Automated analysis of immunocompetent ce lls revealing their subpopulation composition , investigation of cell ensembles and/or other micro-objects with fluorescent labels .

The goal of the analysis is to get statistic distributions of cell individual parameters in two fluorescence spectral ranges simultaneously.

The analytical results obtained with the instrument are similar to results of flow cytometry experiments .

•  Сell suspension will be put into the hemocytometer (count chamber) in the field-of-view (FOV) of a microscope objective and two video cameras. A great number of FOV-images are automatically recorded and processed in series at the rate about 1 image per second. The FOV's are changed by computer control of a microscopic stage.

•  The sample is analyzed in two fluorescence spectral ranges simultaneously by using replaceable band-pass filters for dyes FITC , PE , PI. Digital spectral compensation of fluorescence is provided.

•  Automatic cell detection and evaluation of their parameters are performed in real-time mode by using image recognition algorithms in processing and analyzing the FOV-images (Image Cytometry method).

•  The analysis results are presented as histograms and two-parameter dot plots.

•  The software allows for data gating (selecting arbitrary areas of the plots for analysis), superposition and combination the results, identification the gated data directly on the sample images, and raw data (images) storage for off-line analyses.

Transmitted light measurements: Size : average cell diameter in m m Transparency: integral transparency of a cell as compared with the buffer solution.   Laser measurements (473 nm): Fluorescence intensity of each cell in two spectral ranges. The instrument is equipped with a set of band-pass filters: 532 nm, 576 nm, 673 nm.

Four parameters are determined for each cell individually.

Laboratory clinical diagnostics in immunology, allergology and oncology including immunophenotyping; medical and biological investigations using fluorescent labels.

RESULTS OF LYMPHOCYTE ANALYSIS WITH ДСКФ-01

Determination of T and B lymphocytes relative contents in peripheral blood of a conditionally healthy adult donor by using the ДСКФ-01 instrument. Fluorescence intensity FL1 corresponds to label CD3;
FL2 corresponds to CD19.

•  The instrument is free of cell flow-through transportation systems, as well as of laser beam focusing and scanning systems. This makes the instrument compact and easy in operation, and also decreases significantly its cost.

•  The instrument operation philosophy is based on Image Cytometry. Detection of cells and measurement of their individual parameters are performed by using on-line image processing and analysis via the image recognition algorithms.

•  It is possible to visualize the sample under study in the digital microscope mode (1 µm resolution).

•  It is possible to observe the sample parameters variations in time.

•  It is possible to re-analyze the data of experiments performed earlier, which are stored in the raw image archive. The analysis results, as well as the raw data for experiments of the same type, may be combined in order to improve statistical representativeness.

Developer: Institute for Analytical Instrumentation RAS.

Methodological principles of the instrument application are being developed jointly with State Research Center "Russian FMBA Immunology Institute".