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At least 757 records · Page 42Linked to original sources
PubMed1990

Sequential analysis of the thymocyte differentiation in fully allogeneic bone marrow chimera in mice. II. Further characterization of the CD4+ or CD8+ single positive thymocytes.

Differentiation of CD4+8- and CD4-8+ single-positive (SP) thymocytes in fully allogeneic bone marrow chimeras were investigated using multicolor cytometric analysis. The proportion of CD3+ cells in CD4+ SP population derived from donor mice considerably increased between day 12 and 14 after bone marrow transplantation (BMT), and gradually increased thereafter. The proportion of V beta 8+ cells in the CD3+CD4+ population remained constant (around 20%) at each period, suggesting that alpha and beta chains were used as TCR. The proportion of J11d+ cells in the CD4+ SP thymocytes transiently increased from day 12 to 14 and decreased thereafter, even though almost half of CD4+ SP cells were still dull J11d+ at day 35 after BMT. When CD8+ SP populations were analyzed, the proportion of CD3+ cells was very small until day 18. Thereafter, the proportion considerably increased and reached a maximum (83.2%) at day 21. The proportion of V beta 8+ cells in the CD3+ CD8+ SP population fell within range between 20 and 30%. However, before day 18, most of the V beta 8+ cells were dull positive, while after day 21 the majority were bright V beta 8+. Further, CD8+ SP cells at day 12, 14 and 18 were largely bright J11d+. After day 21, however, the proportion of bright J11d+ cells rapidly decreased. Similar results were obtained when the sequence of appearance of CD4+ and CD8+ SP cells was compared among bright CD3+, bright V beta 8+ or J11d- mature populations. The CD4+ SP cells regularly appeared earlier than CD8+ SP cells in the mature populations. These findings indicate that a considerable heterogeneity exists within both CD4+ and CD8+ SP populations and that the differentiation process for CD4+ SP cells precedes that for CD8+ SP cells.

Animals↗
PubMed1996

Laser treatment of tattoos in darkly pigmented patients: efficacy and side effects.

BACKGROUND: Many modalities for the treatment of tattoos and pigmented lesions produce a greater risk of complications in Fitzpatrick types V and VI skin because of an increased incidence of adverse pigmentary changes and keloidal scarring. In fair-skinned persons Q-switched lasers have proved effective in removing pigmented lesions and tattoos without scarring. OBJECTIVE: This study was conducted to determine the efficacy and effects of Q-switched lasers on a small series of darkly pigmented patients with tattoos. METHODS: Four patients of Ethiopian origin with facial and neck tribal tattoos were treated with both the Q-switched ruby and Nd:YAG lasers. One black woman with a multicolored tattoo on the mid chest was treated with the Q-switched ruby laser. RESULTS: Clearing of all lesions was seen. The treatments did not result in scarring or permanent pigment changes other than the ones intended. CONCLUSION: Our results indicate that in darkly pigmented patients, Q-switched laser treatment of tattoos can be performed successfully. The longer wavelength Q-switched Nd:YAG laser is recommended when removing tattoos in darker complected persons. A test treatment is advised before treatment of large skin areas.

Adult↗
PubMed2002

Apoptosis of immune cells in the tumor microenvironment and peripheral circulation of patients with cancer: implications for immunotherapy.

Rapid turnover of lymphocytes observed in patients with cancer appears to be driven by increased apoptosis of T lymphocytes or insufficient thymic output of recent thymic emigrants (RTE). Using multicolor flow cytometry and apoptosis assays, we found that CD8+CD95+Annexin+ T cells are dying at a rate that is significantly higher in patients with cancer than in normal controls (NC). CD8+ effector subsets of T cells were particularly vulnerable to apoptosis. Thymic excision circle (TREC) analysis of peripheral blood lymphocytes showed a decreased number of RTE in these patients. Together, the data suggest that a high rate of T-cell turnover might contribute to immunologic imbalance in patients with cancer and have unfavorable effects on immunotherapy, including therapeutic antitumor vaccines.

Apoptosis↗
PubMed2003

M-FISH analysis of chromosome aberrations in human fibroblasts exposed to energetic iron ions in vitro.

Confluent human fibroblast cells were exposed to 6 Gy gamma-rays or 200 MeV/nucleon Fe ions at 0.7 or 3 Gy. The cells were allowed to repair for 24 hours after exposure and chromosomes were collected using a premature chromosome condensation technique with calyculin-A. Chromosome aberrations were analyzed using the multicolor FISH (mFISH) technique that allows identification of both complex and truly incomplete exchanges. Results showed that both doses of the Fe ions produced higher ratios of complex to simple exchanges and lower ratio of complete to incomplete exchanges than the 6 Gy gamma-exposure. The ratios of aberration yields were similar for the two doses of Fe ions. After 0.7 Gy of Fe ions, most complex aberrations were found to involve three or four chromosomes, indicating this is the maximum number of chromosome domains traversed by a single Fe ion track.

Cell Line↗
PubMed2002

The stress-associated acetylcholinesterase variant AChE-R is expressed in human CD34(+) hematopoietic progenitors and its C-terminal peptide ARP promotes their proliferation.

OBJECTIVE: Hematopoietic stress responses involve increases in leukocyte and platelet counts, implying the existence of stress responsive factors that modulate hematopoiesis. Acetylcholinesterase (AChE) is expressed in mammalian neurons and hematopoietic cells. In brain, it responds to stress by mRNA overexpression and alternative splicing, yielding the rare stress-associated "readthrough" AChE-R variant protein. This led us to explore the hematopoietic involvement of AChE-R and its cleavable C-terminal peptide ARP. MATERIALS AND METHODS: AChE mRNA variants were labeled in CD34(+) hematopoietic progenitor cells by in situ hybridization. ARP expression was detected by multicolor flow cytometry. Bromo-deoxyuracil incorporation and viable cell counts served to evaluate the proliferative effects of ARP and suppressive effects of the AChE antisense oligonucleotide AS1 on CD34(+) cells. RESULTS: The distal enhancer, proximal promoter, and first intron of the human AChE gene include consensus binding sites for hematopoietically active and stress-induced transcription factors. CD34(+) cells from human cord blood were found to express all three variant AChE mRNAs, having different intracellular distributions. ARP was found in 5 to 15% of adult peripheral blood, bone marrow, and fetal CD34(+) cells (both committed CD38(+) and uncommitted CD38(-)) and in acute myeloid leukemia blasts. Externally supplied ARP by itself facilitated the proliferation of CD34(+) cells in an antisense suppressible manner. When combined with early-acting cytokines, ARP enhanced survival and expansion of CD34(+) cells up to 28 days in culture. CONCLUSIONS: Our findings support ARP, the C-terminal peptide of AChE-R, as a new hematopoietic growth factor that may promote the myelopoietic expansion and thrombopoiesis characteristic of stress and may be used to enhance the efficiency of ex vivo expansion for bone marrow transplantation.

Acetylcholinesterase↗
PubMed2000

Practical applications of genotypic surveys for forensic STR testing.

Legitimate genotype frequency estimation for multiallelic loci relies on component allele frequencies, as population surveys represent only a fraction of possible DNA profiles. Multilocus genotypes from two ethnic human populations, African American (n=195) and U.S. Caucasian (n=200), were compiled at 13 STR loci that are used worldwide in forensic investigation (D3S1358, vWA, FGA, D16S539, TH01, TPOX, CSF1PO, D8S1179, D21S11, D18S51, D5S818, D13S317, and D7S820). Sex-specific AmpFlSTR multiplexes provided stringent PCR-based STR typing specifically optimized for multicolor fluorescence detection. Heterozygosity at each STR locus ranged from 0.57 to 0.89 and encompassed from seven (TH01) to twenty-one (D21S11) alleles. Homozygosity tests, tests based on the distinct numbers of observed homozygous and heterozygous classes, log likelihood ratio tests, and exact tests assessed that the degree of divergence from theoretical Hardy-Weinberg proportions for all 13 STRs does not have practical consequence in genotype frequency estimation. Departures from linkage equilibrium, between loci, that imposed significance to forensic calculations were not indicated by observed variance of the number of heterozygous loci or Karlin interclass correlation tests. For forensic casework, reliable multilocus profile estimates may be obtained from the product of component genotype frequencies, each calculated through application of the Hardy-Weinberg equation to population database allele frequency estimates reported here. The average probability that two randomly selected, unrelated individuals possess an identical thirteen-locus DNA profile was one in 1.8x10(15) African Americans and one in 3.8x10(14) U.S. Caucasians.

Black or African American↗
PubMed2001

Fluorescent SSCP of overlapping fragments (FSSCP-OF): a highly sensitive method for the screening of mitochondrial DNA variation.

The mtDNA analysis (mtDNA) is increasingly being demanded for forensic purposes due to the fact that many times the use of standard nuclear marker fails to analyze degraded samples (such as bones) and specially for the analysis of hair shafts (a common sample in the crime scene). However, analysis of mtDNA sequencing implies a great lab effort when a high number of samples must be analyzed. The present work introduces a novel and reliable method for the screening of mtDNA variation in the first and second hypervariables (HV1 and HV2) regions which we have denominated fluorescent single strand conformation polymorphism (SSCP) of overlapping fragments (FSSCP-OF). FSSCP-OF is based on the basic theory of SSCP analysis and combines two complementary strategies: the use of PCR amplified overlapping fragments and fluorescent detection technology. The overlap region contains a high percentage (50%) of the d-loop mtDNA variation and for this reason, the probability to detect a polymorphic position by SSCP analysis is clearly increased in comparison to conventional SSCP methods due to the fact that the same polymorphic position is usually placed in a different "relative" position in the two overlapped fragments. The use of multicolor fluorescent technology allows also the multiplex amplification of overlapping fragment and its subsequent analysis in an automatic sequencer. We have analyzed 50 samples of unrelated individuals through the FSSCP-OF technique and we have found that using this methodology the probability to distinguish two samples with different sequences is close to 100%. FSSCP-OF has other important advantages with respect to previous screening methods, such as the automation and standardization of the protocols, which is of special interest for the forensic routine.

Base Sequence↗
PubMed1997

Laser treatment of tattoos.

All three Q-switched laser systems can effectively remove most tattoos with minimal scarring or other adverse sequelae. Despite advances in laser technology, all tattoos cannot be completely eliminated, and several wavelengths remain necessary to optimally treat multicolored tattoos. The major advantage of Q-switched laser irradiation to effect tattoo removal is the low risk of scarring associated with treatment. Limitations include the need for multiple treatment sessions, minimal to incomplete responses in some cases, and the possibility of pigmentary and textural changes. Research continues in an effort to perfect laser removal of tattoos.

Anesthesia, Local↗
PubMed1992

Chromosomal assignment of human YAC clones by fluorescence in situ hybridization: use of single-yeast-colony PCR and multiple labeling.

Alu-PCR provides a convenient tool for amplification of human-specific sequences from yeast DNA containing yeast artificial chromosomes (YAC) clones. PCR products can be labeled nonisotopically and hybridized in situ, and the chromosomal origin of the clones can be determined. This avoids time-consuming gel purification of the yeast artificial chromosome and the low-efficiency procedure of labeling whole yeast DNA containing the YAC. The application of Alu-PCR to single-yeast colonies permits the mapping of YACs at a very early stage of their characterization. In situ hybridization can detect clones with noncontiguous fragments of DNA, and these can be discarded without further time-consuming characterization. To increase further the potential of the method, we show the application of multicolor hybridization techniques.

Base Sequence↗
PubMed2000

Immunophenotypes in adult acute lymphocytic leukemia. Role of flow cytometry in diagnosis and monitoring of disease.

Flow cytometry has revolutionized the study of hematopoietic cells. Immunophenotyping by multiparameter flow cytometry supplements conventional morphologic diagnosis by providing information on cell lineage and differentiation in ALL and helps monitor disease by improving sensitivity in detecting minimal residual disease. The use of multiple MoAbs and multicolor study by flow cytometry has revealed heterogeneity among ALL and mixed-lineage acute leukemia, which are assigned to the same diagnostic categories by morphology. As technology has improved, clinical and research applications of flow cytometry have expanded to include evaluation of nuclear markers, oncogene proteins, apoptosis, cytokine receptors, and drug resistance. Expanded identification of MoAbs against leukemia-specific markers and the use of QFCM be a significant in managing patients with ALL in the future. In addition, flow cytometry and flow cytometric sorting will be combined more and more with other technologies, such as molecular probing or fluorescence in situ hybridization (FISH). The sorting of rare malignant cells based on immunophenotype and subsequent confirmation by PCR or FISH has already been proven feasible. Ultimately, it is hoped that further definition of subgroups of ALL by immunophenotyping using prognostically significant markers and the use of hybrid technologies of flow cytometry and molecular analysis or cytogenetics will improve treatment strategies for patients with ALL.

Adult↗
PubMedDate not supplied

Localization of five somatostatin receptors in the rat central nervous system using subtype-specific antibodies.

The cloning of five members of the somatostatin receptor family, sst1-sst5, as well as two isoforms of the somatostatin receptor 2, sst2A and sst2B, enabled us to generate specific anti-peptide antisera against unique sequences in the carboxyl-terminal tail of each somatostatin receptor subtype. We used these antibodies in multicolor immunofluorescent studies aimed to examine the regional and subcellular distribution of somatostatin receptors in adult rat brain. Several findings are notable: The cloned sst1 receptor is primarily localized to axons, and therefore most likely functions in a presynaptic manner. The cloned sst2 receptor isoforms exhibit strikingly different distributions, however, both sst2A and sst2B are confined to the plasma membrane of neuronal somata and dendrites, and therefore most likely function in a postsynaptic manner. The cloned sst3 receptor appears to be excluded from 'classical' pre- or postsynaptic sites but is selectively targeted to neuronal cilia. The cloned sst4 receptor is preferentially distributed to distal dendrites, and therefore most likely functions postsynaptically. The cloned sst5 receptor was not detectable in the adult rat brain, however, prominent sst5 expression was found in the pituitary. Furthermore, sst1-containing axons either co-contained somatostatin or were closely apposed by somatostatin-positive terminals in a regional-specific manner. Neuronal somata and dendrites containing either sst2A, sst2B or sst4 were found to exist in close proximity, although not necessarily synaptically linked, to somatostatin-positive terminals. Together, in the central nervous system the effects of somatostatin are mediated by several different receptor proteins which are distributed with considerable regional overlap. However, there appears to be a high degree of specialization among somatostatin receptor subtypes with regard to their subcellular targeting. This subtype-selective targeting may be the underlying principal of organization that allows somatostatinergic modulation of neuronal activity via both pre- and postsynaptic mechanisms.

Animals↗
PubMed2003

Plasmon resonant particles for biological detection.

Several recent advances in the optical observation, fabrication, and bioconjugation of nanometer-sized gold or silver colloids have produced a robust new class of label. These plasmon resonant particle (PRP) conjugates have several important advantages: they are ultra-bright, so the light scattered from the individual particles can be viewed using a simple optical microscope system with a white light illumination source; they do not photo-bleach; PRPs can be prepared that preferentially scatter light of a chosen color; and it is possible to prepare bioconjugated PRPs that are stable in solution. These properties, and the automation of PRP identification, discrimination, and counting, have enabled the development of ultrasensitive, multicolor, and multiplex applications in the life science field.

Biomarkers↗
PubMed2002

Luminescent quantum dots for multiplexed biological detection and imaging.

Recent advances in nanomaterials have produced a new class of fluorescent labels by conjugating semiconductor quantum dots with biorecognition molecules. These nanometer-sized conjugates are water-soluble and biocompatible, and provide important advantages over organic dyes and lanthanide probes. In particular, the emission wavelength of quantum-dot nanocrystals can be continuously tuned by changing the particle size, and a single light source can be used for simultaneous excitation of all different-sized dots. High-quality dots are also highly stable against photobleaching and have narrow, symmetric emission spectra. These novel optical properties render quantum dots ideal fluorophores for ultrasensitive, multicolor, and multiplexing applications in molecular biotechnology and bioengineering.

Biological Assay↗
PubMed1996

Engineering green fluorescent protein for improved brightness, longer wavelengths and fluorescence resonance energy transfer.

BACKGROUND: Variants of the green fluorescent protein (GFP) with different colors would be very useful for simultaneous comparisons of multiple protein fates, developmental lineages and gene expression levels. The simplest way to shift the emission color of GFP is to substitute histidine or tryptophan for the tyrosine in the chromophore, but such blue-shifted point mutants are only dimly fluorescent. The longest wavelengths previously reported for the excitation and emission peaks of GFP mutants are 488 and 511 nm, respectively. RESULTS: Additional substitutions, mainly in residues 145-163, have improved the brightness of the blue-shifted GFP mutants with histidine and tryptophan in place of tyrosine 66. Separate mutations have pushed the excitation and emission peaks of the most red-shifted mutant to 504 and 514 nm, respectively. At least three different colors of GFP mutants can now be cleanly distinguished from each other under the microscope, using appropriate filter sets. A fusion protein consisting of linked blue- and green-fluorescent proteins exhibits fluorescence resonance energy transfer, which is disrupted by proteolytic cleavage of the linker between the two domains. CONCLUSIONS: Our results demonstrate that the production of more and better GFP variants is possible and worthwhile. The production of such variants facilitates multicolor imaging of differential gene expression, protein localization or cell fate. Fusions between mutants of different colors may be useful substrates for the continuous in situ assay of proteases. Demonstration of energy transfer between GFP variants is an important step towards a general method for monitoring the mutual association of fusion proteins.

Amino Acid Sequence↗
PubMed1997

Color appearance depends on the variance of surround colors.

BACKGROUND: The perceived color at each point in a visual scene depends on the relationship between light signals from that point, and light signals from surrounding areas of the scene. In the well known phenomenon of simultaneous color contrast, changing the overall brightness or hue of an object's surround induces a complementary shift in the perceived brightness or hue of the object's color. Color contrast is thought to contribute to color constancy with changes in illumination. RESULTS: We report a new type of simultaneous color contrast, in which changing only the variance (i.e. contrasts and saturations), but not the mean, of colors in a test spot's surround induces a complementary shift in the perceived contrast and saturation of the test spot's color. Objects appear much more vivid and richly colored against low-contrast, gray surrounds than against high-contrast, multicolored surrounds. CONCLUSIONS: Color appearance depends not just on the mean color of the surround, but also on the distribution of surround colors about the mean. This novel form of simultaneous color contrast is inconsistent with a variety of models of color appearance, including those based on sensitivity regulation at the receptor level, and those in which the effects of complex surrounds on color appearance can be reduced to adaptation to the illuminant or induction from a homogeneous 'equivalent surround'. It tends to normalize the gamut of perceived colors in each visual scene and may also contribute to color constancy under viewing conditions that affect contrast.

Color Perception↗
PubMed2002

Trichromatic concept optimizes MAD experiments in synchrotron X-ray crystallography.

The trichromatic concept is a new synchrotron beamline design that optimizes MAD experiments by reducing systematic experimental errors with three-colored and coaxial synchrotron X-ray beams produced by a tandem vertical undulator and trichromator. The concept enables rapid and flexible switching of three defined wavelengths, and extends the flexibility of experimental design for MAD data collection. Thus, we can collect MAD data taking into account time series effects such as radiation damage. The data based on the trichromatic concept gave a better quality electron density map than data collected by conventional methods. It was also revealed that multicolor diffraction using dichromatic or trichromatic X-ray beams is effective in rapid MAD data collection.

Bacterial Proteins↗
PubMed2003

Microbial whole-cell sensing systems of environmental pollutants.

The past decade has witnessed the development of a novel class of tools for environmental monitoring: genetically engineered microorganisms 'tailored' to respond in a dose-dependent manner to changes in environmental conditions. Recent advances in the field include the expansion of available reporter functions with multicolored fluorescent proteins, a broadening of the detected chemical effects such as the availability of nutrients and enhancement of the spectrum of reporter microorganisms to include cyanobacteria, yeast and fungi. Most importantly, the stage has been set for the incorporation of such cells into various whole-cell array formats on silicon chips, optic fibres and other configurations. The future of such multiplex detection and analysis systems seems bright.

Biosensing Techniques↗
PubMed2002

CENP-B is not critical for meiotic chromosome segregation in male mice.

Centromere protein B (CENP-B) is a constitutive protein that binds to a highly conserved 17bp motif located at most mammalian centromeres. To determine whether disruption of this gene affects chromosome segregation in male germ cells, we evaluated the frequencies of disomic and diploid sperm in CENP-B heterozygous and homozygous null mice using the mouse epididymal sperm aneuploidy (m-ESA) assay, a multicolor FISH method with probes for chromosomes X, Y and 8. The specificity and sensitivity of the m-ESA assay was demonstrated using Robertsonian (2.8) translocation heterozygotes as positive controls for sperm aneuploidy. Our results show that the frequencies of disomic and diploid sperm did not differ significantly between CENP-B heterozygous and homozygous null mice (P> or = 0.5) or from 129/Swiss isogenic mice (P> or = 0.5) and B6C3F1 mice (P> or = 0.2). These findings indicate that CENP-B does not have an essential role during chromosome segregation in male meiosis.

Aneuploidy↗