Flow cytometric determination of DNA content in isolated nuclei of cereals

Genome ◽  
1988 ◽  
Vol 30 (4) ◽  
pp. 565-569 ◽  
Author(s):  
E. Hülgenhof ◽  
R. A. Weidhase ◽  
R. Schlegel ◽  
A. Tewes
Keyword(s):  
Flow Cytometry ◽  
Dna Content ◽  
Internal Standard ◽  
Aegilops Speltoides ◽  
Isolated Nuclei ◽  
Triticum Timopheevi ◽  
Flow Cytometric ◽  
Nuclear Isolation ◽  
Cereal Plants

Isolated nuclei from cereal plants were used for quantitative determination of DNA per nucleus by flow cytometry. The technique is based on enhanced fluorescence of ethidium bromide and olivomycin when they bind to DNA. Nuclei were isolated from protoplasts derived from leaves of seedlings. The diploid Hordeum vulgare cv. Trumpf was used as an internal standard. Analysis of nuclei from several cultivars of hexaploid wheat (Triticum aestivum), Triticum durum var. hordeiforme, T. durum var. alexandrinum, Triticum araraticum, Triticum timopheevi, Triticum monococcum, Aegilops speltoides, Secale cereale, and one hexaploid and one octoploid triticale revealed significant intra- and inter-specific differences in DNA values per 2C nucleus. The advantages of the procedure are discussed along with its utilization for quantitative DNA measurement in Gramineae.Key words: nuclear isolation, flow cytometry, DNA content, cereals.

scholarly journals NUCLEAR DNA CONTENT OF DENDROBIUM ORCHID SPECIES AS DETERMINED BY LASER FLOW CYTOMETRY

HortScience ◽  
1996 ◽  
Vol 31 (3) ◽  
pp. 322c-322
Author(s):  
W.E. Jones ◽  
A.R. Kuehnle ◽  
K. Arumuganathan
Keyword(s):  
Flow Cytometry ◽  
Dna Content ◽  
Nuclear Dna ◽  
Evolutionary Relationship ◽  
Internal Standard ◽  
Nuclear Dna Content ◽  
Orchid Species ◽  
Group Assignment ◽  
Taxonomic Groups

Flow cytometry (FC) has proven to be an efficient and reliable method to estimate nuclear DNA content (genome size) in quantifiable units useful for genetic and molecular biology studies. This method also makes possible determination of the variation in nuclear DNA content between related taxa, which gives insights into the process of speciation. In this study, DNA content was determined in nuclei isolated from leaves of 21 Dendrobium species representing each of the major taxonomic groups used in the Univ. of Hawaii breeding program. Nuclei were mechanically isolated, stained with the nucleic acid-specific fluorochrom propidium iodide, and DNA content determined using a Coulter Epics 753 laser flow cytometer. Chicken erythrocyte nuclei (2C = 2.33 pg DNA) were used as an internal standard for direct comparative measurement. The mean diploid genome (2C) values for Dendrobium species ranged from 3.36 to 5.06 pg. Genome sizes were evaluated for possible use as discrete characters for taxonomic group assignment and compared to previous data on breeding compatibility and evolutionary relationship between species.

Prognostic Value of Flow Cytometry in Bladder Tumours

1993 ◽  
Vol 60 (2) ◽  
pp. 152-157
Author(s):  
D. Grassi ◽  
M. De Siati ◽  
N. Franzolin
Keyword(s):  
Flow Cytometry ◽  
Transitional Cell Carcinoma ◽  
Cell Carcinoma ◽  
Dna Content ◽  
Prognostic Value ◽  
Transitional Cell ◽  
Prognostic Information ◽  
Carcinoma Of The Bladder ◽  
Bladder Tumours

During this last decade, flow cytometry (FCM) has been widely investigated and employed in assessing the DNA content of bladder tumours. The prognostic value of FCM is recognised by the majority of investigators, above all when it concerns superficial transitional cell carcinoma of the bladder. The determination of ploidy and the degree of aneuploidy seem to offer valuable prognostic information. The Authors have reviewed Literature on this subject, identifying three different categories of studies that analyse the relation of FCM to cytohystological characterisation, to the clinical behaviour of the tumours and to the patients’ survival.

scholarly journals Flow cytometric analysis of herpes simplex virus type 1 susceptibility to acyclovir, ganciclovir, and foscarnet.

1997 ◽  
Vol 41 (12) ◽  
pp. 2686-2692 ◽  
Author(s):  
I Pavić ◽  
A Hartmann ◽  
A Zimmermann ◽  
D Michel ◽  
W Hampl ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Cytopathic Effect ◽  
Flow Cytometric Analysis ◽  
Flow Cytometric ◽  
Resistant Strains ◽  
Simplex Virus

We established a quantitative flow cytometric method for determination of herpes simplex virus type 1 (HSV-1) susceptibility to acyclovir (ACV), ganciclovir, and foscarnet in vitro. Susceptibility was defined in terms of the drug concentration which reduced the number of cells expressing HSV-1 glycoprotein C (gpC) with a fluorescence intensity of > or =10(2) by 50% (IC50). Flow cytometry allowed us to use a high (1.0) as well as a low (0.005) multiplicity of infection, and determination of the IC50 was possible after one or more viral replicative cycles. IC50s were dependent on virus input and on time postinfection. In mixture experiments, 1 to 2% resistant viruses added to a sensitive strain could be detected. The results obtained by flow cytometry showed a good qualitative correlation with those achieved by cytopathic effect inhibitory assay. However, flow cytometry might detect more quantitative differences in drug susceptibility, especially among resistant strains, as confirmed also by determination of intracellular drug phosphorylation. The mean IC50s for ACV-sensitive strains were 0.45 to 1.47 microM, and those for ACV-resistant strains were between 140 and 3,134 microM. Flow cytometric analysis was fast and accurate, automatizable, and highly reproducible. Flow cytometry may be a more powerful tool than standard cytopathic effect-based assays and could have advantages for the detection of low levels of drug resistance or mixtures of sensitive and resistant virus strains.

scholarly journals Measurement of cellular DNA mass by flow microfluorometry with use of a biological internal standard.

1978 ◽  
Vol 26 (2) ◽  
pp. 145-148 ◽  
Author(s):  
E Tannenbaum ◽  
M Cassidy ◽  
O Alabaster ◽  
C Herman
Keyword(s):  
Dna Content ◽  
Internal Standard ◽  
Cell Populations ◽  
Experimental Cell ◽  
Flow Microfluorometry ◽  
Chicken Erythrocytes ◽  
Cellular Dna ◽  
Biological Standard

Use of a biological standard (chicken erythrocytes) mixed with experimental cell populations allows control of all variables in flow microfluorometric determination of DNA content. These variables include both staining and instrument procedures. In addition, the use of a biological standard allows determination of cellular DNA mass in unperturbed cell populations. DNA mass measured by FMF technique correlates closely with values reported in the literature that used biochemical techniques.

scholarly journals Preliminary Characterization of a Monoclonal Antibody (AS-2) against Cell Cycle Related Proteins

1998 ◽  
Vol 17 (2) ◽  
pp. 93-101
Author(s):  
Stefano Nigro ◽  
Anna Rapallo ◽  
Angela Di Vinci ◽  
Elio Geido ◽  
Roberto Orecchia ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Molecular Weight ◽  
Monoclonal Antibody ◽  
Flow Cytometry ◽  
Dna Content ◽  
Staining Pattern ◽  
Isolated Nuclei ◽  
Erythroleukemia Cell ◽  
Related Proteins

A monoclonal antibody (AS-2) raised by using isolated nuclei from a human erythroleukemia cell line as immunogen is described.AS-2 was of IgM type and recognized proteins present in both isolated cytoplasms and nuclei. The molecular weight of the AS-2 recognized proteins in the cytoplasm was 200 kDa and 70 and 60 kDa in the nucleus. The relative amount of these proteins were measured simultaneously with DNA content by flow cytometry. We found the highest protein content (or stainability) for both cells and nuclei in late-G1, S and G2, at approximately the same level, and the lowest content in M and early-G1. Sorting based on DNA content and AS-2 associated fluorescence helped identifying the staining pattern of cells and nuclei. Interphase isolated nuclei and cell cytoplasms were characterized by interdispersed staining over the entire surfaces while mitoses showed two dots only. The present preliminary data indicate that the proteins recognized by the AS-2 monoclonal are cell cycle related and suggest that in mitoses they are associated with the centrosomes.

Flow Cytometric and Immunohistochemical Correlations in High Incidence Human Solid Tumors

1997 ◽  
Vol 83 (3) ◽  
pp. 689-697 ◽  
Author(s):  
Donatella Tirindelli Danesi ◽  
Marcello Spanò ◽  
Fabiana Antonini ◽  
Pierluigi AltaVista ◽  
Piera Catalano ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Dna Content ◽  
Rank Correlation ◽  
Breast Cancer Patients ◽  
Dna Index ◽  
Ploidy Levels ◽  
Spearman’S Rank Correlation ◽  
Flow Cytometric ◽  
Human Solid Tumors ◽  
Stem Cell Lines

475 patients with carcinoma at different sites (141 colon-rectum; 102 breast; 50 stomach; 48 kidney; 46 head and neck; 41 bladder; 47 other sites) submitted to surgery have been analyzed after histopathological staging and grading, by flow cytometry (monoparametric DNA content analysis) and immunohistochemistry (p53, c-erbB-2, and PCNA expression). In breast cancer patients the presence of receptors for estrogen (ER) and progesterone (PGR) has also been determined. Flow cytometry-derived parameters were DNA ploidy, fraction of cells in S-phase (SPF), and DNA content heterogeneity (multiclonal stem cell lines with different DNA index and/or more than one subpopulations with different ploidy levels in different samples from the same tumor). Correlations of the results obtained by the different techniques have been attempted by the non-parametric Spearman's rank correlation approach. Significant associations (P «0.05) were found between the histopathological, immunohistochemical and flow cytometric parameters considered in some anatomical regions, such as stomach (p53 vs DNA content aneuploidy and vs heterogeneity), colon-rectum (TNM vs p53 and vs heterogeneity), bladder (grading vs DNA content aneuploidy and vs heterogeneity). Tumor heterogeneity proved to be dependent on the number of tumor samples taken. The results of this preliminary assessment will subsequently be compared with the data obtained from a currently ongoing follow-up survey.

Sign in / Sign up

Export Citation Format

Share Document