Objective: To evaluate the reliability and applicability of the spot-counting system (Cytovision Spot AX workstation) which offers an alternative to the tedious manual analysis of sperm fluorescence in situ hybridization (FISH).
Design: Manual and automatic analyses were performed and compared.
Setting: Universitat Autònoma de Barcelona.
Patient(s): Twenty-four men who asked for information on infertility showing different seminal parameters.
Intervention(s): A semen sample for each patient was collected and prepared for FISH.
Main Outcome Measure(s): A dual-color FISH using specific probes for chromosomes 13 and 21 and a triple-color FISH with centromeric probes for chromosomes 18, X, and Y were used (Vysis). Standard FISH analysis was carried out. Automatic analysis was subsequently performed using a Spot AX system.
Result(s): Overall, we performed 120 comparisons. In 116 out of 120 (96.67%), the percentage of anomalies reported using manual counting fell within the incidence detected using the automatic system. In the remaining comparisons, statistical differences were detected (4 out of 120; 3.33%). Time consumed by the automatic analysis was always higher than the manual one, being influenced by the characteristics of the preparations.
Conclusion(s): The spot-counting system has potential, but before the service is ready to be offered, we still need to overcome some limitations associated with it.
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http://dx.doi.org/10.1016/j.fertnstert.2008.07.1779 | DOI Listing |
J Hum Genet
July 2023
Division of Hematology & Oncology, Department of Medicine, Kyoto Prefectural University of Medicine, Kyoto, Japan.
Three types of chromosomal translocations, t(4;14)(p16;q32), t(14;16)(q32;q23), and t(11;14)(q13;q32), are associated with prognosis and the decision making of therapeutic strategy for multiple myeloma (MM). In this study, we developed a new diagnostic modality of the multiplex FISH in immunophenotyped cells in suspension (Immunophenotyped-Suspension-Multiplex (ISM)-FISH). For the ISM-FISH, we first subject cells in suspension to the immunostaining by anti-CD138 antibody and, then, to the hybridization with four different FISH probes for genes of IGH, FGFR3, MAF, and CCND1 tagged by different fluorescence in suspension.
View Article and Find Full Text PDFMethods Mol Biol
March 2019
Cellular Technology Ltd., Shaker Heights, OH, USA.
Over the past decade, ELISPOT has become a highly implemented mainstream assay in immunological research, immune monitoring, and vaccine development. Unique single cell resolution along with high throughput potential sets ELISPOT apart from flow cytometry, ELISA, microarray- and bead-based multiplex assays. The necessity to unambiguously identify individual T and B cells that do, or do not co-express certain analytes, including polyfunctional cytokine producing T cells has stimulated the development of multi-color ELISPOT assays.
View Article and Find Full Text PDFMethodsX
September 2017
Weldon School of Biomedical Engineering, Purdue University, United States.
Advanced molecular probing techniques such as single molecule fluorescence in situ hybridization (smFISH) or RNAscope can be used to assess the quantity and spatial location of mRNA transcripts within cells. Quantifying mRNA expression in large image sets usually involves automated counting of fluorescent spots. Though conventional spot counting algorithms may suffice, they often lack high-throughput capacity and accuracy in cases of crowded signal or excessive noise.
View Article and Find Full Text PDFMech Dev
December 2017
Stowers Institute for Medical Research, 1000 E 50th St, Kansas City, MO 64110, USA; Department of Anatomy and Cell Biology, University of Kansas School of Medicine, Kansas City, Kansas 66160, USA. Electronic address:
During collective cell migration individual cells display diverse behaviors that complicate our understanding of group cell decisions of direction and cohesion. In vivo gene and protein expression analyses would shed light on the underlying molecular choreography. However, this information has been limited due to difficulties to integrate single cell detection methods and the simultaneous readout of multiple signals deep within the embryo.
View Article and Find Full Text PDFInt J Mol Sci
July 2013
Department of Cell Biology, Physiology and Immunology, Universitat Autònoma de Barcelona, 08193 Bellaterra, Spain.
Phosphorylation of the H2AX protein is an early step in the double strand break (DSB) repair pathway; therefore, phosphorylated histone (γH2AX) foci scoring is widely used as a measure for DSBs. Foci scoring is performed either manually or semi-automatically using hand-operated capturing and image analysis software. In general, both techniques are laborious and prone to artifacts associated with manual scoring.
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