Increased cell-free DNA is associated with oxidative damage in patients with schizophrenia.

Cell-free DNA (cfDNA) has been found to be elevated in patients with schizophrenia (SZ), potentially derived from activated apoptosis, but the underlying mechanisms remain unknown. Moreover, whether the concentrations of cfDNA are altered with disease stage has not been investigated, which limits its clinical application as an auxiliary diagnostic marker for SZ. Using an improved fluorescence correlation spectroscopy (FCS) method that does not require DNA extraction, we measured the molar concentrations of cfDNA in plasma samples of 191 patients with SZ, 78 patients with mood disorders (MD) and 65 healthy controls (HC).

Simultaneously Monitoring Multiple Autophagic Processes and Assessing Autophagic Flux in Single Cells by Fluorescence Cross-Correlation Spectroscopy.

Autophagy is a widely conserved and multistep cellular catabolic process and maintains cellular homeostasis and normal cellular functions the degradation of some harmful intracellular components. It was reported that high basal autophagic activity may be closely related to tumorigenesis. So far, the fluorescence imaging technique has been widely used to study autophagic processes, but this method is only suitable for distinguishing autophagosomes and autolysosomes.

Objective scanning-based fluorescence cross-correlation spectroscopy (Scan-FCCS) for studying the fusion dynamics of protein phase separation.

Protein phase separation plays a very important role in many biological processes and is closely related to the occurrence and development of some serious diseases. So far, the fluorescence imaging method and fluorescence correlation spectroscopy (FCS) have been frequently used to study the phase separation behavior of proteins. Due to the wide size distribution of protein condensates in phase separation from nano-scale to micro-scale in solution and living cells, it is difficult for the fluorescence imaging method and conventional FCS to fully reflect the real state of protein phase separation in the solution due to the low spatio-temporal resolution of the conventional fluorescence imaging method and the limited detection area of FCS.

CRDBP Protein Phase Separation and Its Recruitment to β-Catenin Protein in a Single Living Cell.

The Coding Region Determinant-Binding Protein (CRDBP) is a carcinoembryonic protein, and it is overexpressed in various cancer cells in the form of granules. We speculated the formation of CRDBP granules possibly through liquid-liquid phase separation (LLPS) processes due to the existence of intrinsically disordered regions (IDRs) in CRDBP. So far, we did not know whether or how phase separation processes of CRDBP occur in single living cells due to the lack of methods for studying intracellular protein phase separation.

Deubiquitination Detection of p53 Protein in Living Cells by Fluorescence Cross-Correlation Spectroscopy.

Deubiquitination is a reverse post-translational modification of ubiquitination and plays significant roles in various signal transduction cascades and protein stability. The p53 is a very important tumor-suppressor protein and closely implicates more than 50% of human cancers. Although extracellular studies on the deubiquitination of p53 were reported, the process of p53 deubiquitination in living cells due to the shortage of an efficient in situ method for single living cells is still not clear.

Resonance Light-Scattering Correlation Spectroscopy and Its Application in Analytical Chemistry for Life Science.

Resonance light-scattering correlation spectroscopy (RLSCS) is a new single-particle detection method with its working principle being like fluorescence correlation spectroscopy (FCS). RLSCS is obtained by autocorrelation function analysis on the measured fluctuation of the resonance light scattering (RLS) intensity occurring within a subfemtoliter volume when a single nanoparticle (such as gold nanoparticles (NPs) or silver (SNPs)) freely diffuses through the volume. The RLSCS technique can detect such parameters as concentration, diffusion coefficient (translation and rotation), etc.

monitoring of the ubiquitination of newly synthesized proteins in living cells by combining a click reaction with fluorescence cross-correlation spectroscopy (FCCS).

Newly synthesized proteins are closely related to a series of biological processes, including cell growth, differentiation, and signaling. The post-translational modifications (PTMs) of newly synthesized proteins help maintain normal cellular functions. Ubiquitination is one of the PTMs and plays a prominent role in regulating cellular functions.

measurement of autophagic flux by fluorescence correlation spectroscopy.

Autophagy is a fundamental and phylogenetically conserved self-degradation process and plays a very important role in the selective degradation of deleterious proteins, organelles, and other macromolecules. Although flow cytometry and fluorescence imaging techniques have been used to assess autophagic flux, we remain less able to monitor autophagic flux in a highly sensitive, robust, and well-quantified manner. Here, we reported a new method for real-time and quantitatively monitoring autophagosomes and assessing autophagic flux in living cells based on fluorescence correlation spectroscopy (FCS).

quantitative measurements on MMP-9 activity in single living cells by single molecule fluorescence correlation spectroscopy.

Matrix metalloproteinase-9 (MMP-9) plays an important role in tumor progression. It is of great significance to establish a sensitive assay strategy for MMP-9 activity in single living cells. Here a novel single molecule spectroscopy method based on the fluorescence correlation spectroscopy (FCS) technique was proposed for measuring the MMP-9 activity at different locations within single living cells, using a fluorescent specific peptide and a reference dye as dual probes.

Probing the Protein Corona of Nanoparticles in a Fluid Flow by Single-Particle Differenced Resonance Light Scattering Correlation Spectroscopy.

The protein corona of nanoparticles (NPs) plays a crucial role in determining NPs' biological fates. Here, a novel measurement strategy was proposed to in situ investigate the protein corona formed in the NPs with the home-built dual-wavelength laser-irradiated differenced resonance light scattering correlation spectroscopy (D-RLSCS) technique, combined with the modified generation method of the D-RLSCS curve. With the measurement strategy, the dissociation constants and the binding rates between proteins and gold nanoparticles (GNPs) were determined based on the binding-induced ratiometric diffusion change of NPs (the ratio of characteristic rotational diffusion time to translational one), using the formation of the protein corona of bovine serum albumin (BSA) or fibrinogen (FIB) on gold nanoparticles as a model.

Quantification of mRNA in Single Cells Based on Dimerization-Induced Photoluminescence Nonblinking of Quantum Dots.

Photoluminescence (PL) intermittency (or "blinking") is a unique characteristic of single quantum dot (QD) emission. Here, we report a novel single-molecule detection strategy for the intracellular mRNA of interest using the mRNA-induced nonblinking QD dimers as probes. The working principle of the method is that the DNA hybrid of the target DNA (or mRNA) with a biotin-modified ssDNA probe can induce two blinking streptavidin-modified QDs (SAV-QDs) conjugated.

Brightness Analysis per Moving Particle: Analysis of Alkaline Phosphatase in Living Cells.

quantitative analysis of enzymes such as phosphatase is important to understand a number of involved biological processes ranging from various metabolisms to signal transduction and cellular regulation. In this paper, a novel measurement strategy was proposed to detect alkaline phosphatase (ALP) activity in different locations within single living cells. The principle is based on the measurement of the resonance light scattering brightness ratio (SBR) per moving nanoparticle that forms in an ALP-related chemical reaction.

The Theoretical Model, Method, and Applications of Scattering Photon Burst Counting Based on an Objective Scanning Technique.

Scattering photon burst counting (SPBC) is a single-particle detection method, which is based on measuring scattering photon bursting of single nanoparticles through a detection volume of <1 fL. Although SPBC has been used for bioassays and analysis of nanoparticles, it is necessary to establish its theoretical model and develop a new detection mode in order to further enhance its sensitivity and enlarge its application fields. In this paper, we proposed a theoretical model for the confocal SPBC method and developed a novel SPBC detection mode using the fast objective scanning technique.

Single-Particle Catalytic Analysis by a Photon Burst Counting Technique Combined with a Microfluidic Chip.

Single-particle catalytic analysis plays an important role to understand the catalytic mechanism of nanocatalysts. Currently, some methods are used to study the relationship between single-particle catalytic activity and morphology. However, there is still lack of a simple and rapid analysis method for evaluating the catalytic activity of an individual nanocatalyst that freely moves in solution.

In Situ Assay of Proteins Incorporated with Unnatural Amino Acids in Single Living Cells by Differenced Resonance Light Scattering Correlation Spectroscopy.

Site-specific incorporation of unnatural amino acids (UAAs) into target proteins (UAA-proteins) provides the unprecedented opportunities to study cell biology and biomedicine. However, it is a big challenge to in situ quantitatively determine the expression level of UAA-proteins due to serious interferences from autofluorescence, background scattering, and different viscosity in living cells. Here, we proposed a novel single nanoparticle spectroscopy method, differenced resonance light scattering correlation spectroscopy (D-RLSCS), to measure the UAA-proteins in single living cells.

Simultaneously monitoring endogenous MAPK members in single living cells by multi-channel fluorescence correlation spectroscopy.

The mitogen-activated protein kinase (MAPK) pathway is a major module for cellular signal transduction. The dysregulation of the MAPK pathway has been involved in the pathogenesis of multiple diseases ranging from cancers to chronic inflammations. So far, we have not fully understood the influences of external factors and signaling networks on the MAPK pathway due to the lack of in situ methods for simultaneous detection of multiple kinases in the pathway in living cells.

Studying Homo-oligomerization and Hetero-oligomerization of MDMX and MDM2 Proteins in Single Living Cells by Using In Situ Fluorescence Correlation Spectroscopy.

Protein oligomerization plays a very important role in many physiological processes. p53 acts as a key tumor suppressor by regulating cell cycle arrest, DNA repair, and apoptosis, and its antitumor activity is regulated by the hetero- and homo-oligomerization of MDMX and MDM2 proteins. So far, some traditional methods have been utilized to study the oligomerization of MDMX and MDM2 in vitro, but they have not clarified some controversial issues or whether the extracellular results can represent the intracellular results.

Analyses of p73 Protein Oligomerization and p73-MDM2 Interaction in Single Living Cells Using In Situ Single Molecule Spectroscopy.

Protein oligomerization and protein-protein interaction are crucial to regulate protein functions and biological processes. p73 protein is a very important transcriptional factor and can promote apoptosis and cell cycle arrest, and its transcriptional activity is regulated by p73 oligomerization and p73-MDM2 interaction. Although extracellular studies on p73 oligomerization and p73-MDM2 interaction have been carried out, it is unclear how p73 oligomerization and p73-MDM2 interaction occur in living cells.

Artificial Aquaporin That Restores Wound Healing of Impaired Cells.

Artificial aquaporins are synthetic molecules that mimic the structure and function of natural aquaporins (AQPs) in cell membranes. The development of artificial aquaporins would provide an alternative strategy for treatment of AQP-related diseases. In this report, an artificial aquaporin has been constructed from an amino-terminated tubular molecule, which operates in a unimolecular mechanism.

In Situ Study of Interactions between Endogenous c- mRNA with CRDBP in a Single Living Cell by Combining Fluorescence Cross-Correlation Spectroscopy with Molecular Beacons.

mRNA-protein interactions play key roles in facilitating various biological functions in gene expression regulations and even the progression of diseases. However, it is still a challenge to directly monitor mRNA-protein interactions in a single living cell at present. Herein, we propose a new strategy for real-time studying of mRNA-protein interactions in a single living cell using fluorescence cross-correlation spectroscopy (FCCS) and molecular beacon (MB) labeling techniques.

Controllable "Clicked-to-Assembled" Plasmonic Core-Satellite Nanostructures and Its Surface-Enhanced Fluorescence in Living Cells.

The assembly of noble-metal core-satellite (CS) nanostructures is an appealing means to control their plasmonic properties for their applications such as surface-enhanced fluorescence or Raman scattering. However, till now there is a lack of some rapid or convenient methods to construct stable CS nanostructures. Here, we proposed a "clicked-to-assembly" strategy based on the fast and specific "click chemistry" reaction between trans-cyclooctene (TCO) and 1,2,4,5-tetrazine (Tz).

Fluctuation correlation spectroscopy and its applications in homogeneous analysis.

Fluctuation correlation spectroscopy (FCS) is a single-molecule/particle detection technique based on measuring signal fluctuations in a highly focused detection volume. Multiple-parameter information can be obtained from the FCS measurement including the amplitude, characteristic diffusion time of correlation curve, and brightness of the adopted probes. The multiple-parameter change is related with physical or chemical change occurring in the probes.

In situ study of RSK2 kinase activity in a single living cell by combining single molecule spectroscopy with activity-based probes.

Protein phosphorylation is a very important regulatory mechanism in a majority of biological processes, and the determination of protein kinase activity plays a key role in the pathological study and drug development of kinase-related diseases. However, it is very challenging to in situ study endogenous protein kinase activity in a single living cell due to the shortage of in vivo efficient methods. Here, we propose a new strategy for direct determination of protein kinase activity in a single living cell by combining single molecule fluorescence correlation spectroscopy (FCS) with activity-based probes (ABPs).

Chiral ligand-induced photoluminescence intermittence difference of CdTe quantum dots.

Semiconductor quantum dots (QDs) are a new nano-material, and their unique optical properties have become a focal point of research in both academia and industry. In this study, we studied photoluminescence (PL) intermittence (or 'blinking') behaviors of individual QDs prepared with different chiral ligands by using single molecule microscopy and single molecule fluorescence correlation spectroscopy (FCS). We found that the chirality of N-isobutyryl-d/l-cysteine (D/L-NIC) as surface stabilizers significantly influences PL blinking behaviors of cadmium telluride (CdTe) QDs synthesized in aqueous solution.

Analysis of the concentrations and size distributions of cell-free DNA in schizophrenia using fluorescence correlation spectroscopy.

Cell-free DNA (cfDNA), which is primarily released following cell death, has been described and developed to serve as an effective biomarker in autoimmune diseases which may share the pathogenesis with schizophrenia. In this study, we hypothesized and explored whether the concentrations and size distributions of cfDNA are abnormal in schizophrenia. A total of 65 patients with schizophrenia (SZ), 29 patients with mood disorders (MD) and 62 matched healthy controls (HC) were included in the study.