bluuebunny/crossref_metadata_2025_split huggingface dataset:
Localization of a mouse centromeric DNA repeat in interphase nuclei
Authors: Disteche Christine M. , Adler David A. 
Published: August 2005

AbstractThe position of a mouse DNA repeat located near the centromere of mouse chromosomes X, 11, 13, and 17 was examined in interphase nuclei of bone marrow and fibroblast cells by in situ hybridization of 3H‐ or biotin‐labeled DNA probe 70‐38. In most laboratory mouse strains this probe recognizes a single repeat cluster (DXWas70) close to the centromere of the mouse X chromosome. In a few mouse strains, a second locus (D11Was70, D13Was70, or D17Was70, depending on the mouse strain) is located near the centromere of an autosome. In interphase nuclei from mouse strains with the X‐linked locus only, two distinct sites of hybridization were found in female mice and one in male mice. These two sites remained separated during thedifferent phases of the cell cycle (G1, early S, late S, and G2) as demonstrated by in situ hybridization of the probe to flow‐sorted nuclei. In interphase nuclei from mouse strains with both the X‐linked locus and an autosomal locus, four distinct sites of hybridization were found in female mice and three in male mice. Further analysis of loci DXWas70 and D17Was70 showed that these loci were often located in the outer region of nuclei from bone marrow and fibroblast cells.
Multiple fluorescence in situ hybridization
Authors: Nederlof P. M. , van der Flier S. , Wiegant J. , Raap A. K. , Tanke H. J. , Ploem J. S. , van der Ploeg M. 
Published: August 2005

AbstractA method for multiple fluorescence in situ hybridization is described allowing the simultaneous detection of more than three target sequences with only three fluorescent dyes (FITC, TRITC, AMCA), respectively emitting in the green, red, and blue.This procedure is based on the labeling of (DNA) probes with more than one hapten and visualisation in multiple colors. The possibility to detect multiple targets simultaneously is important for prenatal diagnosis and the detection of numerical and/or structural chromosome aberrations in tumor diagnosis. It may form the basis for an in situ hybridization based chromosome banding technique.
System for flow sorting chromosomes on the basis of pulse shape
Authors: Bartholdi Marty F. , Parson Jimmie D. , Albright Kevin A. , Cram L. Scott 
Published: August 2005

AbstractSorting on the basis of the complex features resolved by chromosome slit‐scan analysis requires rapid and flexible pulse shape acquisition and processing for determining sort decisions before droplet breakoff. Fluorescence scans of chromosome morphology contain centromeric index and banding information suitable for chromosome classification, but these scans are often characterized by variability in length and height and require sophisticated data processing procedures for identification. Setting sort criteria on such complex morphological data requires digitization and subsequent computation by an algorithm tolerant of variations in overall pulse shape.We demonstrate here the capability to sort individual chromosomes based on their morphological features measured by slit‐scan flow cytometry. To do this we have constructed a sort controller capable of acquiring an 128 byte chromosome waveform and executing a series of numerical computations resulting in an area‐based centromeric index sort decision in less than 2 ms. The system is configured in a NOVIX microprocessor, programmed in FORTH, and interfaced to a slit‐scan flow cytometer data acquisition system. An advantage of this configuration is direct control over the machine state during program execution for minimal processing time. Examples of flow sorted chromosomes are shown with their corresponding fluorescence pulse shapes.
Study of X chromosome abnormality in XX males using bivariate flow karyotype analysis and flow sorted dot blots
Authors: Carter N. P. , Ferguson‐Smith M. E. , Affara N. A. , Briggs H. , Ferguson‐Smith M. A. 
Published: August 2005

AbstractWe have used bivariate flow karyotype analysis to quantify aberrant X chromosome size in 11 XX males. With one exception, the patients could be grouped into those with an X homologue difference greater than normal (Group A, n = 3) and into those whose X homologue difference could not be distinguished from female controls (Group B, n = 7). The range of sizes of the aberrant X chromosome in Y‐sequence positive patients agrees with the variable nature of the X‐Y interchange in these individuals as determined by the use of Y‐specific DNA probes and Southern blotting analysis. In one patient it was possible to sort separately the normal and the X‐Y interchanged homologues for dot blot analysis. The presence of Y sequences and an increased dose of the zinc finger gene, ZFY, were detected in the X‐Y interchanged homologue. In preliminary studies of 5 male and 6 female controls, it was noted that a consistent difference between the two X homologues in females was found which could not be totally explained by errors of the fitting procedure. We suggest that this difference could be due to X inactivation and that the two X homologues in females might be distinguishable.
Light scattering measurement in an arc lamp‐based flow cytometer
Authors: Steen Harald B. 
Published: August 2005

AbstractThe epi‐illumination optics employed in most arc lamp‐based flow cytometers may be modified so as to produce a dark‐field configuration which facilitates highly sensitive detection of both forward and large angle light scattering in an instrument with a “jet on open surface” flow chamber. Forward scattering is detected at angles upwards from about 2°, while large angle scattering includes angles above 18°. Theoretical considerations suggest that large angle scattering measured around 20° may be as efficient as that measured at 90° for the purpose of distinguishing cells on the basis of intracellular structure. This was supported by the finding that dual parameter light scattering histograms of leukocyte suspensions obtained with the arc lampbased instrument were closely similar to such histograms recorded with a laserbased instrument with the large angle detector at 90°. Different species of bacteria could be distinguished by means of the dual parameter light scattering device, as could different species of sea algae. The sensitivity of the device is sufficient to measure 0.2 μm polystyrene particles in both forward and large angle scattering.
Vitality measurement using spectrum shift in Hoechst 33342 stained cells
Authors: Ellwart Joachim W. , Dörmer Peter 
Published: August 2005

AbstractA bivariate flow cytometric technique has been developed in which Hoechst 33342 stained vital cells are discriminated from early damaged cells by differences in their fluorescence emission spectra. A discrete population of cells with intact cell membranes passing from vital to dead could be identified by using propidium iodide exclusion in combination with a concentration‐independent spectrum shift of Hoechst fluorescence to longer wavelengths. The appearance after injury of two subsets of cells with intact membranes representing vital and early damaged cells, respectively, indicates two types of binding of Hoechst 33342 to these cells, presumably resulting from differences in chromatin structure. The method was tested in murine and human hemopoietic cell lines by cytotoxic treatment with cytosine arabinoside and interleukin 3 deprivation in factor‐dependent lines. The power of the method was demonstrated by a cell cycle analysis of the early damaged cells after both cytotoxic treatment and growth factor deprivation.
Flow cytometric analysis of recombinant murine GM‐CSF (rmuGM‐CSF) induced changes in the distribution of specific cell populations in vivo
Authors: Pennline Kenneth J. , Pellerito Frances , DaFonseca Manuel , Monahan Peggy , Siegel Marvin I. , Smith Sidney R. 
Published: August 2005

AbstractThe changes in the distribution of granulocytes, monocytes, and lymphocytes in various tissue compartments following subcutaneous (SC) administration of recombinant murine GM‐CSF (rmuGM‐CSF) in vivo was determined by flow cytometry in time course studies. Balb/c mice were given single, daily SC injections of 1 or 4 μg of rmuGM‐CSF for 10 days. Flow cytometric analysis was performed on bone marrow (BMC), peritoneal exudate (PEC), and peripheral blood (PBC) cell preparations from mice treated for 1, 3, and 10 days. Dual fluorescence was employed to gate on leukocytes (T200+) and analyze for Ig+, Thy 1.2+, MAC+, and 8C5+ (granulocytes) cells. The analyses indicated that SC‐rmuGM‐CSF increased the percentage of 8C5+ cells in PBC after 1 day of treatment. However, significant changes in the cell composition of PEC and BMC were not observed until day 10 of treatment and included increases in 8C5+ cells and the myeloid cell population, respectively. Side scatter analysis (cell density) of PBC and PEC indicated that the percentage of the granulocytic cell population increased significantly in rmuGM‐CSF treated mice. The changes observed in PEC and BMC appeared to be doserelated whereas those observed in PBC were not. These data clearly demonstrate the utility of flow cytometric analyses for detecting selective effects of cytokines on cell populations that are involved in host defense mechanisms.
Flow cytometric detection of multifocal DNA aneuploid cell populations in mastectomy specimens containing a primary breast carcinoma
Authors: van Dam Peter A. , van Bockstaele Dirk R. , Keersmaeckers Guy H. , Uyttenbroeck Frans L. 
Published: August 2005

AbstractDNA flow cytometry was used to study the presence of DNA aneuploid cell populations in macroscopically normal glandular tissue in mastectomy specimens from 30 patients with breast cancer. In the 13 patients with a DNA diploid primary tumor, no DNA aneuploidy could be found in any of the 39 distant specimens assessed. However, DNA aneuploid cell populations were demonstrated in four of the 17 (23%) patients with a primary DNA aneuploid carcinoma and in seven out of 54 (13%) distant tissue samples (P = 0.02). In all cases the DNA index of the DNA aneuploid cells found in the distant samples was identical to that of the primary tumor. The replicate aneuploid DNA indices and histologic controls taken in parallel very strongly suggest that these distant DNA aneuploid cell populations are metastases.
Introduction to flow cytometry data file standard
Authors: Dean Phillip N. , Bagwell C. Bruce , Lindmo Tore , Murphy Robert F. , Salzman Gary C. 
Published: August 2005

AbstractThe Data File Standards Committee of the Society for Analytical Cytology presents a Standard to be used for the storage of data associated with flow cytometric measurements. The Standard specifies a format that provides for the inclusion of all information necessary to fully describe: (1) the instrument used for the measurement; (2) the sample measured; (3) the data obtained; and (4) the results of analysis of the data. The Committee and the Society for Analytical Cytology point out that the use of this Standard by all those individuals and companies that generate or use data taken with flow cytometers or generate methods of analysis for the data will encourage the sharing of such data and methods of analysis.
Quantitative imaging of immunocytochemical (pap) estrogen receptor staining patterns in breast cancer sections
Authors: Sklarew R. J. , Bodmer S. C. , Pertschuk L. P. 
Published: August 2005

Abstract“Receptogram Analysis” has been developed as a pattern‐oriented approach for predicting endocrine response in breast cancer based upon quantification of the estrogen receptor immunocytochemical assay (ERICA), using a Quantimet Imaging System. Response prediction was evaluated in 58 stage III and IV patients receiving endocrine therapy (primarily Tamoxifen). The Receptogram is a composite of the univariate distributions of nuclear receptor content, IOD(S), and concentration (MOD), and their bivariate contour plot; where (S) is the calculated nuclear radius in section. MOD distributions were classified into four types based upon peak modality and kurtosis (I–IV), and contour plots were classified into four subtypes (A–D) based upon contour slope. Patients failing therapy were ERICA ‐ or their receptogram revealed co‐existent ER+ and ER− tumor cells (type II), highly skewed MOD distributions lacking defined peaks (type IV), or contours with nearly horizontal slope (type C). Response was realized in 9/16 type I patients, with a single positive MOD peak, and in 9/15 type III patients, with discrete, multimodal MOD peaks. In contrast, 0/8 type II, 0/12 type IV, and 0/10 type C patients were responders. Receptogram analysis was superior to cytosol assay (DCC) as a response discriminant: positive predictive value, 53% vs. 33%; negative predictive value, 100% vs. 75%; sensitivity, 100% vs. 83%; specificity, 68% vs. 23%; and accuracy, 78% vs. 41%, respectively. Alternately, patients were assigned to potentially responsive or nonresponsive groups based upon thresholded mean receptor parameters: field MOD, mean nuclear MOD (NMOD), and mean NMOD(PF) where PF is the ER+ nuclear fraction. While these parameters correlated with DCC (r = .72, 0.69, and 0.69), they were only marginally better in predictive value.
System for acquisition and real‐time processing of multidimensional slit‐scan flow cytometric data
Authors: Robinson R. D. , Wheeless D. M. , Hespelt S. J. , Wheeless L. L. 
Published: August 2005

AbstractThe high‐speed sampling requirements of multidimensional slit‐scan signals (cell contours) have typically required custom hardware. This specialized hardware has often lacked the flexibility to adapt to varying instrument setups and experimental requirements. A hardware and software system capable of sampling multiple slit‐scan cell contours at rates of up to 40 MHz with 10‐bit resolution is described. It utilizes commercially available CAMAC transient recorders, a Digital Equipment Corp. PDP‐11/83 computer, and custom hardware for signal conditioning and trigger generation. The modular design of the software system allows various hardware options with minimal additional coding. Real‐time digital processing checks each cell contour for multiple peaks; extracts morphological features such as width, height, and area; accumulates gated histograms of these data; and optionally saves the derived data, selected contours, or both into list mode files on disk.
Flow cytometric evaluation of the cytotoxicity of novel antiviral compounds
Authors: Nassiri M. Reza , Hudson Jerry L. , Pudlo Jeffrey S. , Birch Gary M. , Townsend Leroy B. , Drach John C. 
Published: August 2005

AbstractTwo acyclic analogs of bromotubericidin were tested for cytotoxic effects on uninfected cells by monitoring cell growth and measuring cell cycle perturbations using flow cytometry. As reported elsewhere, 5‐bromotubercidin analogs in which ribose was replaced by 2‐hydroxyethoxymethyl (compound 102) or by 1,3‐dihydroxypropoxymethyl (compound 183) were potent inhibitors of human cytomegalovirus (HCMV) replication in vitro (Pudlo et al.: Journal of Medicinal Chemistry 31:2086–2092, 1988). Because these compounds also inhibited the growth of uninfected cells, we performed kinetic studies with an established neoplastic line of human cells (KB) using flow cytometry. Growth of KB cells treated with either compound 102 or 183 were inhibited in a dose‐dependent manner. Growth inhibition by compound 183, however, was not fully expressed for at least 24 h. DNA analysis by flow cytometry showed that a 4‐h incubation with 10 μM compound 102 caused a decrease of cells in G2/M phase. Cells began to accumulate in early S phase by 12 h of incubation, leading to mid S phase accumulation at 21 h. Compound 183 at 10 μM slightly decreased the number of cells in G2/M phase after a 4‐h incubation, and led to accumulation of DNA in S phase after a 12‐h incubation. By 24 and 30 h, DNA histograms appeared similar to those of control cells but with a slight accumulation of the population in early S phase. In separate experiments, drugs were removed following a 24‐h incubation. After removal of compound 102, KB cell growth resumed with a normal population doubling time. In contrast, the effects of compound 183 were not reversible, suggesting the two compounds acted by different biochemical mechanisms.
Evidence for false aneuploid peaks in flow cytometric analysis of paraffin‐embedded tissue
Authors: Joensuu Heikki , Alanen Kalle A. , Klemi Pekka J. , Aine Risto 
Published: August 2005

AbstractIt has recently been shown that bimodal histograms with false aneuploid peaks may be obtained by DNA flow cytometry from histologically normal tissue allowed to autolyze. To investigate if such peaks can be generated from surgically excised archival tissue, 198 paraffin blocks from 179 patients containing histologically normal spleen (n = 65), liver (n = 26), thyroid (n = 32), pancreas (n = 19), salivary gland (n = 49), or lymph node tissue (n = 7), obtained from the archives of two university pathology departments, were analyzed for nuclear DNA content. The great majority (n = 160, 83.8%) of the 191 interpretable histograms had a single symmetrical G1 peak; and 8 histograms, all produced from liver tissue had a tetraploid pattern. A slight or a prominent repeatable deviation in the G1 peak outline was present in 14 (7.3%) cases. A peak resembling an aneuploid G1 peak with a DNA index (DI) ranging from 1.14 to 1.38 was repeatedly produced from 9 (4.7%) blocks containing histologically normal or inflamed splenic (n = 3), pancreatc (n = 3), liver (n = 1), thyroid (n = 1), or lymph node (n = 1) tissue. The three abnormal peaks produced from pancreatic tissue were rounded in shape and resembled closely the ones that can be obtained from autolytic pancreatic tissue, and the six remaining extra peaks were all fused with the “diploid” peak. In conclusion, false peaks, probably caused by degradation of the nuclear contents during formalin fixation or before it, may rarely be obtained from surgical paraffin‐embedded samples.
Establishing optimal lymphocyte gates for immunophenotyping by flow cytometry
Authors: Loken Michael R. , Brosnan Jeanne M. , Bach Bruce A. , Ault Kenneth A. 
Published: August 2005

AbstractIt is possible accurately to distinguish lymphocytes from other leukocyte populations in peripheral blood using the combination of fluorescence associated with CD45/CD14 and forward and orthogonal light scatter. By identifying the cell population of interest based on immunofluorescence, a light scattering window can then be drawn to include all (≥98%) of the lymphocytes. In this manner, maximal recovery of the lymphocytes within a sample can be consistently obtained. The combination of light scattering and immunofluorescence can also be used to define the purity of the gate. The identification of nonlymphocytes within the light scattering gate can then be used to establish an accurate denominator for the percent lymphocytes stained. Once the optimal data acquisition gate has been established and characterized, it is possible to correct subsequent analyses with that particular sample since the reactivity of monoclonal antibodies on monocytes and granulocytes can be accounted for once the nonlymphocytes have been identified as being within the acquisition gate.
Selective cloning of hybridoma cells for enhanced immunoglobulin production using flow cytometric cell sorting and automated laser nephelometry
Authors: Marder Philip , Maciak Ronald S. , Fouts Rebecca L. , Baker Robert S. , Starling James J. 
Published: August 2005

AbstractTechniques for selective cloning of murine hybridoma cells by flow cytometric cell sorting and use of automated laser nephelometry to determine the resultant clones' immunoglobulin secretion levels are described. Using a commercially available attachment to a fluorescence‐activated cell sorter, individual hybridoma cells were successfully distributed into microtiter wells in an automated manner based on their forward angle light scatter properties and their reaction to fluorescein‐conjugated anti‐mouse‐IgG. The techniques were used to estimate successfully the frequency of immunoglobulin‐secreting cells in established cultures. In addition, heterogeneity of cell surface immunoglobulin expression was observed and utilized as a criterion for flow sorting of new hybridoma variants. In these studies, clones derived from high (anti‐IgG) intensity sorting regions yielded cultures with enhanced immunoglobulin secretion levels, as determined by automated laser nephelometry. Furthermore, the surface immunoglobulin phenotype of the derived clones was conserved in subsequent progeny. Finally, it was established that inclusion of propidium iodide in the hybridoma cell sorting mixtures improved cloning efficiency by facilitating enhanced discrimination and elimination of nonviable cells. Our results indicate that flow cytometric‐assisted single cell deposition provides positive attributes of several traditional hybridoma cloning techniques and, in addition, furnishes a tool for steering the cloning process toward selection of enhanced immunoglobulin producing cultures.
Increased light scattering resolution facilitates multidimensional flow cytometric analysis
Authors: Terstappen Leon W. M. M. , Mickaels Ronald A. , Dost Richard , Loken Michael R 
Published: August 2005

AbstractMultidimensional flow cytometry identifies cell populations as clusters in a space created by the analysis of multiple parameters simultaneously. Optimal use of this multidimensional space requires each of the individual parameters to provide additional information for cell population discrimination as well as maximum utilization of the dynamic range available for each parameter. In this study we improve the visualization of the information present in light scattering signals from leukocytes to facilitate multidimensional flow cytometric analysis. Optimization of cell preparation techniques are essential to obtain high resolution light scattering signals that give complete separation of the granulocytes, monocytes, and granular and nongranular lymphocytes. The angle at which the forward scattered light was collected was modified to enhance the separation between leukocyte populations. Although orthogonal light scattering signals separate granular and nongranular lymphocytes, the resolution and dynamic range could not be displayed using linear or logarithmic functions. By applying a polynomial function to the orthogonal light scattering signals, all leukocyte populations could be displayed while maintaining high resolution. The combination of high resolution light scattering with a nonlinear display resulted in an equally spaced distribution of the cell populations distinguished by correlating forward and orthogonal light scattering signals. Using this approach, peripheral blood neutrophils, eosinophils, basophils, monocytes, and granular and nongranular lymphocytes were shown to occupy distinct locations in the correlation of orthogonal and forward light scattering. Surprisingly,the basophilic granulocytes were located close to granular lymphocytes and monocytes rather than near neutrophils and eosinophils.