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PubMed1990

Control cells for image cytometric DNA analysis of esophageal tissue and the influence of preoperative treatment.

The nuclear DNA content of 4,960 normal cells in 30 esophagectomy specimens was assessed by image cytometry. In relation to intermediate squamous epithelial cells, the mean transmission values were 25.6% higher for basal squamous epithelial cells, 10.1% higher for fibroblasts and 16.2% lower for lymphocytes. The interslide coefficient of variation (CV) was similar for intermediate and basal squamous epithelial cells, 20.2% and 18.9%, respectively, while fibroblasts and lymphocytes displayed lower interslide CVs, 10.5% and 8.8%, respectively. In spite of their high interslide CVs, the mean DNA values of intermediate and basal squamous epithelial cells had the highest intraslide correlation. This correlation indicates that the preparative and analytical differences between slides that result in alterations in mean DNA values affect squamous epithelial cells of similar origin, size and shape in an analogous manner. Basal squamous epithelial cells replicate their DNA and contain more than the diploid amount; therefore, normal intermediate squamous epithelial cells are suggested as control cells to define the 2c reference value for further DNA studies on squamous epithelial lesions of the esophagus. Preoperative irradiation, with or without chemotherapy, did not alter the DNA values of the investigated control cells.

Adenocarcinoma↗
PubMedDate not supplied

Image cytometry DNA analysis of invasive squamous cell carcinoma of the esophagus.

The Feulgen-DNA content of squamous carcinoma cell nuclei from the human esophagus was assessed in punch biopsies from 47 untreated patients. Forty-four of the 47 biopsies (93.6%) demonstrated aneuploid cell populations, and the remaining 3 (6.4%) were non-diploid. Previous studies have demonstrated that in esophageal dysplasias adjacent to invasive squamous cell carcinoma, DNA in single cells is substantially altered. Thus the process of esophageal carcinogenesis can be monitored not only by histological changes, but also by DNA aberrations in single cells. Quantitative DNA measurement appears, therefore, to be a complement to the histological evaluation of esophageal lesions with suspected, but not unequivocal, evidence of neoplastic growth.

Adult↗
PubMed1974

[Anti-DNA analysis].

Explore the source record for details and available documents.

Antibodies, Antinuclear↗
PubMed1998

[DNA image cytometry in urothelial carcinomas. A pilot study of a new application module of the Lucia G DNA analysis system].

OBJECTIVE: To introduce and test a standardized measurement protocol for DNA image cytometry into a newly developed LUCIA G system application program. Contribution to a discussion concerning the possibility of using the histology sections for DNA image cytometry. STUDY DESIGN: Routine biopsy material from 18 urinary bladder cancer cases was analyzed (Grade 1: n = 6, Grade 2: n = 6, Grade 3: n = 6), both in histological and in cytoseparated preparations. More than one hundred diagnostic and more than 30 reference cells were measured. DNA features (mean DNA value, 2cDI, Böcking's grade of malignancy) and histogram types in Auer's classification (AI-IV) were compared. RESULTS: Most carcinomas measured were aneuploid (two thirds G1, all G2, G3). The most common histogram type was A IV. Malignancy grade was mostly slightly higher in cytoseparated material, the shape of the histogram was analogous. Two carcinomas looking euploid in histology sections were proven aneuploid in cytoseparation. CONCLUSION: More convenient tissue section measurement provided useful DNA profile information in most cases. Those without proven aneuploidy should be additionally measured in cytoseparations. An experienced pathologist meeting strictly measurement protocol demands is an important part of the measurement system.

Carcinoma, Transitional Cell↗
PubMed1990

[Flow cytofluorometric DNA analysis in fresh bladder specimens and its correlations between cytology and mitotic index].

From 1986 to 1987, cell materials of 50 patients with a suspicious history of bladder tumor were determined by flow cytofluorometric DNA measurement and compared with conventional cytogenetic analysis in the institution. The frequency of DNA aneuploid was found 47.6 per cent positive in 42 consecutive cases investigated by means of flow cytometry whereas in 47 cases by cytology and 42 cases by mitotic index (MI) profile a comparable 44.7% and 52.5% of positive findings were observed respectively, showing that there was good correlation between flow cytometry and conventional cytology. In a combined analysis of these 3 potential indices for tumor marker a 74 per cent positive rate could be reached and a good consistency of the 3 indices was observed by 69.7%. The study of DNA distribution pattern by FCM would definitely be of clinical significance in daily practice only if a disagreement appears a reevaluation should then be made. Cytological studies require a high level of skill and experience and it is time-consuming with subjective bias, comparing to FCM which sever as a valuable method with much more diagnostic sensitivity, objectivity and rapidity. Flow cytometry is now developed to the point as a particularly valuable adjuvant for bladder tumor detection.

Adult↗
PubMed1986

DNA microspectrophotometry of bone sarcomas in tissue sections as compared to imprint and flow DNA analysis.

The aim of the present study was to establish an upper limit of diploidy for microspectrophotometric (MSP) DNA measurements in sections of mesenchymal tissue analyzing DNA data of a large number of normal cell populations. The reliability of this upper limit of diploidy for discriminating between diploid and hyperploid bone sarcomas was tested by analyzing the same tumors by MSP in imprint preparations and flow cytometry (FCM). The median DNA value of control cells in tissue sections was given arbitrary value of DNA index (DI) 1.0, denoting the diploid DNA content. The proportion of cells with DNA values exceeding DI 1.25 (greater than DI 1.25) was determined for each normal cell population. The maximum percentage of cells with DNA values exceeding DI 1.25, encountered by analysis of 91 normal cell populations in tissue sections, was 31%. This percentage was set as an upper limit of diploidy. Hence, tumors with a higher percentage of cells greater than DI 1.25 were classified as hyperploid. When we applied this criterion, 31 of 36 sarcomas analyzed by MSP in tissue sections were hyperploid, which was in complete agreement with FCM and MSP in imprints of the same tumors. Apart from discriminating between diploid and hyperploid tumors, an attempt was made to determine peak DNA values of sarcomas analyzed in tissue sections. Peak DNA values, as defined by a minimum of 30% of the cells within a class width of DI 0.25, could be determined for 23 of 36 tumors. These peak DNA values correlated well with corresponding peaks obtained by FCM.(ABSTRACT TRUNCATED AT 250 WORDS)

Bone Neoplasms↗
PubMed2000

Laser scanning cytometry can complement the flow cytometric DNA analysis in paraffin-embedded cancer samples: a paradigmatic case.

Archival studies on paraffin-embedded tumor samples are often complicated by difficulty obtaining a reliable diploid DNA standard. Nontumor cells, e.g., inflammatory and stromal cells, most often found interspersed among tumor cells, would represent a solution to this problem. Unfortunately, there is an inherent difficulty to positively identifying tumor cells in paraffin-embedded specimens. Using an aneuploid paraffin-embedded breast cancer sample, we show here that laser scanning cytometer (LSC) in conjunction with flow cytometry can help to address this issue. Following standard protocols, the tissue was deparaffinized and rehydrated, and the nuclei mechanically isolated before being exposed to propidium iodide. An aliquot served for single-parameter flow cytometric analysis, and the remaining cells were cytocentrifuged onto a microscope slide and LSC analysis was performed. The DNA histogram profiles generated by the two approaches were comparable and both showed the presence of cell populations with different DNA content. To assess the nature of these subsets, we performed a correlated measurement of DNA content and chromatin organization at the single-cell level by LSC. This allowed the identification of several subsets of nuclei. Slides were then stained with Giemsa and the nature of these subsets was assessed morphologically by exploiting the relocating capability of LSC. Inflammatory and stromal cells, residual diploid epithelial cells, and hyperdiploid tumor cells-each characterized by a peculiar coordinate pattern of DNA content and chromatin organization-could be positively identified. Diploid, nontumor cells can then be used as an internal standard for DNA ploidy.

Breast Neoplasms↗
PubMed2002

Evaluation of prognostic factors following flow-cytometric DNA analysis after cytokeratin labelling: I. Breast cancer.

In gynecologic oncology valid prognostic factors are necessary to estimate the course of disease and to define biologically similar subgroups for analysis of therapeutic efficacy. The presented study is a prospective study concerning prognostic significance of DNA ploidy and S-phase fraction in breast cancer following enrichment of tumor cells by cytokeratin labelling. Epithelial cells were labeled by FITC-conjugated cytokeratin antibody (CK 5, 6, 8, and CK 17) prior to flow cytometric cell cycle analysis in 327 fresh specimens of primary breast cancer. Univariate analysis in breast cancer detected the prognostic significance of DNA-ploidy, S-phase fraction and CV (coefficient of variation) of G(0)G(1)-peak of tumor cells for clinical outcome, especially for nodal-negative patients. Multivariate analysis could not confirm prognostic evidence of DNA-ploidy and S-phase fraction. In conclusion, in breast cancer no clinical significance for determination of DNA-parameters was found.

Breast Neoplasms↗
PubMed2004

Protein and DNA analysis for the prenatal diagnosis of alpha2-laminin-deficient congenital muscular dystrophy.

Congenital muscular dystrophies (CMD) are characterized by neonatal hypotonia and/or artrogriposis associated with a dystrophic muscle biopsy. The CMD1A form is caused by a deficiency of the alpha2 chain of laminin 2 (LAMA2 gene at 6q2), a protein present in the basal lamina of muscle fibers, in Schwann cells, epidermis, and in fetal trophoblastic tissue. This allows its study for prenatal diagnosis in the chorionic villous (CV), which was performed in a family with one deceased affected CMD1A child. Immunohistochemical analysis of the CV using antibodies against the C- and N-terminal domains of the alpha2-laminin protein showed a normal positive labeling for both antibodies in the "at-risk" CV, which did not differ from the normal control CV. The integrity of the CV membrane was confirmed through the analysis with antibodies against alpha1, beta1, and gamma1 laminins. DNA study using markers flanking the 6q2 region showed that the affected patient and the "at-risk" fetus did not share the same haplotype. Therefore, the fetus was considered normal through both methodologies, which was confirmed after the birth of a clinically normal male baby. As the LAMA2 gene is very large and the spectrum of mutations causing disease is wide, the analysis of the protein in muscle biopsy has been largely used for the diagnosis. Besides, the possibility to detect it in the chorionic villous, mainly using positive markers, also offers a powerful tool for prenatal diagnosis.

Antibodies, Monoclonal↗
PubMed1999

Comparison of randomly amplified polymorphic DNA analysis and pulsed-field gel electrophoresis for typing of Moraxella catarrhalis strains.

Randomly amplified polymorphic DNA (RAPD) and pulsed-field gel electrophoresis (PFGE) for the analysis of 13 Moraxella catarrhalis isolates, 11 successive strains isolated from sputa of five children and 2 isolates obtained the same day from twins, were compared. RAPD and PFGE both yielded nine types from the 13 isolates, showing a chronic colonization with one strain in three patients and a successive colonization with different strains in two patients. The promising results obtained with RAPD should be confirmed with a larger number of strains, but RAPD seems as suitable as PFGE for the typing of M. catarrhalis.

Adolescent↗
PubMed1993

Flow and image cytometry for DNA analysis in bladder washings: improved concordance by using internal reference for flow.

Using flow or image cytometry, we compared the DNA distribution of cells from bladder washings from 52 patients with bladder cancer. For image cytometry, urothelial nuclei (recognized visually) were analyzed for DNA content using polymorphonuclear nuclei as internal diploid reference. For flow cytometry, two methods can be used: either all cells can be analyzed, as commonly performed, or urothelial cells can be analyzed alone, after specific detection. In this flow cytometry study, cells were doubly stained for panurothelial antigens T16 and for DNA. All the cells were first analyzed using peripheral lymphocytes as an external reference; 79% of the results were similar with results obtained from image analysis. For discordant results, flow-cytometric data were reprocessed to identify immunologically stained urothelial cells; one additional case became concordant with image cytometry when only urothelial cells were analyzed, with lymphocytes as diploid reference; a better concordance (94%) was found when the nonurothelial cells of the samples served as a diploid reference instead of peripheral lymphocytes. This suggests that we achieved an improvement of the flow-cytometric evaluations for ploidy assessment, and we conclude that, on these conditions, flow or image analysis can be considered as equivalent methods for DNA content studies of tumors.

Adult↗
PubMed1989

Differentiation between papillary and nonpapillary renal cell carcinomas by DNA analysis.

Recent studies have shown that the deletion of chromosome 3p or the loss of DNA sequences at 3p is generally associated with the development of renal cell carcinoma (RCC). However, chromosome analysis of some papillary RCCs suggested that this type of tumor differs genotypically from the most common nonpapillary RCCs. Therefore, by using cytogenetic and molecular genetic approaches, we examined human papillary and nonpapillary RCCs for the loss of heterozygosity or homozygosity at the short arm of chromosome 3. The constitutional heterozygosity for the DNF15S2 locus and for one allele of the c-erbA beta and the c-raf-1 proto-oncogenes was lost in nonpapillary RCCs, whereas both alleles were retained in each papillary RCC analyzed. We conclude that the loss of DNA sequences at the chromosome 3p region is a genomic change occurring consistently in nonpapillary RCCs, but never occurring in papillary RCCs.

Carcinoma, Renal Cell↗
PubMed1999

[Gastrointestinal stromal tumors: prognostic value of DNA analysis with flow cytometry and of classical clinicopathologic parameters].

BACKGROUND: Biological behavior of gastrointestinal stromal tumours (GIST) is still difficult to predict. The aim of this study was to analyze the prognostic valve of DNA study by flow cytometry in addition to the classical clinicopathologic parameters. PATIENTS AND METHODS: 82 GIST were investigated to assess by multivariable analysis the prognostic significance of classical morphologic and clinical features and DNA flow-cytometric analysis. RESULTS: Features significantly correlated with bad prognosis were: pain, recurrences, metastasis, small intestine tumours, invasion of adjacent tissues, size > 6 cm, infiltrative pattern, high cellularity, high-grade of atypia, mitosis count > 2/50 high powered field, necrosis, DNA aneuploid-hyperploid or near-diploid, S and SG2M-phase of aneuploid/s population/s > 11% and average S-phase > 11%. Multivariable analysis showed that mitotic activity was the only independent variable indicating poor prognosis. CONCLUSIONS: In this series of GIST mitosis count was the only independent prognostic factor, being more important than flow-cytometric variables like DNA ploidy and cellular cycle values.

Adolescent↗
PubMed1993

Chorionic DNA analysis for the prenatal diagnosis of familial hypercholesterolaemia.

Prenatal diagnosis for familial hypercholesterolaemia (FH) was performed by using restriction fragment length polymorphisms (RFLPs) of the LDL receptor gene on chorionic villi DNA taken during the 10th week of pregnancy. Both parents were FH heterozygotes and had previously had a healthy son and an FH homozygous son. Two RFLPs were informative in this family and revealed that the fetus was unaffected by FH. At birth the child was found to have an LDL cholesterol level of 30 mg/dl and a normal LDL receptor activity in cultured umbilical cord fibroblasts. RFLP analysis on chorionic villi DNA is highly recommended for all heterozygous FH couples in whom the LDL receptor gene mutation/s is/are still to be characterized.

Cells, Cultured↗
PubMed2003

DNA analysis of the genes encoding acidocin LF221 A and acidocin LF221 B, two bacteriocins produced by Lactobacillus gasseri LF221.

Lactobacillus gasseri LF221, an isolate from the feces of a child, produces two bacteriocins. Standard procedures for molecular techniques were used to locate, clone and sequence the fragments of LF221 chromosomal DNA carrying the acidocin LF221 A and B structural genes, respectively. Sequencing analysis revealed the gene of acidocin LF221 A to be an open reading frame encoding a protein composed of 69 amino acids, including a 16-amino-acid N-terminal extension. The acidocin LF221 B gene was found to encode a 65-amino-acid bacteriocin precursor with a 17-amino-acid N-terminal leader peptide. DNA homology searches showed similarities of acidocin LF221 A to brochocin B, lactococcin N and thermophilin B, whereas acidocin LF221 B exhibited some homology to lactacin F and was virtually identical to gassericin X. The peptides encoded by orfA1 and orfB3 showed characteristics of class II bacteriocins and are suspected to be the complementary peptides of acidocin A and B, respectively. orfA3 and orfB5 are proposed to encode putative immunity proteins for the acidocins. Acidocin LF221 A and acidocin LF221 B are predicted to be members of the two-component class II bacteriocins, where acidocin LF221 A appears to be a novel bacteriocin. L. gasseri LF221 is being developed as a potential probiotic strain and a food/feed preservative. Detailed characterization of its acidocins is an important piece of background information useful in applying the strain into human or animal consumption. The genetic information on both acidocins also enables tracking of the LF221 strain in mixed populations and complex environments.

Amino Acid Sequence↗
PubMed2003

Molecular cloning and DNA analysis of a gene encoding alpha mating pheromone from the yeast Saccharomyces naganishii.

A DNA fragment encoding the precursor peptide for alpha mating pheromone was isolated from the S. naganishii genome based on the amino acid sequence of the mature pheromone. The precursor peptide contains three copies of the pheromone. Hydrophobicity analysis of the precursor peptide revealed an N-terminal signal sequence for translocation into the lumen of the endoplasmic reticulum and several signals for a series of secretion-related processes. However, upstream regulatory sequences necessary for expression of the S. cerevisiae alpha mating pheromone gene were not found, suggesting the divergence of systems that regulate alpha mating pheromone gene expression in S. naganishii and S. cerevisiae. Hybridization of a probe corresponding to the S. naganishii alpha mating pheromone nucleotide sequence to S. naganishii chromosomal DNA revealed a single gene located on either chromosome VI or VII. The S. naganishii alpha mating pheromone sequence has been deposited in the DDBJ/EMBL/GenBank data library under Accession No. AB086431.

Amino Acid Sequence↗