A temperature-independent model of dual calibration standards for onsite and point-of-care quantification analyses via electrophoresis titration chip.

Electrophoresis titration chip (ETC) is a versatile tool for onsite and point-of-care quantification analyses because it affords naked-eye detection and a straightforward quantification format. However, it is vulnerable to changes in environmental temperature, which regulates the electrophoretic migration by affecting the ion mobility and the target recognition by influencing the enzyme activity. Therefore, the quantification accuracy of the ETC tests was severely compromised. Rather than using the dry bath or heating/cooling units, we proposed a facile model of dual calibration standards (DCS) to mathematically eliminate the effects of temperature on quantification accuracy. To verify our model, we deployed the ETC device at different temperatures ranging from 5 to 40 °C. We further utilized the DCS-ETC to determine the protein content and uric acid concentration in real samples outside the laboratory. All the experimental results showed that our model significantly stabilized the quantification recovery from 35.31-153.44 % to 99.38-103.44 % for protein titration; the recovery of uric acid titration is also stable at 96.25-106.42 %, suggesting the enhanced robustness of the ETC tests. Therefore, DCS-ETC is a field-deployable test that can offer reliable quantification performance without extra equipment for temperature control. We envision that it is promising to be used for onsite applications, including food safety control and disease diagnostics.

[Detection and analysis of moving reaction boundary-based electrophoresis distance using smartphone images].

Electrophoresis titration (ET) based on the moving reaction boundary (MRB) theory can detect the analyte contents in different samples by converting content signals into distance signals. However, this technique is only suitable for on-site qualitative testing, and accurate quantification relies on complex optical equipment and computers. Hence, applying this method to real-time point-of-care testing (POCT) is challenging. In this study, we developed a smartphone-based ET system based on a visual technique to achieve real-time quantitative detection. First, we developed a portable quantitative ET device that can connect to a smartphone; this device consisted of five components, namely, an ET chip, a power module, a microcontroller, a liquid crystal display screen, and a Bluetooth module. The device measured 10 cm×15 cm×2.5 cm, weighed 300 g, and was easy to hold. Thus, it is suitable for on-site testing with a run time of only 2-4 min. An assistant mobile software program was also developed to control the device and perform ET. The colored electrophoresis boundary can be captured using the smartphone camera, and quantitative detection results can be obtained in real time. Second, we proposed a quantitative algorithm based on ET channels. The software was used to recognize the boundary migration distance of three channels, a standard curve based on two given contents of the standards was established using the two-point method, and the content of the test sample was calculated. Human serum albumin (HSA) and uric acid (UA) were used as a model protein and biosample, respectively, to test the performance of the detection system. For HSA detection, different HSA solutions were mixed with a polyacrylamide gel (PAG) stock solution, phenolphthalein was added as an indicator, and sodium persulfate and tetramethyl ethylenediamine (TEMED) were used to promote polymerization to form a gel. For UA detection, agarose gel was filled into the ET channel, the UA sample, urate oxidase, and leucomalachite green were added into the anode cell and incubated for 20 min. ET was then performed. The fitting goodness (R2) values of HSA and UA were 0.9959 and 0.9935, respectively, with a linear range of 0.5-35.0 g/L and a log-linear range of 100-4000 µmol/L. The limits of detection for HSA and UA were 0.05 g/L and 50 µmol/L, respectively, and the corresponding relative standard deviations (RSDs) were not greater than 2.87% and 3.21%, respectively. These results demonstrate that the detection system has good accuracy and sensitivity. Clinical samples collected from healthy volunteers were used as target blood samples, and the developed system was used to measure serum total protein and UA levels. Serum samples from five volunteers were selected, standard curves of total serum protein and UA were established, and the test results were compared with hospital standard testing results. The relative errors for serum total protein and UA were less than 6.03% and 6.21%, respectively, and the corresponding RSDs were less than 3.72% and 5.84%, respectively. These findings verify the accuracy and reliability of the proposed detection system. The smartphone-based ET detection system introduced in this paper presents several advantages. First, it enables the portable real-time detection of total serum protein and UA. Second, compared with traditional ET strategies based on colored boundaries, it does not rely on optical detection equipment or computers to obtain quantitative detection results; as such, it can reduce the complexity of the operation and provide portability and real-time metrics. Third, the detection of two biomarkers, serum total protein and UA, is achieved on the same device, thereby improving the multitarget detection potential of the ET method. These advantages render the developed method a promising detection platform for clinical applications and real-time POCT.

Effects of Mind-Body Exercises for Osteoporosis in Older Adults: A Systematic Review and Meta-analysis of Randomized Controlled Trials.

Introduction: Osteoporosis is a major cause of fractures and even life-threatening fractures in the elderly. Mind-body exercise is a beneficial intervention to improve flexibility, control body balance and reduce pain. We aimed to evaluate the effects of physical and mental exercise on osteoporosis in the elderly. Methods: Randomized controlled trials (RCTs) focusing on mind-body exercises for osteoporosis were included. Web of Science, PubMed, Science Direct, Medline, Cochrane Library, China National Knowledge Infrastructure (CNKI), and Wanfang were searched from inception to January 2023. Outcomes included bone mineral density (BMD), bone mineral content (BMC), body balance (BB), pain, indicators of bone metabolism (BMI), lower extremity function, fearing level, and quality of life (QOL). The quality of study reporting was rated by 2 reviewers independently, and Review Manager software (version 5.3) was used for meta-analysis. Results: Thirty-nine trials with 2325 participants were included. The pooled results showed that mind-body exercises have encouraging effect on elderly people with osteoporosis, especially in aspects of BMD, BMC, QOL, improving the function of lower extremity, reducing pain and fearing level. While, dance and eight-section brocade could not improve the quality of life,or dance and eight-section brocade have no effect on BMD. Conclusions: Mind-body exercises may have potential efficacy for osteoporosis in the elderly. However, due to the poor methodological quality of the included trials, more clinical trials with precise methodological design and rigorous reporting are needed.

[Determination of human serum total protein via electrophoresis titration and capacitively coupled contactless conductivity detection].

Serum total protein refers to the sum of all proteins in the serum, and its content determination is relevant to human health monitoring and disease diagnosis. However, existing detection techniques present a number of limitations; for example, the Kjeldahl method suffers from the negative effects of interfering substances such as non-protein nitrogen (NPN). Although the electrophoresis titration (ET) method has solved interference problems to some extent, the current ET technique relies on optical detection methods, which increases the tediousness of the operation. This study addresses the challenge of accurate serum total protein detection by combining the traditional ET technique with capacitively coupled contactless conductivity detection (C4D). The research contributions of this work are multifold. First, it presents the first development of an ET-C4D detection system, which consists of six components: an ET power module, an ET chip, a C4D sensing module, a detection module, a data acquisition card, and software. The developed system can capture the conductivity of substances in the channel using the software developed by our laboratory during ET. The detection system can be used to quantify the total protein content in human serum without the addition of specific labeling reagents or using optical detection equipment, and its running time is approximately 300 s. Second, this research proposes the corresponding principle of the system. Under an electric field, ion migration results in different pH levels before and after the boundary, leading to a protein surface charge difference. The maintenance of the electrical neutrality of the substances in the detection channel is related to the protein surface charge; therefore, the ion concentration distribution of the substances in the detection channel changes as the protein surface charge varies. A plot of conductivity as a function of running time showed an "inverted clock shape", first falling and then rising. Owing to the addition of different types and concentrations of proteins, the microenvironment of the entire system changes, resulting in different changes in conductivity. Third, the performance of the detection system was tested using human serum albumin (HSA) standard protein, which was mixed with polyacrylamide gel (PAG) mother liquor, riboflavin, etc., and irradiated under ultraviolet light for 10 min to form a gel. The ET experiments were then carried out. The shape of the conductivity curve was consistent with the proposed principle, and the higher the HSA concentration, the lower the conductivity curve trough, followed by a lagged time of the trough. Quantitative analysis of the conductivity signals showed that the linear range was 0.25-3.00 g/L, with a linearity of up to 0.98. The limit of detection (LOD) was 0.01 g/L, the relative standard deviation (RSD) was 1.90%, and the relative error of the test values was <7.20%, indicating the good detection stability and sensitivity of the system. Clinical samples collected from healthy volunteers were used as target blood samples for serum total protein content measurement using our detection system. Blood samples from a volunteer were used to obtain a standard curve, and the serum samples of other four volunteers were selected for ET-C4D and biuret detection. The results showed that the relative errors between the two methods were within 4.43%, indicating the accuracy and reliability of the detection system. The advantages of the ET-C4D detection system proposed in this paper are as follows: (i) ET-C4D realizes the rapid detection of total serum protein content based on the ET technique; (ii) compared with the traditional protein ET technique, the ET-C4D method does not rely on specific labeling components or optical detection equipment, thereby reducing the complexity of the operation; and (iii) the output signal of ET-C4D can be used for quantitative analysis with excellent analytical performance and high accuracy. These merits highlight the potential of the developed system for clinical application and biochemical analysis.

[In-site electrophoretic elution of excessive fluorescein isothiocyanate from fluorescent particles in gel for image analysis].

The sensitivity, accuracy, and efficiency of fluorescent particle detection can be improved by purifying the fluorescent-dye-labeled particles. In this study, an in-site model of electrophoretic elution (EE) was developed for the facile and efficient removal of unconjugated fluorescent dyes after labeling reactions, thereby facilitating the sensitive fluorescent imaging of proteins captured by microbeads. First, bovine serum albumin (BSA) and magnetic beads (MBs) were chosen as the model protein and particles, respectively, and an MBs-BSA complex was synthesized by mixing the beads with the BSA solution. Second, excessive fluorescein isothiocyanate (FITC) was added to the EP tube with MBs-BSA suspension for the fluorescent labeling of BSA, and a labeled compound was obtained after 8-h incubation in the dark at 4 ℃. The unpurified MBs-BSAFITC was obtained by removing the supernatant, leaving 5 µL of the residual solution in the EP tube. The obtained MBs-BSAFITC solution was added to a 50-µL phosphate buffer solution (PBST, containing 0.01% Triton X-100, pH 7.4). Third, gel suspension was prepared by mixing the MBs-BSAFITC solution with the low-gelling-temperature agarose gel (10 g/L) and filled into an electrophoresis channel. To demonstrate the high efficiency of the in-site model of EE for removing excessive FITC, a 10-mm hydrogel segment was prepared using MBs-BSAFITC sandwiched between two blank hydrogels and filled into a 50-mm-long electrophoresis tube (outer diameter: 5 mm; inner diameter: 3 mm) for the EE. Subsequently, the filled channel was set in an electrophoresis device to construct the in-site EE model. The particle size of the MBs was larger than the pore size of the gel, and the fluorescent beads were physically immobilized in the gel while the excessive FITC was removed from the channel by electrophoresis. Before an EE run, the original fluorescence image of the target gel was captured using a CCD camera. After the 30-min EE (50 V, 6 mA, pH 7.4 PBS), the fluorescence image was also recorded by the CCD camera. The fluorescent images were converted to a grayscale intensity map. To simplify the calculation, a simple fluorescent image analysis method was developed. The side view of the grayscale intensity map is a two-dimensional plot of peaks. Each peak indicates a fluorescent spot at a given position along the length of the channel when the distribution density of the particles is low, and the peak value is the grayscale intensity of the fluorescent spot. The statistical peak numbers and values can be used to approximate fluorescent spots on the image. After image processing and calculations, 27.8% of the average grayscale intensity of the fluorescent spot was retained, comparing the average gray value of the bright spot before and after EE, and 97.6% of excessive FITC in the channel was cleared, obtained by calculating the decreased background fluorescence grayscale intensity after EE. The particle-to-background signal ratio (P/B ratio, PBr) increased from 1.08 to 12.2 after EE with an exposure time of 1.35 s. In addition, different exposure times were explored during the fluorescence detection. Increasing the exposure time from 1.35 to 2.35 s enhanced PBr from 12.2 to 15.5, which could effectively increase the signal-to-noise ratio. An appropriate increase in exposure time also allowed the detection of many weak fluorescent particles that were previously undetectable, indicating increased sensitivity of the fluorescence detection. The EE model has the following advantages: (i) increase in specificity by eluting FITC absorbed to the surface of beads; (ii) high efficiency in the removal of free FITC with more than 97% clearance; (iii) rapid decrease in noise in the mass hydrogel (within 30 min). This method can be used in beads/spots-based immunoassay in gel, immuno-electrophoresis, and fluorescent staining of protein/nucleic acid bands in gel electrophoresis.

Roles of acyl-CoA-binding proteins in plant reproduction.

Acyl-CoA-binding proteins (ACBPs) constitute a well-conserved family of proteins in eukaryotes that are important in stress responses and development. Past studies have shown that ACBPs are involved in maintaining, transporting and protecting acyl-CoA esters during lipid biosynthesis in plants, mammals, and yeast. ACBPs show differential expression and various binding affinities for acyl-CoA esters. Hence, ACBPs can play a crucial part in maintaining lipid homeostasis. This review summarizes the functions of ACBPs during the stages of reproduction in plants and other organisms. A comprehensive understanding on the roles of ACBPs during plant reproduction may lead to opportunities in crop improvement in agriculture.

Interactions between plant lipid-binding proteins and their ligands.

Lipids participate in diverse biological functions including signal transduction, cellular membrane biogenesis and carbon storage. Following de novo biosynthesis in the plastids, fatty acids (FAs) are transported as acyl-CoA esters to the endoplasmic reticulum where glycerol-3-phosphate undergoes a series of acyl-CoA-dependent acylation via the Kennedy pathway to form triacylglycerols for subsequent assembly into oils. Alternatively, newly synthesized FAs are incorporated into phosphatidylcholine (PC) by a PC:acyl-CoA exchange process defined as "acyl editing". Acyl-CoA-binding proteins (ACBPs) at various subcellular locations can function in lipid transfer by binding and transporting acyl-CoA esters and maintaining intracellular acyl-CoA pools. Widely distributed in the plant kingdom, ACBPs are found in all eukaryotes and some eubacteria. In both rice and Arabidopsis, six forms of ACBPs co-exist and are classified into four groups based on their functional domains. Their conserved four-helix structure facilitates interaction with acyl-CoA esters. ACBPs also interact with phospholipids as well as protein partners and function in seed oil regulation, development, pathogen defense and stress responses. Besides the ACBPs, other proteins such as the lipid transfer proteins (LTPs), annexins and lipid droplet-associated proteins are also important lipid-binding proteins. While annexins bind Ca2+ and phospholipids, LTPs transport lipid molecules including FAs, acyl-CoA esters and phospholipids.

Oxylipin signaling in salt-stressed soybean is modulated by ligand-dependent interaction of Class II acyl-CoA-binding proteins with lipoxygenase.

Plant lipoxygenases (LOXs) oxygenate linoleic and linolenic acids, creating hydroperoxy derivatives, and from these, jasmonates and other oxylipins are derived. Despite the importance of oxylipin signaling, its activation mechanism remains largely unknown. Here, we show that soybean ACYL-COA-BINDING PROTEIN3 (ACBP3) and ACBP4, two Class II acyl-CoA-binding proteins, suppressed activity of the vegetative LOX homolog VLXB by sequestering it at the endoplasmic reticulum. The ACBP4-VLXB interaction was facilitated by linoleoyl-CoA and linolenoyl-CoA, which competed with phosphatidic acid (PA) for ACBP4 binding. In salt-stressed roots, alternative splicing produced ACBP variants incapable of VLXB interaction. Overexpression of the variants enhanced LOX activity and salt tolerance in Arabidopsis and soybean hairy roots, whereas overexpressors of the native forms exhibited reciprocal phenotypes. Consistently, the differential alternative splicing pattern in two soybean genotypes coincided with their difference in salt-induced lipid peroxidation. Salt-treated soybean roots were enriched in C32:0-PA species that showed high affinity to Class II ACBPs. We conclude that PA signaling and alternative splicing suppress ligand-dependent interaction of Class II ACBPs with VLXB, thereby triggering lipid peroxidation during salt stress. Hence, our findings unveil a dual mechanism that initiates the onset of oxylipin signaling in the salinity response.

[Determination of absolute mobility and dissociation constant of lovastatin using capillary electrophoresis and empirical equation of ion mobility].

In capillary electrophoresis, determination of the basic physical and chemical properties of compounds, such as absolute mobility (m0) and dissociation constant (pKa), is of great practical significance. This is because the aforementioned properties are often used for the qualitative or quantitative analyses of the relevant compounds toward their application as potential drugs. Lovastatin is a potential drug candidate that can reduce the levels of cholesterol and low-density lipoprotein cholesterol in the blood, as well as prevent atherosclerosis and coronary heart disease. For a more convenient and rapid investigation of the properties and applications of lovastatin, it is necessary to determine its m0 and pKa values. However, existing research on capillary electrophoresis for lovastatin and other related drugs focus on their quantitative determination, and their action mechanism and functions. Unfortunately, there are very few studies aimed at the determination of the m0 and pKa values of lovastatin. Based on related studies, this paper herein proposed a novel method to determine m0 and pKa of lovastatin. The present study mainly included a calculation method and experimental verification. The calculation method was based on capillary zone electrophoresis (CZE) and the empirical formula of ion mobility. First, on the basis of the empirical formula, the calculation formula for m0 was derived from the relationship between the actual mobility (mact), effective mobility (meff) and m0. Second, for a monovalent acid (HA), according to the calculation formula for m0 part, considering the hydrogen ion concentration as the independent variable and the reciprocal of meff as the dependent variable, a straight line was obtained on the coordinate axis. From the slope of this straight line, the dissociation equilibrium constant Ka was obtained directly, and pKa was calculated easily. After the derivation of m0 and pKa in the theoretical part, the feasibility and reliability of this method were verified by using it to determine the m0 and pKa values of several organic acids and bases (barbituric acid, benzoic acid, benzylamine, phenol, and m-cresol) in the experimental part. Note that for the buffer system with pH<6.0, reverse capillary electrophoresis was used for the determination of pKa, because this technique helped shorten the migration time and facilitates the detection of analytes that could not reach the cathode. After obtaining m0 and pKa, the theoretical reference values for these parameters were obtained by PeakMaster 5.1. The experimental data were well consistent with the theoretical m0 and pKa values. The standard deviation (SDs) of m0 and pKa were less than 6.0% and 6.2%, respectively. From the correlation coefficient (R) of the linear regression equation, it was found that the linear regression lines of pKa fit well, indicating the excellent reliability of this method. Finally, with this simple and reliable method, dimethyl sulfoxide (DMSO) was used as a marker for electroosmotic flow to determine the m0 and pKa values of lovastatin (-1.70×10-8 m2/(V·s) and 9.00, respectively). This method is suitable for the determination of m0 and pKa of acidic and basic analytes. The method has high accuracy and is expected to play an indispensable role in drug analysis.

[Speculation of hemoglobin A3 peak position in clinical cation exchange high performance liquid chromatogram of the diabetic blood sample with microarray isoelectric focusing].

Hemoglobin A1c (HbA1c) is a major component of glycated hemoglobin in human red blood cells. It has been proven to be a significant biomarker for the diagnosis of diabetes; its content in fresh red cells in diabetes blood reflects the average level of blood glucose over the previous three months. Thus, HbA1c level has been used for the assessment of long-term glycemic control in diabetes; the level of 6.5% HbA1c has been certified as a critical cut-off for the diabetes diagnosis. The current commonly used method for HbA1c quantification is based on cation-exchange high performance liquid chromatography (CX-HPLC). The method has advantages such as high stability, rapidity, and automation, but there are still some unidentified peaks of Hb species in CX-HPLC (VARIANT Ⅱ system); in particular, the presence of HbA3 (a glutathiolated Hb) affects the accurate determination of HbA1c. HbA3 is usually present in healthy adult blood samples at 2%-4%, but the concentration of HbA3 increases due to the protection of erythrocytes from oxidation, resulting in decreased HbA1c. However, the relative location of the HbA3 peak in the CX-HPLC clinical chromatogram has not been established. To address this issue, we extracted Hb species from fresh blood samples obtained from a hospital in an anaerobic environment to avoid possible redox reactions of Hb and glutathione. After the extraction, the Hb samples were analyzed using two methods: a low-resolution CX-HPLC (5/50 mm column) currently used for diabetes diagnosis and a high-resolution cationic exchange HPLC (Mono-S 5/50 mm column), to identify the peak corresponding to HbA3. The CX-HPLC analysis of fresh blood samples indicated that the unknown peak P3 located between HbA1c and HbA0 peaks corresponded to the HbA3 peak between HbA1c and HbA0 in the Mono-S-HPLC. Microarray isoelectric focusing (IEF) was used for the micro-preparation of HbA3, HbA1c, and HbA0 in healthy blood samples; then, the micro-prepared species of HbA3, HbA1c, and HbA0 were individually identified via Mono-S-HPLC. The results of the CX-HPLC, Mono-S-HPLC, and microarray IEF experiments indicated that the P3 peak might correspond to HbA3. To confirm this, glutathiolated Hb samples were synthesized via acetylphenylhydrazine and analyzed using both the Mono-S- and CX-HPLC systems. The results showed that the content of both glutaminated hemoglobin of HbA3 in Mono-S-HPLC and P3 in CX-HPLC increased, implying the peak of P3 with the retention time of 1.50 min in CX-HPLC was the peak corresponding to HbA3 in Mono-S-HPLC and microarray IEF. Based on the above experiments and our previous results, the influence of HbA3 on both the analysis of HbA1c in blood samples and the diabetes diagnosis needs to be considered and discussed. The study results are significant for the tentative assignment of peak P3 and for offering more information on diabetes diagnosis using CX-HPLC in the clinical setting.

In silico Analysis of Acyl-CoA-Binding Protein Expression in Soybean.

Plant acyl-CoA-binding proteins (ACBPs) form a highly conserved protein family that binds to acyl-CoA esters as well as other lipid and protein interactors to function in developmental and stress responses. This protein family had been extensively studied in non-leguminous species such as Arabidopsis thaliana (thale cress), Oryza sativa (rice), and Brassica napus (oilseed rape). However, the characterization of soybean (Glycine max) ACBPs, designated GmACBPs, has remained unreported although this legume is a globally important crop cultivated for its high oil and protein content, and plays a significant role in the food and chemical industries. In this study, 11 members of the GmACBP family from four classes, comprising Class I (small), Class II (ankyrin repeats), Class III (large), and Class IV (kelch motif), were identified. For each class, more than one copy occurred and their domain architecture including the acyl-CoA-binding domain was compared with Arabidopsis and rice. The expression profile, tertiary structure and subcellular localization of each GmACBP were predicted, and the similarities and differences between GmACBPs and other plant ACBPs were deduced. A potential role for some Class III GmACBPs in nodulation, not previously encountered in non-leguminous ACBPs, has emerged. Interestingly, the sole member of Class III ACBP in each of non-leguminous Arabidopsis and rice had been previously identified in plant-pathogen interactions. As plant ACBPs are known to play important roles in development and responses to abiotic and biotic stresses, the in silico expression profiles on GmACBPs, gathered from data mining of RNA-sequencing and microarray analyses, will lay the foundation for future studies in their applications in biotechnology.

Investigations of Lipid Binding to Acyl-CoA-Binding Proteins (ACBP) Using Isothermal Titration Calorimetry (ITC).

Isothermal titration calorimetry (ITC) is a quantitative, biophysical method to investigate intermolecular binding between biomolecules by directly measuring the heat exchange in the binding reaction. The assay is carried out in solution when the molecules interact in vitro. This allows to determine values for binding affinity (Kd), binding stoichiometry (n), as well as changes in Gibbs free energy (ΔG), entropy (ΔS), and enthalpy (ΔH). This method also addresses the kinetics of enzymatic reactions for a substrate during conversion to a product. ITC has been used to study the interactions between proteins and ligands such as those of acyl-CoA-binding proteins (ACBPs) and acyl-CoA thioesters or ACBPs with protein partners. ITC has also been used in investigating interactions between antiserum and antigen, as well as those involving RNA and DNA and other macromolecules. We describe the methods used to isolate and purify a recombinant rice ACBP (OsACBP) for ITC. To study OsACBP binding to long-chain acyl-CoA thioesters, a microcalorimeter was used at 30 °C, and the ligand (acyl-CoA thioesters or a protein partner in the first cell), was mixed with the ACBP protein solution in a second cell, for more than 40 min comprising 20 injections. Subsequently, the binding parameters including the heat-release data were analyzed and various thermodynamic parameters were calculated.

Model, Simulation, and Experiments on Moving Exchange Boundary via Ligand and Quantum Dots in Chip Electrophoresis.

Herein, the quench model of the moving exchange boundary (MEB) was first created via a ligand of 5,5'-dithiobis(2-nitro-benzoic acid) (DTNB) and group of 3-mercaptopropionic acid (MPA) capped on QDs, and then the recovery model was formed via MPA and 2-nitro-5-thiobenzoic acid (TNB) capped on QDs. The theory on MEB dynamics and width was developed based on the two reversible models, the simulation was conducted for the illumination of MEB, and the protocol was described for the MEB runs. The experiments revealed that (i) the quench model could be created via DTNB and MPA capped on QDs and the recovery one could be in situ formed via MPA and TNB capped on QDs, showing the feasibility of MEB models; (ii) the simulations on MEB dynamics and width were in coincidence with the theoretic predictions, showing the validity of two models; and (iii) the experiments demonstrated the validity of models, predictions, and simulations. The models and theory have potential for development of a biosensor, nanoparticle characterization, separation science, and an affinity assay of ligand-QDs.

Crystal structure of the rice acyl-CoA-binding protein OsACBP2 in complex with C18:3-CoA reveals a novel pattern of binding to acyl-CoA esters.

Acyl-CoA-binding proteins (ACBPs) are a family of proteins that bind acyl-CoA esters at a conserved acyl-CoA-binding domain. ACBPs maintain intracellular acyl-CoA pools to regulate lipid metabolism. Here, we report on the structure of rice OsACBP2 in complex with C18:3-CoA ester. The residues Y33, K34 and K56 of OsACBP2 play a crucial role in binding the CoA group, while residues N23, L27, K52 and Y55 in one molecule of OsACBP2 cooperate with L27, L28, A59 and A62 from another anchoring the fatty acyl group. Multiangle light scattering assays indicate that OsACBP2 binds C18:3-CoA as a monomer. The first complex structure of a plant ACBP binding with C18:3-CoA is therefore presented, providing a novel model for the interaction between an acyl-CoA ester and the acyl-CoA-binding domain(s).

A novel neoantigen discovery approach based on chromatin high order conformation.

BACKGROUND: High-throughput sequencing technology has yielded reliable and ultra-fast sequencing for DNA and RNA. For tumor cells of cancer patients, when combining the results of DNA and RNA sequencing, one can identify potential neoantigens that stimulate the immune response of the T cell. However, when the somatic mutations are abundant, it is computationally challenging to efficiently prioritize the identified neoantigen candidates according to their ability of activating the T cell immuno-response. METHODS: Numerous prioritization or prediction approaches have been proposed to address this issue but none of them considers the original DNA loci of the neoantigens from the perspective of 3D genome. Based on our previous discoveries, we propose to investigate the distribution of neoantigens with different immunogenicity abilities in 3D genome and propose to adopt this important information into neoantigen prediction. RESULTS: We retrospect the DNA origins of the immuno-positive and immuno-negative neoantigens in the context of 3D genome and discovered that DNA loci of the immuno-positive neoantigens and immuno-negative neoantigens have very different distribution pattern. Specifically, comparing to the background 3D genome, DNA loci of the immuno-positive neoantigens tend to locate at specific regions in the 3D genome. We thus used this information into neoantigen prediction and demonstrated the effectiveness of this approach. CONCLUSION: We believe that the 3D genome information will help to increase the precision of neoantigen prioritization and discovery and eventually benefit precision and personalized medicine in cancer immunotherapy.

DeepAntigen: a novel method for neoantigen prioritization via 3D genome and deep sparse learning.

MOTIVATION: The mutations of cancers can encode the seeds of their own destruction, in the form of T-cell recognizable immunogenic peptides, also known as neoantigens. It is computationally challenging, however, to accurately prioritize the potential neoantigen candidates according to their ability of activating the T-cell immunoresponse, especially when the somatic mutations are abundant. Although a few neoantigen prioritization methods have been proposed to address this issue, advanced machine learning model that is specifically designed to tackle this problem is still lacking. Moreover, none of the existing methods considers the original DNA loci of the neoantigens in the perspective of 3D genome which may provide key information for inferring neoantigens' immunogenicity. RESULTS: In this study, we discovered that DNA loci of the immunopositive and immunonegative MHC-I neoantigens have distinct spatial distribution patterns across the genome. We therefore used the 3D genome information along with an ensemble pMHC-I coding strategy, and developed a group feature selection-based deep sparse neural network model (DNN-GFS) that is optimized for neoantigen prioritization. DNN-GFS demonstrated increased neoantigen prioritization power comparing to existing sequence-based approaches. We also developed a webserver named deepAntigen (http://yishi.sjtu.edu.cn/deepAntigen) that implements the DNN-GFS as well as other machine learning methods. We believe that this work provides a new perspective toward more accurate neoantigen prediction which eventually contribute to personalized cancer immunotherapy. AVAILABILITY AND IMPLEMENTATION: Data and implementation are available on webserver: http://yishi.sjtu.edu.cn/deepAntigen. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Arabidopsis cytosolic acyl-CoA-binding proteins function in determining seed oil composition.

As plant seed oils provide animals with essential fatty acids (FAs), genes that regulate plant lipid metabolism have been used in genetic manipulation to improve dietary seed oil composition and benefit human health. Herein, the Arabidopsis thaliana cytosolic acyl-CoA-binding proteins (AtACBPs), AtACBP4, AtACBP5, and AtACBP6 were shown to play a role in determining seed oil content by analysis of atacbp (atacbp4, atacbp5, atacbp6, atacbp4atacbp5, atacbp4atacbp6, atacbp5atacbp6, and atacbp4atacbp5atacbp6) seed oil content in comparison with the Col-0 wild type (WT). Triacylglycerol (TAG) composition in electrospray ionization-mass spectrometer (ESI-MS) analysis on atacbp6 seed oil showed a reduction (-50%) of C58-TAGs in comparison with the WT. Investigations on fatty acid composition of atacbp mutants indicated that 18:2-FA accumulated in atacbp6 and 18:3-FA in atacbp4, both at the expense of 20:1-FA. As TAG composition can be modified by acyl editing through phosphatidylcholines (PC) and lysophosphatidylcholines (LPC), total PC and LPC content in atacbp6 mature seeds was determined and ESI-MS analysis revealed that LPC had increased (+300%) at the expense of PC. Among all the 14 tested PC species, all (34:1-, 34:2-, 34:3-, 34:4-, 34:5-, 34:6-, 36:2-, 36:3-, 36:5-, 36:6-, 38:2-, 38:3-, and 38:4-PCs) but 36:4-PC were lower in atacbp6 than the WT. In contrast, all LPC species (16:0-, 18:1-, 18:2-, 18:3-, and 20:1-LPC) examined were elevated in atacbp6. LPC abundance also increased in atacbp4atacbp5, but not atacbp4 and atacbp5. Interestingly, when LPC composition in atacbp4atacbp5 was compared with atacbp4 and atacbp5, significant differences were observed between atacbp4atacbp5 and each single mutant, implying that AtACBP4 and AtACBP5 play combinatory roles by affecting LPC (but not PC) biosynthesis. Furthermore, PC-related genes such as those encoding acyl-CoA:lysophphosphatidylcholine acyltransferase (LPCAT1) and phospholipase A2 alpha (PLA2α) were upregulated in atacbp6 developing seeds. A model on the role of AtACBP6 in modulating TAG through regulating LPCAT1 and PLA2α expression is proposed. Taken together, cytosolic AtACBPs appear to affect unsaturated TAG content and are good candidates for engineering oil crops to enhance seed oil composition.

The overexpression of rice ACYL-CoA-BINDING PROTEIN2 increases grain size and bran oil content in transgenic rice.

As Oryza sativa (rice) seeds represent food for over three billion people worldwide, the identification of genes that enhance grain size and composition is much desired. Past reports have indicated that Arabidopsis thaliana acyl-CoA-binding proteins (ACBPs) are important in seed development but did not affect seed size. Herein, rice OsACBP2 was demonstrated not only to play a role in seed development and germination, but also to influence grain size. OsACBP2 mRNA accumulated in embryos and endosperm of germinating seeds in qRT-PCR analysis, while ß-glucuronidase (GUS) assays on OsACBP2pro::GUS rice transformants showed GUS expression in embryos, as well as the scutellum and aleurone layer of germinating seeds. Deletion analysis of the OsACBP2 5'-flanking region revealed five copies of the seed cis-element, Skn-I-like motif (-1486/-1482, -956/-952, -939/-935, -826/-822, and -766/-762), and the removal of any adversely affected expression in seeds, thereby providing a molecular basis for OsACBP2 expression in seeds. When OsACBP2 function was investigated using osacbp2 mutants and transgenic rice overexpressing OsACBP2 (OsACBP2-OE), osacbp2 was retarded in germination, while OsACBP2-OEs performed better than the wild-type and vector-transformed controls, in germination, seedling growth, grain size and grain weight. Transmission electron microscopy of OsACBP2-OE mature seeds revealed an accumulation of oil bodies in the scutellum cells, while confocal laser scanning microscopy indicated oil accumulation in OsACBP2-OE aleurone tissues. Correspondingly, OsACBP2-OE seeds showed gain in triacylglycerols and long-chain fatty acids over the vector-transformed control. As dietary rice bran contains beneficial bioactive components, OsACBP2 appears to be a promising candidate for enriching seed nutritional value.

[A literature analysis of power frequency electric field testing data].

OBJECTIVE: To analyze the literature on power frequency electric field testing data and to propose views and suggestions for current testing. METHODS: The literature on power frequency electric field testing data published in the previous years was searched to identify 306 articles involving 193 valid testing data. Mann-Whitney test and Wilcoxon W test were used for analyzing the testing data. RESULTS: The classification of data was carried out according to one quarter of occupational exposure limit (1.25 kV/m), one half of the exposure limit (2.5 kV/m), and the exposure limit (5 kV/m). The structure of testing data showed a significant difference between the non-power facility group and the power facility group (P<0.05). CONCLUSION: As occupational hazard factors, the radiation exposure from power frequency electric field is extensive. However, the power frequency electric field testing data in actual workplaces except high-voltage power facilities are far less than the occupational exposure limit with little harmfulness. There is a phenomenon of excessive testing at present.

Turbulence-driven bootstrap current in low-collisionality tokamaks.

Neoclassical bootstrap current is expected to provide a significant fraction of the equilibrium plasma current in tokamak reactors. Here we report a novel mechanism through which a bootstrap current may be driven even in a collisionless plasma. In analogy with the neoclassical mechanism, in which the collisional equilibrium established between trapped and passing electrons produces a steady state current, we show that resonant scattering of electrons by drift wave microturbulence provides an additional means of determining the equilibrium between trapped and passing electrons and thus driving a bootstrap current. Employing a linearized Fokker-Planck collision operator, the plasma current in the presence of both collisions and resonant electron scattering is computed, allowing for the relative strength of these two mechanisms to be quantified as a function of collisionality and fluctuation amplitude.