Use of automated microscopy for the detection of disseminated tumor cells in bone marrow samples.

The use of automated microscopy has reached the maturity necessary for its routine use in the clinical pathology laboratory. In the following study we compared the performance of an automated microscope system (MDS) with manual method for the detection and analysis of disseminated tumor cells present in bone marrow preparations from breast carcinoma patients. The MDS System detected rare disseminated tumor cells among bone marrow mononuclear cells with higher sensitivity than standard manual microscopy. Automated microscopy also proved to be a method of high reproducibility and precision, the advantage of which was clearly illustrated by problems of variability in manual screening. Accumulated results from two pathologists who had screened 120 clinical slides from breast cancer patients both by manual microscopy and by use of the MDS System revealed only two (3.8%) missed by the automatic procedure, whereas as many as 20 out of 52 positive samples (38%) were missed by manual screening.

Enrichment, identification and analysis of fetal cells from maternal blood: evaluation of a prenatal diagnosis system.

In this study we evaluated the performance of a system for the enrichment, identification and analysis of fetal cells in maternal peripheral blood. Blood samples were collected from women after chorionic villus sampling and enriched for the presence of nucleated erythrocytes using a three-step procedure, namely: (a) centrifugation to separate nucleated red blood cells (NRBCs) from the majority of red blood cells (RBCs) and white blood cells (WBCs); (b) selective lysis of the remaining maternal RBCs; (c) separating the NRBCs from the remaining WBCs in a three-layer density gradient. Fetal cells were identified by using a monoclonal antibody against the gamma-chain of fetal haemoglobin (anti-HbF) and a nuclear stain (DAPI). Additionally, to further increase the specificity of the identification, and to eliminate some of the undesired staining by maternal leukocytes, a fluorescent antibody (CD45) was added. The sex chromosome complement of the cells was determined by fluorescence in situ hybridization (FISH) with X and Y-specific probes and the results were compared with the karyotypes obtained after analysis of chorionic villi. Using the described method, in all cases where the woman was carrying a male fetus (n=18) at least one XY cell was found, while no male cells were found in women carrying a female fetus. However, in the majority of cases with a male fetus (n=11) female HbF positive cells were found indicating the presence of maternal nucleated erythrocytes. The study demonstrates that the combination of anti-HbF and CD45 is a useful, but not fully specific, marker for fetal NRBCs and that additional markers are needed.

Strategies for rare-event detection: an approach for automated fetal cell detection in maternal blood.

This article explores the feasibility of the use of automated microscopy and image analysis to detect the presence of rare fetal nucleated red blood cells (NRBCs) circulating in maternal blood. The rationales for enrichment and for automated image analysis for "rare-event" detection are reviewed. We also describe the application of automated image analysis to 42 maternal blood samples, using a protocol consisting of one-step enrichment followed by immunocytochemical staining for fetal hemoglobin (HbF) and FISH for X- and Y-chromosomal sequences. Automated image analysis consisted of multimode microscopy and subsequent visual evaluation of image memories containing the selected objects. The FISH results were compared with the results of conventional karyotyping of the chorionic villi. By use of manual screening, 43% of the slides were found to be positive (>=1 NRBC), with a mean number of 11 NRBCs (range 1-40). By automated microscopy, 52% were positive, with on average 17 NRBCs (range 1-111). There was a good correlation between both manual and automated screening, but the NRBC yield from automated image analysis was found to be superior to that from manual screening (P=.0443), particularly when the NRBC count was >15. Seven (64%) of 11 XY fetuses were correctly diagnosed by FISH analysis of automatically detected cells, and all discrepancies were restricted to the lower cell-count range. We believe that automated microscopy and image analysis reduce the screening workload, are more sensitive than manual evaluation, and can be used to detect rare HbF-containing NRBCs in maternal blood.

[An automated unit for manual photometry and morphometry]. / Avtomatizirovannaia ustanovka dlia ruchnoi fotometrii i morfometrii.

An automated unit for manually controlled photometry and morphometry is described. It enables the operator to carry out photometrical measurements according to the plug-method and morphometrical measurements with the help of an object-micrometer or some kind of a grid. The data obtained are stored in the "Elektronika D3-28" microcomputer to be statistically evaluated after the end of sampling. New variables can be obtained defined by the user as a product of some source variables or their inverses. The data and control structures of the system are described in detail.

Morpho-functional principle of neuroendocrine system analysis.

A new approach to the analysis of the neuroendocrine system (NES) is suggested. It is based on the fact of structural and metabolic determination of any effect on cell and cell aggregates. The principle of a common communication channel in the NES is formulated and a possible method of its formalization is proposed.

[Modeling the cell cycle of complex populations]. / Modelirovanie kletochnogo tsikla slozhnykh populiatsii.

A model of cell cycle kinetics of complex populations is developed, by using systems of difference equations, and simulated on the computer. This model provides theoretical curves of labeled mitoses (PLM) percentage, labeling index and grain count histograms for different time intervals after 3H-thymidine administration. The PLM curve corresponding to a mixed population is a weighed sum of PLMs for populations, comprising the mixture, the weights being determined by parameters of populations. The model appeared to give a good agreement with the PLM curves obtained elsewhere.