Variability of DNA analysis by image cytometry

Reeder, Jay E.; C, Cox; Ll, Wheeless; Flint, Andrew; Liebert, Monica; Hb, Grossman

doi:10.1002/(sici)1097-0320(19970601)28:2<176::aid-cyto12>3.0.co;2-n

Cited by 7 publications

(6 citation statements)

References 3 publications

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“…Studies using automated image analysis in classi cation of neoplasms (70) have shown a sensitivity of 85% to 99%, a speci city of 86% to 98% and a positive predictive value of 87% to 97%. Variability in automated image analysis (42) has been due to specimen variability in systematic sampling in 92% of the cases and specimen variability in selective sampling in 67%. Technical variability has been responsible for 10% of the differences.…”

Section: Discussionmentioning

confidence: 99%

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Collagen Matrix in Development and Progression of Experimentally Induced Respiratory Neoplasms in the Hamster

Laitakari

Stenbäck

2001

Toxicol Pathol

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Intratracheal instillations of 7H-dibenzo(c, g)carbazole (DBC), a tobacco smoke component, into Syrian golden hamsters, resulted in preneoplastic lesions and benign and malignant respiratory neoplasms. Neoplastic progression was associated with specific changes in the extracellular matrix (ECM), dependent on the stage of tumor development. DBC-induced tracheobronchial squamous metaplasia was associated with an increase in collagen type I and type III deposition in the subepithelial ECM, as observed by computer-assisted image analysis of immunohistochemical staining for the aminoterminal propeptides of collagen type I (PINP) and collagen type III (PIIINP). Increased collagen matrix synthesis was detected in dysplasia by in situ hybridization of alpha1(I) mRNA for collagen I and alpha1(III) mRNA for collagen type III after continued exposure to DBC. In well-differentiated squamous cell carcinomas with an expansive growth pattern, collagen deposition increased, as did fiber size. In moderately differentiated neoplasms, basement membrane (BM) destruction and invasion was associated with a destructive growth pattern and decreases in collagen synthesis and the deposition of new collagen. Preserved deposition of mature collagen was detected by staining for the telopeptide of collagen type I propeptide. In less differentiated tumors, ECM development was minimal, with few and small fibers, possibly explaining the rapid development of these neoplasms. Transforming growth factor beta (TGFbeta1) immunoreactivity was increased in hyperplastic epithelium and well differentiated neoplasms and decreased in dysplasia and less differentiated squamous cell carcinomas, while TGFbeta2 and TGFbeta3 expression was also distinct in neoplastic cells. Collagen synthesis and epithelial differentiation were associated with an increased number of myofibroblasts in the ECM and with increased TGFbeta3 immunoreactivity in differentiated cells and in the matrix. The nature of the composition of the ECM was related to neoplastic growth and progression when analyzed by computer-associated image analysis, revealing alterations in collagen structure, size, and shape.

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Section: Discussionmentioning

confidence: 99%

“…Clinical applications in tumor prognostication have also been proposed. Variability in automated image analysis occurs, and Reeder et al (42) reported signi cant differences to be due to specimen variability in sampling. Studies involving controlled conditions and de ned endpoints are few.…”

Section: Introductionmentioning

confidence: 99%

Collagen Matrix in Development and Progression of Experimentally Induced Respiratory Neoplasms in the Hamster

Laitakari

Stenbäck

2001

Toxicol Pathol

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“…The OD lymphocyte nuclei from each section served as an internal control (diploid reference). The resultant DNA histograms were analysed using previously described methods (Raju et al 1993;Cajulis et al 1995;Reeder et al 1997). The G0/G1 peak was identified visually in each lesion, and the mean, standard deviation (SD) and coefficient of variation (CV) values were calculated.…”

Section: Methodsmentioning

confidence: 99%

DNA study of bladder papillary tumours chemically induced by N‐butyl‐N‐(4‐hydroxybutyl) nitrosamine in Fisher rats

Oliveira

Adega

Palmeira

et al. 2006

Int J Experimental Path

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To examine DNA abnormalities in bladder papillary tumours induced by N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) in female rats, using image cytometric DNA analysis and cytogenetics. Thirty female rats were exposed to BBN in their drinking water for 20 weeks. One group of 10 animals served as controls. The animals exposed to BBN were killed at a rate of two per week, with the bladder being collected under aseptic conditions and those tumours with exophytic growth removed. The nuclear DNA content of the tumours was evaluated using image cytometric analysis. In two rats part of the tumour pieces was stipulated for culturing. Cytogenetic analysis was performed on at least 30 cells from each cell population and on both tumours. Papillary carcinomas were classified as low grade and high grade. DNA ploidy studies were carried out on 28 low-grade and 21 high-grade papillary carcinomas. Histograms obtained by image analysis showed that a normal urothelium was diploid; 28.6% and 100% of low-and high-grade papillary carcinomas were aneuploid respectively. Both tumours used for cell culture showed multiple numerical and structural chromosome alterations and several marker chromosomes. Image cytometric DNA analysis proved to be a good and reliable method for examining DNA alterations in papillary bladder carcinomas. The present findings establish that the DNA content is statistically different between low-grade and high-grade papillary carcinomas and that deviation from the diploid number is markedly higher in the high-grade ones. In addition, the occurrence of marker chromosomes seems to be related to the aggressiveness of the tumour.

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“…Most software packages include data filters to automatically eliminate objects touching the edge of the field or falling outside a specified size range. Consistency in the criteria employed for exclusion is important for minimizing variability in results obtained by different investigators (e.g., Thunnissen et al 1996Thunnissen et al ,1997Reeder et al 1997;Vilhar et al 2001).…”

Section: Outline Of Methodologymentioning

confidence: 99%

“…As with flow cytometry, the use of image analysis technology in DNA quantification began in cancer diagnosis (e.g., Borgiani et al 1994;Fischler et al 1994). Given the extreme importance of accuracy in such an application, it is not surprising that the fidelity of the technique has been scrutinized with exceptional rigour (e.g., Böcking et al 1995;Thunnissen et al 1996Thunnissen et al ,1997Reeder et al 1997;Puech and Giroud 1999). Comparisons have shown that flow cytometry and image analysis provide similar efficacy of DNA quantification for diagnostic purposes (e.g., Bertino et al 1994;Borgiani et al 1994;Marcos et al 1998).…”

Section: Image Analysis Densitometrymentioning

confidence: 99%

From Pixels to Picograms

Hardie

Gregory

Hebert

2002

J Histochem Cytochem.

226

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S U M M A R YThe study of genome size variation is important from a number of practical and theoretical perspectives. For example, the long-standing "C-value enigma" relating to the more than 200,000-fold range in eukaryotic genome sizes is best studied from a broad comparative standpoint. Genome size data are also required in detailed analyses of genome structure and evolution. The choice of future genome sequencing projects will be dependent on knowledge regarding the sizes of genomes to be sequenced, and so on. To date, genome size data have been acquired primarily by Feulgen microdensitometry or flow cytometry. Each has several advantages but also important limitations. In this review, we provide a practical guide to the new technique of Feulgen image analysis densitometry. The review is designed for those interested in genome size measurements but not extensively experienced in histochemistry, densitometry, or microscopy. Therefore, relevant historical and technical background information is included. For easy reference, we provide recipes for required reagents, guidelines for cell staining, and a checklist of steps for successful image analysis. We hope that the accuracy, rapidity, and cost-effectiveness of Feulgen image analysis demonstrated here will stimulate further surveys of genome sizes in a variety of taxa.

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Variability of DNA analysis by image cytometry

Cited by 7 publications

References 3 publications

Collagen Matrix in Development and Progression of Experimentally Induced Respiratory Neoplasms in the Hamster

Collagen Matrix in Development and Progression of Experimentally Induced Respiratory Neoplasms in the Hamster

DNA study of bladder papillary tumours chemically induced by N‐butyl‐N‐(4‐hydroxybutyl) nitrosamine in Fisher rats

From Pixels to Picograms

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