SEARCH · Search PubMed

Results for “Single cell”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

SourceResearch connectionFrom yearThrough year

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 361 records · Page 20Linked to original sources

PubMed2025

MaxComp: Predicting single-cell chromatin compartments from 3D chromosome structures.

The genome is organized into distinct chromatin compartments with at least two main classes, a transcriptionally active A and an inactive B compartment, broadly corresponding to euchromatin and heterochromatin. Chromatin regions within the same compartment preferentially interact with each other over regions in the opposite compartment. A/B compartments are traditionally identified from ensemble Hi-C contact frequency matrices using principal component analysis of their covariance matrices. However, defining compartments at the single-cell level from sparse single-cell Hi-C data is challenging, especially since homologous copies are often not resolved. To address this, we present MaxComp, an unsupervised method, for inferring single-cell A/B compartments based on 3D geometric considerations in single-cell chromosome structures-derived either from multiplexed FISH-omics imaging or 3D structure models derived from Hi-C data. By representing each 3D chromosome structure as an undirected graph with edge-weights encoding structural information, MaxComp reformulates compartment prediction as a variant of the Max-cut problem, solved using semidefinite graph programming (SPD) to optimally partition the graph into two structural compartments. Our results show that the population average of MaxComp single-cell compartment annotations closely matches those derived from ensemble Hi-C principal component analysis, demonstrating that compartmentalization can be recovered from geometric principles alone, using only the 3D coordinates and nuclear microenvironment of chromatin regions. Our approach reveals widespread cell-to-cell variability in compartment organization, with substantial heterogeneity across genomic loci. When applied to multiplexed FISH imaging data, MaxComp also uncovers relationships between compartment annotations and transcriptional activity at the single-cell level. In summary, MaxComp offers a new framework for understanding chromatin compartmentalization in single cells, connecting 3D genome architecture, and transcriptional activity with the cell-to-cell variations of chromatin compartments.

Chromatin↗

PubMed2002

Study of the mutagenicity of "Hypiran" and "Sankol" in human blood cells and comparison with hydrogen peroxide by single cell gel electrophoresis (SCGE) or comet assay.

Due to the increased use and availability of herbal medicinal products, it has been necessary to perform investigations pertaining to their toxicological safety. The aim of this study was to use the rapid and sensitive technique of Single Cell Gel Electrophoresis (SCGE) or "Comet Assay" to assess the mutagenic potential of two such products: "Hypiran" and "Sankol." The results were compared against positive (hydrogen peroxide) and negative (no mutagen) control groups. The instances of DNA damage were observed in 25 randomly selected cells and viewed under a fluorescence microscope following electrophoresis and staining with ethidium bromide. Both microscopic inspection and calculation of the number of affected/damaged cells were encouraging and were comparable to the observations of previous studies using SCGE. The total instances of cell damage in the presence of "Hypiran" increased as the dose increased, with all 25 cells exhibiting fragmentation at the highest dose. However, "Sankol" only exhibited some damage (8-12% of cells) at all doses. The results obtained suggest that in vitro, "Hypiran" produces dose dependent DNA damage and therefore may possess some mutagenic potential. Compared to the positive control, no significant DNA damage was observed with "Sankol" and hence this compound may have no mutagenic potential. However, further study is required before a full evaluation of these products can be made.

Comet Assay↗

PubMed1987

Single cell translocations in couples with multiple spontaneous abortions.

Single cell translocations have been previously reported to occur in normal lymphocyte cultures. Comparison of the frequency of these in 140 individuals referred for a history of multiple miscarriages and 415 individuals referred for a history of multiple miscarriages and 415 individuals referred for other reasons showed a significantly greater number of single cell translocations in the former group. This group also had a significantly greater number of other types of single cell structural abnormalities. Increased chromosome instability, chromosome mosaicism, residual fetal trophoblasts, and immunological differences are discussed in considering the possible etiology.

Abortion, Habitual↗

PubMed2026

Single-cell vector copy number analysis of phenotypically defined long-term hematopoietic stem cells for gene therapy safety assessment.

Hematopoietic stem cell (HSC)-based gene therapy has emerged as a transformative approach for the treatment of genetic diseases; however, accurate evaluation of vector copy number (VCN) remains critical for ensuring safety. Conventional bulk VCN assays, including quantitative PCR (qPCR) and droplet digital PCR (ddPCR), do not resolve clonal heterogeneity and cannot identify rare high-VCN cells that may contribute disproportionately to insertional mutagenesis risk. Here, we developed an accessible single-cell VCN profiling method by combining fluorescence-activated cell sorting (FACS) of phenotypically defined long-term HSCs (Lineage- CD34+ CD38- CD90+ CD45RA- cells) with whole-genome amplification followed by conventional qPCR. This approach enabled resolution of VCN distributions at single-cell level using standard laboratory techniques. Notably, single-cell analysis revealed a high VCN tail that bulk VCN analysis could not resolve. Furthermore, in a humanized mouse transplantation model, single-cell VCN profiling demonstrated that overall VCN distributions could be analyzed after engraftment, although inter-donor and inter-mouse variability was observed. Collectively, this method provides a rapid, cost-effective, and phenotypically resolved strategy for assessing VCN heterogeneity in gene-modified HSCs. Single-cell VCN profiling offers complementary insights beyond conventional bulk assays and may enhance preclinical safety evaluation of gene and cell therapy products.

lentiviral vector↗

PubMed2025

Decoding heterogeneous single-cell perturbation responses.

Understanding how cells respond differently to perturbation is crucial in cell biology, but existing methods often fail to accurately quantify and interpret heterogeneous single-cell responses. Here we introduce the perturbation-response score (PS), a method to quantify diverse perturbation responses at a single-cell level. Applied to single-cell perturbation datasets such as Perturb-seq, PS outperforms existing methods in quantifying partial gene perturbations. PS further enables single-cell dosage analysis without needing to titrate perturbations, and identifies 'buffered' and 'sensitive' response patterns of essential genes, depending on whether their moderate perturbations lead to strong downstream effects. PS reveals differential cellular responses on perturbing key genes in contexts such as T cell stimulation, latent HIV-1 expression and pancreatic differentiation. Notably, we identified a previously unknown role for the coiled-coil domain containing 6 (CCDC6) in regulating liver and pancreatic cell fate decisions. PS provides a powerful method for dose-to-function analysis, offering deeper insights from single-cell perturbation data.

Single-Cell Analysis↗

PubMed2005

Study of T-cell costimulatory blockade in vivo at a single-cell level.

The critical role of costimulatory signals in T-cell activation and the complexity of T-cell costimulatory pathways involved make a detailed understanding of this system a challenging task. By taking advantage of the unique chemical properties of CFSE, we and others have developed an in vivo model that allows quantitative analysis of T-cell activation at a single-cell level. This model involves labeling of donor T-cells with the tracking dye CFSE and adoptively transferring into lethally irradiated allogeneic hosts. T-cells proliferating in the host mice can be explicitly analyzed upon recovery. By using mice deficient for certain costimulatory molecules as a source of donor cells or by treating the host mice with reagents that block certain costimulatory pathways, this CFSE model is extremely useful in studying the role of T-cell costimulatory signals in activation, survival, and effector differentiation of alloreactive T-cells in vivo.

Adoptive Transfer↗

PubMed2005

Assessment of DNA damage in Down Syndrome patients by means of a new, optimised single cell gel electrophoresis technique.

Single cell gel electrophoresis (SCGE), also known as comet assay is a widely used method to detect DNA damage. Its use is nonetheless subjected to some pitfalls, due to differences in experimental set-up, to operator-dependent variability and to quantification of the comets, which is usually accomplished by visual scoring or by image-analysis software. Biological variability in the extent of DNA damage must be taken into account particularly regarding in vivo studies. In the present paper we propose an improved methodology where major features are: a) cryopreservation of lymphocytes collected at different time points and simultaneous analysis in a single run; b) use of an internal control on each slide; c) development of a custom-made software with semi - automated image analysis in order to overcome operator dependent variability. Cryopreservation was accomplished by storing lymphocytes in liquid nitrogen in a solution commonly used for preserving vital cells to be reinfused. We found that this procedure did not alter DNA after 2 and 4 months of storage. The use of quality control from a batch of aliquoted lymphocytes from a healthy donor on each slide, enabled to highlight possible experimental anomalies as well as verify inter-experimental variability. Moreover, by using a newly developed software able to automatically recognise comets we minimised operator-dependent variability in the scoring process. This improved methodology is proposed for longitudinal in vivo studies and in the present work its application made it possible to assess a significant increase of DNA in pediatric Down Syndrome patients compared to healthy controls of the same age.

Comet Assay↗

PubMed2006

Kinetics of single cells: observation and modeling of a stochastic process.

The successive generation times for single cells of Escherichia coli K-12 were measured as described by A. Elfwing, Y. LeMarc, J. Baranyi, and A. Ballagi (Appl. Environ. Microbiol. 70:675-678, 2004), and the histograms they generated were used as empirical distributions to simulate growth of the population as the result of the multiplication of its single cells. This way, a stochastic birth model in which the underlying distributions were measured experimentally was simulated. To validate the model, analogous bacterial growth curves were generated by the use of different inoculum levels. The agreement with the simulation was very good, proving that the growth of the population can be predicted accurately if the distribution of the first few division times for the single cells within that population is known. Two questions were investigated by the simulation. (i) To what extent can we say that the distribution of the detection time, i.e., the time by which a single-cell-generated subpopulation reaches a detectable level, can be identified with that of the lag time of the original single cell? (ii) For low inocula, how does the inoculum size affect the lag time of the population?

Cell Division↗

PubMed2000

[Study on the amplification uniformity of whole genome in a single cell by degenerate oligonucleotide primed polymerase chain reaction].

OBJECTIVE: To investigate the uniformity of whole genome amplification from a single cell by degenerate oligonucleotide primed polymer ase chain reaction (DOP-PCR). METHODS: Amplification of the whole genomic DNA from a single cell with XX, XY, XO, XXY, +13 or +21 was performed by DOP-PCR. Comparative genomic hybridization (CGH) of a single cell DOP-PCR product against the genomic DNA or a single cell DOP-PCR product from normal male as carried out respectively. RESULTS: The average profile of fluorescence intensive ratio in CGH with the genomic DNA as the reference was greatly fluctuated and the standard deviation in about 30% haploid was beyond the normal limits. False positive hyperrepresentation was existed in X chromosome while trisomy 13 and 21 were failed to be detected. However, the distributions of the mean and the standard deviation of the ratio in the CGH with DOP-PCR product as the reference were acceptable. The copy number changes of chromosome X, Y, 13 and 21 were demonstrated. CONCLUSION: Although the single cell DOP-PCR product is not completely uniform, it would be reliable to study the whole genome of a single cell by controlling the conditions of PCR and selecting the suitable control reference because the unequal amplification is unrandom.

Female↗

PubMed1994

Lymphocyte predominance Hodgkin's disease: lineage and clonality determination using a single-cell assay.

Lymphocyte predominance Hodgkin's disease (LPHD) is a clinically indolent condition. Although there is evidence that the putative neoplastic cell in this disease, the "L&H" cell, is of B-cell lineage, there is conflicting data concerning the clonality of these cells. Our study was aimed at clarifying the issue of lineage and clonality of the L&H cells of LPHD using a single-cell assay. Four cases of LPHD were studied. To circumvent the difficulties of obtaining fresh tissue and to be able to study representative cases, a new method was developed to obtain single-cell suspensions of L&H cells from archival formalin-fixed paraffin-embedded tissue. Single L&H cells were identified by morphology and immunostaining for epithelial membrane antigen, isolated using a micropipette, and subjected to polymerase chain reaction (PCR) amplification of the complematarity determining region 3 (CDR3) of the Ig heavy chain (IgH) gene, which is B-cell clone-specific. The PCR products were size-fractionated by polyacrylamide gel electrophoresis and representative products were directly sequenced. Single T cells and small B cells were also isolated from the tissues and used as negative and positive controls, respectively. In all four cases of LPHD, the IgH CDR3 of single L&H cells could be amplified. Within each case, the IgH CDR3 of single L&H cells was found to be of different length or of different sequence. Therefore, our results provide strong evidence for the B-cell origin of the L&H cells and the polyclonal nature of LPHD.

Antigens, CD↗

PubMed2005

Single cell manipulation, analytics, and label-free protein detection in microfluidic devices for systems nanobiology.

Single cell analytics for proteomic analysis is considered a key method in the framework of systems nanobiology which allows a novel proteomics without being subjected to ensemble-averaging, cell-cycle, or cell-population effects. We are currently developing a single cell analytical method for protein fingerprinting combining a structured microfluidic device with latest optical laser technology for single cell manipulation (trapping and steering), free-solution electrophoretical protein separation, and (label-free) protein detection. In this paper we report on first results of this novel analytical device focusing on three main issues. First, single biological cells were trapped, injected, steered, and deposited by means of optical tweezers in a poly(dimethylsiloxane) microfluidic device and consecutively lysed with SDS at a predefined position. Second, separation and detection of fluorescent dyes, amino acids, and proteins were achieved with LIF detection in the visible (VIS) (488 nm) as well as in the deep UV (266 nm) spectral range for label-free, native protein detection. Minute concentrations of 100 fM injected fluorescein could be detected in the VIS and a first protein separation and label-free detection could be achieved in the UV spectral range. Third, first analytical experiments with single Sf9 insect cells (Spodoptera frugiperda) in a tailored microfluidic device exhibiting distinct electropherograms of a green fluorescent protein-construct proved the validity of the concept. Thus, the presented microfluidic concept allows novel and fascinating single cell experiments for systems nanobiology in the future.

Animals↗

PubMed2004

Microdosimetry of astatine-211 single-cell irradiation: role of daughter polonium-211 diffusion.

A microdosimetric analysis of previously published data on 211At-albumin, free 211At, and 211At-C215 irradiation of Colo-205 cells in a slowly rotating single-cell suspension is presented. A custom-built computer program based on the Monte Carlo method was used to simulate the irradiation and the energy deposition in individual cell nuclei. Separate simulations were made for the assumption that the 211Po atom stays in the position where it is created, and that it diffuses away. The mean event number at which 37% of all cells survived, n37, and the frequency mean specific energy per event, zF, were estimated. The Poisson distribution of events and simulated single and multievent distributions of specific energy were used to find the single-cell specific energy at which the probability of survival is reduced to 37%, z37. The calculated single-cell radiosensitivity values show that 211Po atoms, created on a cell surface by the decay of 211At atoms, will diffuse from the cell during its life-span. The increasing distance to the cell nucleus will drastically decrease the probability of the emitted alpha particle to hit the nucleus. This will result in fewer alpha-particle events in the cell nucleus. For dispersed cells, the diffusion of 211Po atoms will reduce the total dose from cell-bound 211At by a factor of 2.

Alpha Particles↗

PubMed2006

Fluorescence-activated single cell sorting of human embryonic stem cells.

Human embryonic stem cells (hESC) are the subject of intense investigation for use in regenerative medicine, in toxicity testing, and as models for the study of human development. Automated cell sorting will enhance the isolation of homogenous pools of differentiated hESCs both for basic studies and for therapeutic applications. Sorting could also be used to deplete undifferentiated, potentially tumourigenic cells. However, hESCs are sensitive to single cell disaggregation and recover poorly when plated at clonal density. Here we report a method for successful semi-automated single cell sorting of hESCs. This method utilizes an ES-specific promoter-transgene construct and automated FACS-based single cell sorting and plating. Clonal recovery in physiologic oxygen (2%) was increased fourfold over room oxygen (21%; p < 0.01). This automated protocol will help to realize proposed hESC strategies that are hampered by low throughput and poor yields.

Alkaline Phosphatase↗

PubMed1985

Characterization of the heterogeneity of R3327 rat prostatic tumors derived from single-cell clones.

Prostatic adenocarcinoma is characterized by cellular diversity, which is well demonstrated in the Dunning R3327 rat prostatic adenocarcinoma. This heterogeneity may arise from epigenetic influences, ie, cellular adaptation or selection, and/or from genetic changes. To investigate the question of genetic instability, four tissue culture cell lines were derived from single cells isolated from the uncloned late (UCL) passage of the Dunning R3327H prostate cell culture. Each of these clonally derived tissue cultures was injected into castrated and intact young adult male rats for tumor production. Uncloned early (UCE) and UCL passage tissue cultures were also propagated as solid tumors. Tumors and the cultures from which they were derived were examined for evidence of phenotypic and genetic changes using morphological and cytometric methods. Transmission and scanning electron microscopy revealed only slight differences among the cell cultures. A single population of diploid cells was demonstrated in each of the cell cultures by propidium iodide staining and subsequent flow cytometric measurement of DNA content/nucleus. Tumors of unicellular as well as multicellular origin exhibited extreme heterogeneity of histological features, both among animals as well as within a single tumor. Tumors were surveyed and tissue types were characterized and cataloged. Clone 3 was generally better differentiated than the others; tumors from castrated animals were better differentiated than those from intact animals. Flow cytometry revealed multiple hyperdiploid cell populations that were variable from one sample to another. We concluded that changes in genotype as well as phenotype occurred in the tumors derived from single cells. Some of these changes may have occurred in the cells while still in culture.

Adenocarcinoma↗

PubMed2005

Chemical cytometry for monitoring metabolism of a Ras-mimicking substrate in single cells.

BACKGROUND: Chemical cytometry is an emerging technology that analyzes chemical contents of single cells by means of capillary electrophoresis or capillary chromatography. It has a potential to become an indispensable tool in analyses of heterogeneous cell populations such as those in tumors. Ras oncogenes are found in 30% of human cancers. To become fully functional products, oncogenic Ras proteins require at least three posttranslational modifications: farnesylation, endoproteolysis, and carboxyl-methylation. Therefore, enzymes that catalyze the three reactions, farnesyltransferase (FTase), endoprotease (EPase), and methyltransferase (MTase), are considered highly attractive therapeutic targets. In this work, we used chemical cytometry to study the metabolism of a pentapeptide substrate that can mimic Ras proteins with respect to their posttranslational modifications in solution. METHODS: Mouse mammary gland tumor cells (4T1) and mouse embryo fibroblasts (NIH3T3) were incubated with a fluorescently labeled pentapeptide substrate, 2',7'-difluorofluorescein-5-carboxyl-Gly-Cys-Val-Ilu-Ala. Cells were washed from the substrate and resuspended in phosphate buffered saline. Uptake of the substrate by the cells was monitored by laser scanning confocal microscopy. Single cells were injected into the capillary, lysed, and subjected to capillary electrophoresis. Fluorescent metabolic products were detected by laser-induced fluorescence and compared with products obtained by the conversion of the substrate by FTase, EPase, and MTase in solution. Co-sampling of single cells with the in-vitro products was used for such comparison. RESULTS: Confocal microscopy data showed that the substrate permeated the plasma membrane and clustered in the cytoplasm. Further capillary electrophoresis and chemical cytometry analyses showed that the substrate was converted into three fluorescently labeled products, two of which were secreted in the culture medium and one remained in the cells. The intracellular product was present at approximately 100,000 molecules per cell. The three metabolic products of the substrate were found to be different from the products of its processing by FTase, EPase, and MTase in solution. CONCLUSIONS: This is the first report of chemical cytometry in the context of Ras-signaling studies. The chemical cytometry method used in this work will find applications in the development of suitable peptide substrates for monitoring enzyme activities in single cells.

Alkyl and Aryl Transferases↗

PubMed2025

ShortCake: an integrated platform for efficient and reproducible single-cell analysis.

SUMMARY: Recent advances in single-cell analysis have introduced new computational challenges. Researchers often need to use multiple analysis tools written in different programming languages while managing version conflicts between related packages within a single workflow. For the research community, minimizing the time spent on environment setup and installation issues is essential. We present ShortCake, a containerized platform that integrates a suite of single-cell analysis tools written in R and Python. ShortCake isolates competing Python tools into separate virtual environments that can be easily accessed within a Jupyter notebook. This enables users to effortlessly transition between various environments, including R, even within a single notebook. Additionally, ShortCake offers multiple "flavors," enabling users to select container images tailored to their specific needs. ShortCake provides a unified environment with fixed versions of various tools, thus streamlining workflows, reducing setup time, and improving reproducibility. AVAILABILITY AND IMPLEMENTATION: The ShortCake image is available on DockerHub (https://hub.docker.com/r/rnakato/shortcake) and Zenodo (DOIs: 10.5281/zenodo.17116765 and 10.5281/zenodo.17118158). The source code is available on GitHub (https://github.com/rnakato/ShortCake).

Single-Cell Analysis↗

PubMed1990

Biomedical and clinical applications of automated single cell electrophoresis.

The automated single cell electrophoresis microscope Parmoquant has been used for the discrimination of lymphocytes and the study of the interaction of substances with cells and synthetic particles. Electrophoretic histograms allowed the determination of changes in the proportion of lymphocyte populations after kidney transplantation, during dialysis treatment, open heart surgery and during pregnancy. Discrimination of leukemic cells on the basis of electrophoresis was used as an additional parameter in diagnosis. In a mouse tumor model, histogram determination enabled the in vivo effect of the tumor necrosis factor on immune cells to be evaluated. Cell electrophoresis was shown to be suitable to detect the influence of antibodies, lectins and bacteria on the cell surface. Protein adsorption was studied on synthetic particles using cell electrophoresis. This method was applied to investigate the phenomena of blood interaction with biomaterials used in artificial organs and to determine differences in the protein composition of serum or other body fluids connected with diseases.

Animals↗

PubMed2026

Early immune dysregulation in Mtb/SIV co-infection resists cART treatment at the single-cell level.

Using single-cell transcriptomics of bronchoalveolar lavage cells from Mtb/SIV co-infected rhesus macaques on cART, we reveal profound immune dysregulation during early SIV co-infection of latent tuberculosis. SIV induces a sharp decline in CD4+ T cells, NK, and NKT cells, with incomplete recovery of Mtb-specific TH1 effector responses despite viral suppression. Instead, a persistent TH17-skewed environment emerges, alongside sustained myeloid inflammation driven by Type I interferon signaling and pro-inflammatory regulators such as KLF6 and NFKB1. Ligand-receptor network analyses demonstrate expanded CD4+ T cell-macrophage crosstalk and loss of immune homeostasis that cART fails to fully restore. These findings expose how SIV remodels the pulmonary immune landscape to impair protective immunity against Mtb, providing a transcriptomic framework to explain TB reactivation in HIV infection. Our work highlights the urgent need for adjunctive immunotherapies to complement cART, aiming to rebalance immune responses and improve TB control in co-infected individuals.

HIV↗