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At least 469 records · Page 26Linked to original sources

PubMed2006

Single cell analysis on microfluidic devices.

There is significant variability among cells of the same type at the single cell level. This variability may be because of external stimuli that vary temporally or spatially among a population of cells. It may also be owing to the nonsynchronized responses of cells to various stimuli. In addition, differences in otherwise similar cells may be generated by genetic mutations acquired by one or more of the cells. Often times multiple biochemical pathways and molecules are involved in such differences. In order to better understand these differences and to detect those rare cells in a large population that may be indicative of early disease states, methods that are capable of rapidly quantifying multiple molecular species in single cells are desired. Microfluidic devices may provide the optimal platform upon which to develop such methods. Microfluidics has the capability of combining the high-throughput manipulation and transport of cells with rapid, high-efficiency separations and high-sensitivity detection. This chapter describes how to fabricate microfluidic devices for the high-throughput manipulation and rapid electrical lysis of single, nonadherent (suspension) cells followed by the injection and separation of the fluorescently labeled cell contents.

Carboxylesterase↗

PubMed1999

Accuracy of centre of pressure measurement using a piezoelectric force platform.

OBJECTIVE: To evaluate the accuracy in determining centre of pressure when using a force platform for stabilometry research. DESIGN: Technical report. BACKGROUND: Previous studies have determined the accuracy of locating the centre of pressure by applying force through a single point. In human stabilometry research however, force is distributed across two feet and the centre of pressure located between the feet. METHODS: Forces were applied to a Kistler force platform using a force applied through a single point as well as when force was distributed between two metal blocks. RESULTS: Errors in calculating centre of pressure were substantially larger when force was applied through a single point than when the force was distributed between two blocks. CONCLUSIONS: Evaluating centre of pressure accuracy by applying force through a single point overestimates the errors inherent in human stabilometry research. RELEVANCE: Evaluating changes in the centre of pressure of human subjects standing on a force platform is a common tool for evaluating control mechanisms of balance. Accurate information regarding the centre of pressure is essential for the validity of this research.

Biomechanical Phenomena↗

PubMed2006

Analysis of the binding of the N-terminal conserved domain of yeast Cbf5p to a box H/ACA snoRNA.

During ribosome biogenesis, the RNA precursor to mature rRNAs undergoes numerous post-transcriptional chemical modifications of bases, including conversions of uridines to pseudouridines. In archaea and eukaryotes, these conversions are performed by box H/ACA small ribonucleoprotein particles (box H/ACA RNPs), which contain a small guide RNA responsible for the selection of substrate uridines and four proteins, including the pseudouridine synthase, Cbf5p. So far, no in vitro reconstitution of eukaryotic box H/ACA RNPs from purified components has been achieved, principally due to difficulties in purifying recombinant eukaryotic Cbf5p. In this study, we present the purification of a truncated derivative of yeast Cbf5p (Cbf5(Delta)p) that retains the highly conserved TRUB and PUA domains. We have used band retardation assays to show that Cbf5(Delta)p on its own binds to box H/ACA small nucleolar (sno)RNAs. We demonstrate that the conserved H and ACA boxes enhance the affinity of the protein for the snoRNA. Furthermore, like its archaeal homologs, Cbf5(Delta)p can bind to a single stem-loop-box ACA RNA. Finally, we report the first enzymatic footprinting analysis of a Cbf5-RNA complex. Our results are compatible with the view that two molecules of Cbf5p interact with a binding platform constituted by the 5' end of the RNA, the single-stranded hinge domain containing the conserved H box, and the 3' end of the molecule, including the conserved ACA box.

Amino Acid Sequence↗

PubMed2025

Scalable single-cell total RNA-seq reveals non-coding programs in immunity, infection, and brain development.

Non-coding RNAs represent a widespread and diverse layer of post-transcriptional regulation across cell types and states, yet much of their diversity remains uncharted at single-cell resolution. This gap stems from the limitations of widely used single-cell RNA-sequencing protocols, which focus on polyadenylated transcripts and miss many short or non-polyadenylated RNAs. Here, we adapted single-cell RNA-sequencing on the 10x Genomics platform to capture a broad complement of coding and non-coding RNAs-including miRNAs, tRNAs, lncRNAs, histone RNAs, and non-adenylated viral transcripts. This approach enabled the discovery of rich, dynamic non-coding RNA programs across immune cells, virally infected hepatocytes, and the developing human brain. In dengue virus-infected hepatocytes, we detect non-adenylated viral transcripts and distinguish active from transcriptionally quiescent infected states, each with distinct host regulatory signatures. In brain tissue, we identify biotype-specific, cell-type-restricted non-coding RNAs, including miRNAs whose expression anticorrelates with predicted targets, consistent with post-transcriptional regulatory relationships. We show that MIR137, one of the strongest GWAS loci associated with schizophrenia and intellectual disability, is expressed specifically in Cajal-Retzius cells, an early-born but transient population that guides subsequent cortical neuron migration. These findings demonstrate the importance of non-coding RNAs in defining cell identity and state, and show how expanded transcriptome coverage can reveal additional layers of gene control-now accessible through practical and scalable single-cell profiling.

Journal Article↗

PubMed2004

The effects of estrogens on learning in rats with chronic brain cholinergic deficiency in a Morris water test. Identification of the "passive swimming" component.

Chronic decreases in brain cholinergic functions due to intraventricular administration of the neurotoxin AF64A were accompanied by increases in the latent period of locating an invisible platform during training of rats in a Morris water test, as compared with control sham-operated animals. Recordings of the animals' movement trajectories using a video camera along with an original computer program (Behavioral Vision) showed that administration of 17beta-estradiol and its synthetic analog J-861 (0.2 mg/kg p.o. daily for seven days before and 10 days after single intraventricular injections of AF64A) improved learning. The directivity of platform search trajectories was assessed quantitatively using a new parameter--trajectory straightness. Introduction of the "passive swimming" parameter allowed periods of immobility in water to be identified within the total latent period in animals after administration of AF64A; 17beta-estradiol but not J-861 "eliminated" these periods. The new parameters (especially trajectory straightness) allowed the ability to learn to be discriminated from decreases in mobility, including mobility losses due to study agents, in the Morris water test.

Adrenal Glands↗

PubMed2027

Systematic Dissection of Key Driver Perturbation Signatures in Single Cells via ECCITE-seq.

CRISPR screens, such as expanded CRISPR-compatible cellular indexing of transcriptomes and epitopes by sequencing (ECCITE-seq), enable the simultaneous measurement of transcriptomes, gRNA identity, and cell-surface protein expression at single-cell resolution to systematically interrogate gene function. This platform provides a powerful and scalable experimental approach for validating disease-associated regulators identified by large-scale association studies and other computational methods, including network-based analyses of multi-omics data. Here, as an example application, we describe an ECCITE-seq framework to characterize the transcriptomic consequences of perturbing multiple neuronal key driver genes associated with Alzheimer's disease (AD) in human-induced pluripotent stem cell (hiPSC)-derived neurons. More broadly, by integrating customized pooled gRNA libraries with different CRISPR effectors across multiple cell types, this approach allows for the assessment of the regulatory impact of candidate genes implicated in development and disease processes.

Humans↗

PubMed2026

Dual-Matrix Platform for Highly Specific Multi-Omics Profiling of Renal Cell Carcinoma.

Multiomics interrogation provides complementary information beyond single-omics approaches for improved disease characterization. To enable such multilayer profiling, we expanded the rapid functionalized mesoporous nanoparticle-coupled laser desorption/ionization mass spectrometry (fMNPLDI-MS) platform by designing two structurally homologous but functionally tailored fMNPs. This design enables efficient acquisition of both serum metabolic and peptide fingerprints from a total of only 2.05 μL of serum, with an LDI MS analysis time of approximately 90 s per sample, while addressing the limitation of single-matrix systems in simultaneously optimizing analytical performance for different biomolecular species. Through statistical analysis and machine learning-based feature selection, an integrated multiomics biomarker panel was established, comprising 5 peptides and 4 metabolites. Notably, this integrated panel outperformed both single-omics panels across all evaluation metrics in the validation set, improving the area under curve from 0.985 to 1.000 and increasing the classification accuracy from 0.947 (metabolites) and 0.930 (peptides) to 0.965, while showing consistent improvements in F1-score, precision, and recall. Collectively, these results demonstrate the robust performance of the dual-matrix design and multiomics integration for renal cell carcinoma classification, with potential relevance for broader applications in complex disease profiling.

Carcinoma, Renal Cell↗

PubMed2006

Unique aggregation of anthrax (Bacillus anthracis) spores by sugar-coated single-walled carbon nanotubes.

There has been significant interest in the binding of anthrax spores by molecular species, but with only limited success. Proteins and more recently peptides were used. However, despite the known presence of carbohydrates on the spore surface, carbohydrate-carbohydrate interactions have hardly been explored likely because of the lack of required specific platform for synthetic carbohydrates. We report the successful use of single-walled carbon nanotubes as a truly unique scaffold for displaying multivalent monosaccharide ligands that bind effectively to anthrax spores with divalent cation mediation to cause significant spore aggregation. The work should have far-reaching implications in development of countermeasure technologies.

Animals↗

PubMed2000

Sequence variation in genes and genomic DNA: methods for large-scale analysis.

The large-scale typing of sequence variation in genes and genomic DNA presents new challenges for which it is not clear that current technologies are sufficiently sensitive, robust, or scalable. This review surveys the current platform technologies: separation-based approaches, which include mass spectrometry; homogeneous assays; and solid-phase/array-based assays. We assess techniques for discovering and typing variation on a large scale, especially that of single-nucleotide polymorphisms. The in-depth focus is the DNA chip/array platform, and some of the published large-scale studies are closely examined. The problem of large-scale amplification is addressed, and emerging technologies for present and future needs are indicated.

Base Sequence↗

PubMed1996

Maximum voluntary weight-bearing by the affected and unaffected legs in standing following stroke.

OBJECTIVE: To compare stroke patients to control subjects for ability to transfer body weight onto the affected and unaffected leg in standing; to investigate intra-session reliability. DESIGN: Comparative clinical study conducted within a single session. BACKGROUND: There is a paucity of quantitative data about maximum voluntary weight-bearing in patients during rehabilitation following stroke. METHODS: A Kistler force platform was used to quantify maximum amount of body weight transferred to a single limb in the lateral and forward directions during weight-shifting. Twelve control subjects matched by gender and age (median 64 years) were compared to 12 inpatient stroke patients after a median of 37 days post-onset. RESULTS: The median score for control subjects was approximately 95% of body weight to each leg in both directions. In contrast, stroke patients transferred less body weight (P<0.01) to the affected leg (65.5% lateral; 54.9% forward) and also to the unaffected leg (85.0% lateral; 80.1% forward). For the stroke patients, transfer of body weight was more challenging in the forward direction than the lateral direction on the affected leg (P<0.05). Relative to individual differences in the stroke group, error due to the repeated measurement process was low. CONCLUSION: The testing procedure was found to discriminate between stroke patients and control patients and had high retest reliability within a single session.

Journal Article↗

PubMed2005

Association of nuclear factor-kappaB in psoriatic arthritis.

OBJECTIVE: To examine the association of single nucleotide polymorphisms (SNP) in the NFKB1 gene, as well as 2 genes in the nuclear factor (NF)-kappaB functional complex (RelA and NFKBIA), in patients with psoriatic arthritis (PsA) from Newfoundland. METHODS: Patients with PsA and controls were genotyped for one 4-base insertion/deletion and 5 SNP in NFKB1, 4 SNP in RelA, and 7 SNP in NFKBIA by time-of-flight mass spectrometry, using the Sequenom platform. Chi-square analysis was used to test the single locus associations between SNP in the NF-kappaB complex and PsA. Associations between multi-locus haplotypes and case or control status were tested using the software PHASE. RESULTS: Two hundred and twenty-four patients with PsA (52% male) and 88 ethnically matched controls (64% male) were genotyped. No association was noted with any of the SNP tested for the single locus associations in NFKB1, RelA, and NFKBIA or with multi-locus haplotypes. In particular, the allele frequency for the NFKB1 -94delATTG was 41.7% in cases and 41.6% in the controls (p = 0.97). CONCLUSION: No association between the NFKB1 -94 ins/delATTG promoter polymorphism or with other NF-kappaB complex SNP in patients with PsA from Newfoundland was observed.

Adult↗

PubMed2004

Locked nucleic acid (LNA) single nucleotide polymorphism (SNP) genotype analysis and validation using real-time PCR.

With an increased emphasis on genotyping of single nucleotide polymorphisms (SNPs) in disease association studies, the genotyping platform of choice is constantly evolving. In addition, the development of more specific SNP assays and appropriate genotype validation applications is becoming increasingly critical to elucidate ambiguous genotypes. In this study, we have used SNP specific Locked Nucleic Acid (LNA) hybridization probes on a real-time PCR platform to genotype an association cohort and propose three criteria to address ambiguous genotypes. Based on the kinetic properties of PCR amplification, the three criteria address PCR amplification efficiency, the net fluorescent difference between maximal and minimal fluorescent signals and the beginning of the exponential growth phase of the reaction. Initially observed SNP allelic discrimination curves were confirmed by DNA sequencing (n = 50) and application of our three genotype criteria corroborated both sequencing and observed real-time PCR results. In addition, the tested Caucasian association cohort was in Hardy-Weinberg equilibrium and observed allele frequencies were very similar to two independently tested Caucasian association cohorts for the same tested SNP. We present here a novel approach to effectively determine ambiguous genotypes generated from a real-time PCR platform. Application of our three novel criteria provides an easy to use semi-automated genotype confirmation protocol.

Automation↗

PubMed1987

Persistence of tolerance to a single dose of ethanol in the selectively-bred alcohol-preferring P rat.

The persistence of tolerance to a single dose of ethanol was examined in the selectively-bred alcohol-preferring P line of rats. Tolerance was measured by a test that required trained rats to jump onto a descending platform to avoid footshock. On day 0, each trained rat received a single IP injection of 2.5 g ethanol/kg body weight and was tested every 15 minutes for recovery to a criterion of 75% of pre-alcohol training performance. The second ethanol injection of 2.5 g/kg and testing were carried out seven days later for one group (n = 12), and 14 days later for another group (n = 12). Tolerance was assessed by the differences in time required to recover to criterion performance and blood alcohol concentrations (BACs) at time of recovery on day 0 vs. day 7 and day 14. The mean recovery times and BACs on day 0 were 156 +/- 5 minutes and 222 +/- 6 mg%, respectively. The group injected on day 7 exhibited shorter recovery times of 113 +/- 4 minutes and higher BACs at recovery of 261 +/- 4 mg%, while the group injected on day 14 did not show any significant differences from the values obtained on day 0. In a second experiment, the persistence of tolerance in P rats was compared with that of rats from the alcohol-nonpreferring NP line and of stock Wistar rats (n = 6/group). All rats were trained and tested for recovery to criterion after 2.5 g ethanol/kg on day 0 as described for the first experiment. The rats were then injected with ethanol and tested for tolerance on three subsequent occasions.(ABSTRACT TRUNCATED AT 250 WORDS)

Alcohol Drinking↗

PubMed1994

Alterations in weight-transfer capabilities in adults with hemiparesis.

BACKGROUND AND PURPOSE: The purposes of this study were (1) to examine the position and displacement in the frontal plane of the body's center of mass (CM) with respect to the base of support during single-leg flexion movements in adults with hemiplegia and (2) to examine their relationship with other clinical scores. SUBJECTS: Fourteen ambulatory adult volunteers with hemiparesis of the right side of the body due to cerebrovascular accident participated in the study. METHODS: Subjects performed single-leg flexion movements with the paretic and nonparetic limbs while standing on two separate force platforms. Motion analysis and force platform data were used to determine the displacement of the CM. RESULTS: Successful performance of the transfer and holding single-limb stance occurred for 48% (to the nonparetic side) and 20% (to the paretic side) of the trials. Lack of success was due to insufficient displacement of the CM (26% of the trials to the nonparetic side and 17% of the trials to the paretic side) or a failure to maintain single-limb stance (26% of all trials to the nonparetic side and 63% of the trials to the paretic side). Overall, the final position of the CM with respect to the single-limb support region did not differ between sides. Successful performance was highly to moderately associated with clinical assessment scores for motor function and balance. Its association with gait velocity, however, was poor. CONCLUSION AND DISCUSSION: A classification scheme that can distinguish between four categories of bipedal to single-limb stance transitions has been established. Issues concerning clinical assumptions pertaining to the relationship between static and dynamic motor dysfunction in adults with hemiparesis are discussed.

Adult↗

PubMed2005

Simultaneous release of hydrophobic and cationic solutes from thin-film "plum-pudding" gels: a multifunctional platform for surface drug delivery?

The release of two compositionally different solutes from a composite gel composed of two different populations of microgel particles embedded in a single bulk gel matrix is described, showing the potential of the "plum-pudding gel" as a multifunctional platform for controlled surface release. One hydrophobic solute (pyrene) and one hydrophobic and charged solute (rhodamine 123) were chosen as the solutes to be released. Hydrophobic microgels composed of 50% N-isopropylacrylamide (NIPAM) and 50% N-tert-butylacrylamide (BAM) were loaded with pyrene, and anionic microgels composed of 30% acrylic acid (AAc), 20% NIPAM, and 50% BAM were loaded with rhodamine 123. The two solute-loaded microgel populations were incorporated into a single bulk gel network, from which the two solutes were released simultaneously and independently. Using this structural motif, solutes that are mutually incompatible can be incorporated into a single matrix with which they may also be incompatible. The electrostatically incorporated solute was released much more slowly than the hydrophobically attracted solute, indicating that the microgel composition can be tailored to the specific solute, and thus control its release rate. The choice of bulk matrix was also found to influence the release rate much more than expected, offering a further control element to the system.

Drug Delivery Systems↗

PubMed2026

shinyDeepGxP: a user-friendly R shiny app for predicting surface protein abundance from scRNA-seq expression using deep learning in blood cells.

MOTIVATION: Understanding accurate immune cell heterogeneity and function in single-cell datasets requires access to protein-level information, which is often unavailable due to experimental limitations. RESULTS: We present shinyDeepGxP, an interactive web application featuring our deep learning model, DeepGxP, for predicting surface protein abundance from single-cell RNA-sequencing (scRNA-seq) data. This platform makes DeepGxP accessible to researchers without programming skills. Users can upload scRNA-seq count matrices and use "Predict Protein" to predict the abundance of 224 biologically relevant surface proteins. shinyDeepGxP provides visualizations to help identify distinct cell populations based on predicted protein profiles. Moreover, users can choose "Explore Model" to reveal key RNA predictors and their associated biological pathways for each protein. Overall, shinyDeepGxP is a user-friendly, freely available web tool that provides protein-level detail for RNA-only single-cell datasets, enabling multimodal discovery without additional experiments. AVAILABILITY AND IMPLEMENTATION: shinyDeepGxP can be launched on https://shiny.crc.pitt.edu/deepgxp/.

Journal Article↗

PubMed2004

A unified view of propagating and localized surface plasmon resonance biosensors.

The intense colors of noble metal nanoparticles have inspired artists and fascinated scientists for hundreds of years. In this review, we describe refractive index sensing platforms based on the tunability of the localized surface plasmon resonance (LSPR) of arrays of silver nanoparticles and of single nanoparticles. Specifically, the color associated with single nanoparticles and surface-confined nanoparticle arrays will be shown to be tunable and useful as platforms for chemical and biological sensing. Finally, the LSPR nanosensor will be compared to traditional, flat surface, propagating surface plasmon resonance sensors.

Biosensing Techniques↗

PubMed2005

Age-dependent differences in lateral balance recovery through protective stepping.

BACKGROUND: Aging appears to present particular problems for lateral balance stability related to falls. Protective stepping is a common strategy for maintaining balance that may be impaired with aging due to changes in neuromusculoskeletal factors. This study assessed the response patterns, kinematics, and single support hip abduction torque during lateral protective stepping for balance recovery in healthy young and elderly adults. METHODS: Ten healthy elderly and 10 younger adults received stepper-motor driven waist-pulls of bipedal stance applied pseudorandomly to either side. Stepping response strategies were quantified with force platforms and motion analysis. FINDINGS: The young responded primarily using a single lateral sidestep with the limb that was initially loaded passively by the waist-pull, while older subjects favored crossover stepping using multiple steps with more inter-limb collisions. When the elderly did use loaded side steps, the steps were longer, slower, and higher and included greater and prolonged lateral trunk motion than in the young. Overall, older subjects produced greater and less rapid stabilizing hip abduction torque during the single support phase. INTERPRETATION: Age-associated differences in lateral balance control through stepping included using a riskier recovery strategy with increased collisions between the limbs, multiple steps, altered first step characteristics and lateral trunk motion during direct sidestepping, and a generally greater support hip torque. The difficulties with lateral balance control in aging may reflect factors such as impaired hip abduction torque-time capacity and lateral trunk mobility/control. Our findings contribute additional knowledge pertaining to the problem of balance dysfunction and falls among the elderly.

Accidental Falls↗