobin - 162, ESR - 4, l - 5.9 105, p - 3, C - 49, l - 39, m - 9.

The study was conducted in accordance with the proposed method. A pure neutrophil population was isolated from venous heparinized blood (25 units of heparin per 1 ml of blood) of a patient (15 ml) by centrifugation on a double density gradient ficoll-verographin (D1-1.077, Da-1,093) according to the Wong-Wilson method for 40 min at 1500 rpm A suspension of granulocytes from the ring formed at the interface between solutions was diluted with 199 medium to a concentration of 5-106 cells (neutrophils) in 1 ml and a suspension of particles of monodispersed polystyrene latex with a diameter of 1.7 m km was introduced into it at the rate of 50 particles per 1 - neutrophil, Suspension tube was placed in a thermostat at 37 ° C for 1 h to obtain latex activated neutrophil supernatant. The contents of the tube were centrifuged at 3000 rpm for 15 minutes to remove latex cells and particles.

Additionally donor monocytes were isolated, for which mononuclear cells isolated from peripheral blood were centrifuged on a double density gradient. The resulting ring of mononuclear cells was aspirated and removed twice by medium 199. Mononuclear cells at a concentration of 106 cells in 1 ml were incubated for 1 h at 37 ° С in test tubes, boots with coverslips at the bottom. Cells adhering to the glass were then washed from lymphocytes with 199 medium and used to study phagocytic activity as target cells for the action of neutrophil secretory products. The control was monocytes incubated in medium 199.

1 ml latex activated neutrophil supernatant (SAN) was added to the test boot tubes. After one-hour incubation, 0.1 ml of a suspension of standardized latex microspheres with a diameter of 1.7 µm in a ratio of 1:20 was added to the test and control tubes. Test tubes

placed for 1 h in a thermostat, after which the monolayer was washed three times with medium 199 and stained according to Romanovsky-Giemsa and the activity (AF) and intensity (IP) of latex phagocytosis by donor monocytes in experiment and control were determined: the number of phagocytic cells was calculated per monolayer, 100, which reflects the activity of phagocytosis, and the number of latex particles absorbed in 100 counted cells, the intensity of phagocytosis:

AF199 31, AFSAN 16, IF199 127, IFSAN 61.

The values of the indices of phagocytosis activity (IAF) and phagocytosis intensity (IIF) were determined:

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IIF, 48 IAF 4u 0.31

The values of both indicators are less than 1.0, which in accordance with the proposed method indicates active pulmonary tuberculosis.

Observation of the patient subsequently confirmed the correctness of the diagnosis.

Example 2. Gabdurakhmanov Z. G., 36 years.

Entered for further examination and treatment. Pathologists in the lungs were detected fluorographically during a physical examination. In the lungs: percussion-pulmonary sound, auscultation - vesicular breathing, no wheezing. Radiographically in the survey photographs, tomograms of the right apex 6-7-8 cm. On the right in the upper lobe there are a few numerical foci of medium intensity, clear contours of small and medium size. On the left, in the 11-11 intercostal space, small foci of medium intensity.

In the blood test at admission: heme. 175, ESR - 13, L - 4.0, e - 2, p - 1, s - 53, l - 35, m - 2. Urine analysis without pathology. Mantoux reaction - 7 mm. In the analysis of sputum by the luminescent method and plating repeatedly BK was not detected.

In accordance with the proposed method AF monocyte donor with SAN patient

amounted to 29, IF - 107; AF of donor monocytes in medium 199-41; IF of donor monocytes in medium 199-132.

IAF, 7 IIF .8

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The values of both indicators are less than 1.0, which made it possible to diagnose the active tuberculosis process in the patient's lungs.

Subsequently, after 2.5 months the diagnosis was clarified for CVCK: focal tuberculosis in / lobe of the right lung, infiltration phase. VK-1 group of dispensary registration.

The accuracy of the proposed method is 96% versus 60% according to the prototype method.

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Claims (1)

SUMMARY OF THE INVENTION A method for diagnosing the activity of pulmonary tuberculosis, including blood sampling, blood cell isolation, incubation in the presence of latex, calculation of indicators of activity and phagocinosis intensity, characterized in that, in order to improve the accuracy of the method, supernatant is isolated after blood cells are incubated, incubated with it, human donor monocytes with the subsequent calculation of indicators, activity and intensity of phagocytosis and with their value below 1, diagnose the active form of tuberculosis.