METHOD FOR ANALYZING ORAL FLUID PREPARATION Russian patent published in 2024 - IPC G01N33/569 G01N33/577 G01N1/30 A61B5/00 

FIELD: medicine.

SUBSTANCE: invention relates to medicine, namely to dentistry, immunology and can be used for visualization of neutrophil extracellular traps in oral fluid. Oral fluid is collected in amount of 1–2 ml, after which it is diluted with a phosphate buffer solution at pH = 7.4 at ratio of 1:1. 2 parts of the diluted oral fluid are transferred onto a slide, 1 part of propidium iodide in concentration of 0.1 mg/ml and 1 part of monoclonal antibodies to CD45, labelled with FITC, preliminarily diluted with phosphate buffer solution at pH = 7.4 at ratio of 1:10 and incubated under a cover glass in dark for 5 minutes at 37 °C. Then the preparation is visualized by means of luminescent microscopy, using light filters: excitation - with wavelength range of 450–480 nm and emission - in the wavelength range of 515–700 nm. Preparation is examined for the presence, with the determination of the percentage: neutrophilic extracellular traps of the type of early netosis; filamentous netosis; cloud-like netosis; leukocytes by type: intact leukocytes without stained nucleus; monocytes without stained nucleus; weakly activated neutrophils; activated neutrophils; hyperactive neutrophils; epithelial cells by type: intact epithelial cells; epitheliocytes with propidium-positive nucleus and propidium-negative cytoplasm; epitheliocytes with propidium-positive nucleus and cytoplasm; layers of epithelial cells. Percentage of epithelial cells with signs of adhesion of microorganisms, epithelial cells without signs of adhesion of microorganisms, microorganisms, free colonies of microorganisms is estimated.

EFFECT: invention provides simultaneous visualization in the oral fluid sample of neutrophil traps of different types with the possibility of counting bacteria captured by formed NLV, intact viable leukocytes, as well as neutrophils in different degrees of activation in the process of a non-forming reaction, visualization, identification and calculation of the percentage of monocytes, visualization of epithelial cells and identification of their morphological equivalents from intact forms to non-viable and counting epithelial cells with signs of adhesion of microorganisms and without signs of adhesion.

1 cl, 5 dwg, 3 ex