Telomere length measurements in leukocyte subsets by automated multicolor flow‐FISH

Baerlocher, Gabriela M.; Lansdorp, Peter M.

doi:10.1002/cyto.a.10064

Cited by 100 publications

(90 citation statements)

References 24 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…An example for telomere analysis utilizing cell suspensions is the use of flow-FISH (60)(61)(62) or high throughput quantitative FISH (HT Q-FISH) (63), allowing for a high throughput of samples. However, these methods are less sensitive (kB-range) than Q-FISH.…”

Section: Discussionmentioning

confidence: 99%

Novel automated three‐dimensional genome scanning based on the nuclear architecture of telomeres

Klewes

Höbsch

Katzir³

et al. 2010

Cytometry Pt A

View full text Add to dashboard Cite

Telomeres, the end of chromosomes, are organized in a nonoverlapping fashion and form microterritories in nuclei of normal cells. Previous studies have shown that normal and tumor cell nuclei differ in their 3D telomeric organization. The differences include a change in the spatial organization of the telomeres, in telomere numbers and sizes and in the presence of telomeric aggregates. Previous attempts to identify the above parameters of 3D telomere organization were semi-automated. Here we describe the automation of 3D scanning for telomere signatures in interphase nuclei based on three-dimensional fluorescent in situ hybridization (3D-FISH) and, for the first time, define its sensitivity in tumor cell detection. The data were acquired with a highthroughput scanning/acquisition system that allows to measure cells and acquire 3D images of nuclei at high resolution with 403 or 603 oil and at a speed of 10,000-15,000 cells h 21 , depending on the cell density on the slides. The automated scanning, TeloScan, is suitable for large series of samples and sample sizes. We define the sensitivity of this automation for tumor cell detection. The data output includes 3D telomere positions, numbers of telomeric aggregates, telomere numbers, and telomere signal intensities. We were able to detect one aberrant cell in 1,000 normal cells. In conclusions, we are able to detect tumor cells based on 3D architectural profiles of the genome. This new tool could, in the future, assist in patient diagnosis, in the detection of minimal residual disease, in the analysis of treatment response and in treatment decisions. '

show abstract

Section: Discussionmentioning

confidence: 99%

Novel automated three‐dimensional genome scanning based on the nuclear architecture of telomeres

Klewes

Höbsch

Katzir³

et al. 2010

Cytometry Pt A

View full text Add to dashboard Cite

show abstract

“…Though there are certain specific FISH applications that have strong and consistent signals, such as telomeric length analysis or the detection of the presence or absence of a Y chromosome, FISH probe intensity variation can be high and signal intensities often approach the detection limits of standard flow cytometry, reducing the reliability of aneuploidy assessment. [27][28][29][30] Imaging flow cytometry is potentially well suited to FISH analysis because the detection limit of imaging flow cytometry improves as the size of the fluorescent signal source decreases. 31 Further, the quantitation of FISH-probed cells for applications such as aneuploidy analysis is accomplished by spot counting rather than relying exclusively on total intensity analysis, making it tolerant of wide variations in probe intensity and more consistent with the standard of practice in clinical FISH assessment.…”

Section: High Throughput Extended Depth Of Field Imaging Of Cells Submentioning

confidence: 99%

Cellular Image Analysis and Imaging by Flow Cytometry

Basiji

Ortyn

Liang

et al. 2007

Clinics in Laboratory Medicine

373

288

View full text Add to dashboard Cite

SynopsisImaging flow cytometry combines the statistical power and fluorescence sensitivity of standard flow cytometry with the spatial resolution and quantitative morphology of digital microscopy. The technique is a good fit for clinical applications by providing a convenient means for imaging and analyzing cells directly in bodily fluids. Examples are provided of the discrimination of cancerous from normal mammary epithelial cells and the high throughput quantitation of FISH probes in human peripheral blood mononuclear cells. The FISH application will be further enhanced by the integration of extended depth of field imaging technology with the current optical system. Keywordsimaging; flow; cytometry; fluorescence; brightfield; darkfield; multispectral Quantifying Cellular Structure in Health and DiseaseThe eukaryotic cell is a highly structured, three-dimensional object containing a wide range of molecular species. The size, shape, and structure of the cell, as well as the abundance, location, and co-location of any of these constituent biomolecules may be of significance in any given clinical situation or research application. For instance, in hematopoiesis, as cells differentiate and mature, different subsets of molecules are expressed that reflect a specialized functional capacity for that unique cell type (e.g., granulocytes vs. lymphocytes). In general the characterization of this array of constituent molecules by imaging or flow cytometry provides insight into the physiological function of any particular cell or alternatively, pathological changes that may have occurred or accrued. In clinical practice and in research settings, cellular evaluation by imaging technologies and flow cytometry provides significant information reflecting the particular cellular phenotype, both normal and pathological. Microscopy provides a wealth of information, but data acquisition rates are slow and analysis is generally subjective. In flow cytometry, data acquisition is rapid and better suited for the evaluation of pathologies present in low frequency, but the data are only intensity-based, thus lacking the morphology that truly lends credence to the analysis.In addition, the assessment and evaluation of cell samples by imaging and flow cytometric techniques is complicated by a number of factors. For instance, changes in a cell type or aCorresponding author for proofs and reprints:

show abstract

“…To assess telomere length in cells, flow-FISH analysis of cells hybridized with a telomere-specific FITC-conjugated peptide nucleic acid probe was performed as described (29,30). Mean telomere length was calculated as described (30) by using cow thymocytes as an internal standard.…”

Section: Measurement Of Telomerase Activity and Telomere Lengthmentioning

confidence: 99%

Dissociation of telomerase activity and telomere length maintenance in primitive human hematopoietic cells

Wang

Warner

Erdmann

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Primitive human hematopoietic cells have low endogenous telomerase activity, yet telomeres are not maintained. In contrast, ectopic telomerase expression in fibroblasts and other cells leads to telomere length maintenance or elongation. It is unclear whether this disparity can be attributed to telomerase level or stems from fundamentally different telomere biology. Here, we show that telomerase overexpression does not prevent proliferation-associated telomere shortening in human hematopoietic cells, pointing to the existence of cell type-specific differences in telomere dynamics. Furthermore, we observed eventual stabilization of telomere length without detectable changes in telomerase activity during establishment of two leukemic cell lines from normal cord blood cells, indicating that additional cooperating events are required for telomere maintenance in immortalized human hematopoietic cells.hematopoiesis ͉ leukemogenesis

show abstract

Telomere length measurements in leukocyte subsets by automated multicolor flow‐FISH

Cited by 100 publications

References 24 publications

Novel automated three‐dimensional genome scanning based on the nuclear architecture of telomeres

Novel automated three‐dimensional genome scanning based on the nuclear architecture of telomeres

Cellular Image Analysis and Imaging by Flow Cytometry

Dissociation of telomerase activity and telomere length maintenance in primitive human hematopoietic cells

Contact Info

Product

Resources

About