Strategies for Rare-Event Detection: An Approach for Automated Fetal Cell Detection in Maternal Blood

Abstract

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.

Automated detection of rare‐event pathogens through time‐gated luminescence scanning microscopy

“…These include both flow cytometry-based techniques and solid-phase image scanning cytometry techniques (3,4,(6)(7)(8). The latter usually uses a planar multielement photodetector, such as a charge-coupled device (CCD) camera, to make images from adjacent elements across the specimen, followed by scene segmentation and use of pattern recognition algorithms to locate the particular targets (9)(10)(11)(12)(13)(14)(15). Although it has the potential to detect rare events in an automatic way, presently, there exist several restrictions that limit its practical application.…”

Prenatal Diagnosis of Genetic Abnormalities Using Fetal CD34+Stem Cells in Maternal Circulation and Evidence They Do Not Affect Diagnosis in Later Pregnancies

“…1-13), who had never been pregnant (Nos. [14][15][16][17][18][19][20][21][22][23][24][25], and who were currently pregnant (Nos. [26][27][28][29][30][31][32][33].…”

“…Many strategies have been designed to improve the recovery, purity, and yield of these fetal cells from maternal blood, but the success rate varies considerably [11][12][13][14][15][16][17][18]. The major limitations to most of these techniques are the few fetal cells which can be sorted from maternal blood samples [19][20][21][22] and the lack of a specific fetal cell marker [23][24][25].…”

Applicability of Four New Antibodies for the Detection of Fetal Nucleated Cells Out of Maternal Blood by FISH Analysis

“…Various attempts for this are in progress [28][29][30]. Furthermore, until the optimal fetal specific antibody has been discovered, it will remain necessary to improve upon the current enrichment methods by examining different density gradients and MACS or FACS [18,31], because these methods are associated with the loss of cells of maternal as well as of fetal origin.…”