Comprehensive performance evaluation of ligand-binding assay-LC-MS/MS method for co-dosed monoclonal anti-SARS-CoV-2 antibodies (AZD7442).

Aims: AZD7442 is a combination SARS-CoV-2 therapy comprising two co-dosed monoclonal antibodies. Materials & methods: The authors validated a hybrid ligand-binding assay-LC-MS/MS method for pharmacokinetic assessment of AZD7442 in human serum with nominal concentration range of each analyte of 0.300-30.0 µg/ml. Results: Validation results met current regulatory acceptance criteria. The validated method supported three clinical trials that spanned more than 17 months and ≥720 analytical runs (∼30,000 samples and ∼3000 incurred sample reanalyses per analyte). The data generated supported multiple health authority interactions, across the globe. AZD7442 (EVUSHELD) was approved in 12 countries for pre-exposure prophylaxis of COVID-19. Conclusion: The results reported here demonstrate the robust, high-throughput capability of the hybrid ligand-binding assay-LC-MS/MS approach being employed to support-next generation versions of EVUSHELD, AZD3152.

The measurement of antibodies in human body fluids (e.g., blood, serum) has historically been tied to laboratory tests that may face operational limitations, including susceptibility to interference from other blood components and a reliance on unique reagents that can take months to produce. As such, there is a pursuit of alternative analytical methods to more accurately detect and measure antibody drugs from complex matrices. In the method, the authors describe different techniques that once combined were used to capture, separate, filter, fragment and then detect and measure the co-dosed antibody drugs. This method has been validated in accordance with current health authority guidelines and has been used to support three clinical trials that spanned more than 17 months; that is, the validated method was used to analyze nearly 30,000 serum samples from more than 2000 patients. Collectively, the results reported here demonstrate the robustness and high-throughput capability of this analytical approach.

Sleep deprivation alters pubertal timing in humans and rats: the role of the gut microbiome.

STUDY OBJECTIVES: Evidence implied that sleeping duration is associated with the timing of puberty and that sleep deprivation triggers early pubertal onset in adolescents. Sleep deprivation can affect metabolic changes and gut microbiota composition. This study investigated the effects of sleep deprivation on pubertal onset and gut microbiota composition in animal models and a human cohort. METHODS: This study comprised 459 boys and 959 girls from the Taiwan Pubertal Longitudinal Study. Sleep duration was evaluated using the self-report Pittsburgh Sleep Quality Index questionnaire. Early sexual maturation was defined by pediatric endocrinologist assessments. Mediation analyses were done to examine the association between sleep parameters, obesity, and early sexual maturation. Besides, Sprague Dawley juvenile rats were exposed to 4 weeks of chronic sleep deprivation. Vaginal opening (VO) and preputial separation (PS) were observed every morning to determine pubertal onset in female and male rats. RESULTS: The sleep-deprived juvenile rats in the sleep-deprived-female (SDF) and sleep-deprived-male (SDM) groups experienced delayed VO (mean VO days: 33 days in control; 35 days in SDF; p-value < 0.05) and PS (mean PS days: 42 days in control; 45 days in SDM; p-value < 0.05), respectively. Relative to their non-sleep-deprived counterparts, the sleep-deprived juvenile rats exhibited lower body weight and body fat percentage. Significant differences in relative bacterial abundance at genus levels and decreased fecal short-chain-fatty-acid levels were identified in both the SDF and SDM groups. In the human cohort, insufficient sleep increased the risk of early sexual maturation, particularly in girls (OR, 1.44; 95% CI: 1.09 to 1.89; p-value < 0.01). Insufficient sleep also indirectly affected early sexual maturation in girls, with obesity serving as the mediator. CONCLUSIONS: Overall, sleep deprivation altered the timing of puberty in both animal and human models but in different directions. In the rat model, sleep deprivation delayed the pubertal onset in juvenile rats through gut dysbiosis and metabolic changes, leading to a low body weight and body fat percentage. In the human model, sleep deprivation led to fat accumulation, causing obesity in girls, which increased the risk of early puberty.

Dual-probe ligation without PCR for fluorescent sandwich assay of EGFR nucleotide variants in magnetic gene capture platform.

A simple, rapid, and highly efficient fluorescent detection technique without PCR through dual-probe ligation with the genetic capture of magnetic beads and reported probe was developed for determination of epidermal growth factor receptor (EGFR) gene exon 19 deletions. The EGFR exon 19 deletion mutation makes up 48% of all mutations associated with anti-tyrosine kinase inhibition sensitivity, and thus, the EGFR nucleotide variant is very important in clinical diagnosis. In this approach, the dual-probe ligation was designed to target exon 19 deletion. The magnetic genetic captured system was then applied to capture the successful dual-probe ligation. Thereafter, a reporter probe which is coupled with 6-fluorescein amidite (6-FAM) was introduced to hybridize with dual-probe ligation product on the surface of streptavidin magnetic beads, and finally, the supernatant was taken for fluorescence measurements for distinguishing mutant types from wild types. After optimization (the RSD of the fluorescent intensity was less than 4.5% (n = 3) under the optimal condition), 20 blind DNA samples from the population were analyzed by this technique and further confirmed by direct sequencing. The results of our assay matched to those from direct sequencing data, evidencing that the developed method is accurate and successful. These 20 blind DNA samples were diagnosed as wild and then spiked with different percentages of the mutant gene to quantify the ratio of the wild and mutant genes. This strategy was also successfully applied to quantify the ratio of the wild and mutant genes with good linearity at the λex/λem of 480 nm/520 nm (r = 0.996), and the limit of detection reached 1.0% mutant type. This simple fluorescent detection of nucleotide variants shows its potential to be considered a tool in biological and clinical diagnosis. Importantly, this strategy offers a universal detection capability for any kind of mutation (point, deletion, insertion, or substitution) in a gene of interest.

One Single Tube Reaction of Aptasensor-Based Magnetic Sensing System for Selective Fluorescent Detection of VEGF in Plasma.

In this study, a simple, easy and convenient fluorescent sensing system for the detection of the vascular endothelial growth factor (VEGF) based on VEGF aptamers, aptamer-complementary fluorescence-labeled probe and streptavidin magnetic beads was developed in one single tube. The VEGF is the most important biomarker in cancer, and it is investigated that the serum VEGF level varied according to the different types and courses of cancers. Hence, efficient quantification of VEGF is able to improve the accuracy of cancer diagnoses and the precision of disease surveillance. In this research, the VEGF aptamer was designed to be able to bind with the VEGF by forming G-quadruplex secondary structures; then, the magnetic beads would capture the non-binding aptamers due to non-steric interference; and finally, the fluorescence-labeled probes were hybridized with the aptamers captured by the magnetic beads. Therefore, the fluorescent intensity in the supernatant would specifically reflect the present VEGF. After an overall optimization, the optimal conditions for the detection of VEGF were as followed, KCl, 50 µM; pH 7.0; aptamer, 0.1 µM; and magnetic beads, 10 µL (4 µg/µL). The VEGF could be well quantified within a range of 0.2-2.0 ng/mL in plasma, and the calibration curve possessed a good linearity (y = 1.0391x + 0.5471, r = 0.998). The detection limit (LOD) was calculated to be 0.0445 ng/mL according to the formula (LOD = 3.3 × σ/S). The specificity of this method was also investigated under the appearance of many other serum proteins, and the data showed good specificity in this aptasensor-based magnetic sensing system. This strategy provided a simple, sensitive and selective biosensing platform for the detection of serum VEGF. Finally, it was expected that this detection technique can be used to promote more clinical applications.

Copper nanoclusters on specific-primer PCR fragments with magnetic capture for the label-free fluorescent sensing of the T315I single nucleotide variant in the BCR-ABL1 gene.

In this study, a simple and facile procedure using the all or none formation of double-stranded DNA-templated copper nanoclusters on specific-primer PCR fragments was designed to fluorescently identify the T315I single nucleotide variant on the BCR-ABL1 gene. Chronic myeloid leukaemia (CML), a disease caused by the BCR-ABL1 fusion of tyrosine kinase, is well known for the T315I mutation that causes tyrosine kinase inhibitors (TKIs) to be resisted due to the alternative structure of the drug-binding site. Therefore, it is an important single nucleotide variant for clinical detection. In this study, only specific functional primers and the digestion of the wild genotype from the T315I mutation site with specific restriction enzymes were designed, and the different digested products could then be captured using magnetic beads. The final products would allow for fluorescent sensing via the all or none formation of double-stranded DNA-templated copper nanoclusters for the detection of the T315I mutation. This study has been successfully applied for identifying wild and mutant homozygotes and the mutant/wild heterozygote of the T315I mutation. It is expected that this analytical system can serve as a tool for the clinical diagnosis of T315I mutations and be applied to real samples of CML patients in the future.

Fluorescent aptasensor based on conformational switch-induced hybridization for facile detection of ß-amyloid oligomers.

Aß oligomers (AßO) are a dominant biomarker for early Alzheimer's disease diagnosis. A fluorescent aptasensor coupled with conformational switch-induced hybridization was established to detect AßO. The fluorescent aptasensor is based on the interaction of fluorophore-labeled AßO-specific aptamer (FAM-Apt) against its partly complementary DNA sequence on the surface of magnetic beads (cDNA-MBs). Once the FAM-Apt binds to AßO, the conformational switch of FAM-Apt increases the tendency to be captured by cDNA-MBs. This causes a descending fluorescence of supernatant, which can be utilized to determine the levels of AßO. Thus, the base-pair matching above 12 between FAM-Apt and cDNA-MBs with increasing hybridizing free energies reached the ascending fluorescent signal equilibrium. The optimized aptasensor showed linearity from 1.7 ng mL-1 to 85.1 (R = 0.9977) with good recoveries (79.27-109.17%) in plasma. Furthermore, the established aptasensor possesses rational selectivity in the presence of monomeric Aß, fibrotic Aß, and interferences. Therefore, the developed aptasensor is capable of quantifying AßO in human plasma and possesses the potential to apply in clinical cases.

A chemometric experimental design with three-step stacking capillary electrophoresis for analysis of five tobacco-specific nitrosamines in cigarette products.

Tobacco-specific nitrosamines (TSNAs) as carcinogens endanger our health and life from cigarette products. However, the safe range of TSNAs levels in commercial cigarette products has not yet been established. For the purpose of safety and supervision, a three-step stacking approach including field amplified sample injection (FASI), sweeping, and analyte focusing by micelle collapse (AFMC), was developed for the simultaneous determination of five TSNAs levels in cigarette products. This approach also involved aspects of chemometric experimental design, including fractional factorial design and central composite design. After the multilevel optimization of the experimental design, the five TSNAs were well separated. The LOD (S/N = 3) values of the N´-nitrosonornicotine (NNN), N´-nitrosoanatabine (NAT), N´-nitrosoanabasine (NAB), 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in the FASI-sweeping-AFMC CE approach were 1.000 ng/mL, 0.500 ng/mL, 0.125 ng/mL, 1.000 ng/mL, and 0.500 ng/mL respectively. The results of relative standard deviation (RSD) and relative error (RE) were all less than 3.35%, demonstrating good precision and accuracy. Finally, this novel approach was further applied to monitor three commercial cigarette products, and a range of 250.1-336.6 ng/g for NNN, 481.6-526.7 ng/g for NAT, 82.2-247.6 ng/g for NAB, 167.7-473.7 ng/g for NNAL, and 39.4-246.7 ng/g for NNK could be observed among these. Based on these results, the novel CE stacking strategy was successfully applied for the analysis of five TSNAs levels in cigarette products and could serve as a tool for assays of quality control of nitrosamines.

A three-step stacking capillary electrophoresis of field-amplified sample injection, sweeping, and micellar collapse for determination of dabigatran and its active metabolite in human plasma.

A three-step stacking capillary electrophoresis (CE) composed of field-amplified sample injection, sweeping, and analyte focusing by micellar collapse (FASI-sweeping-AFMC) was developed to determine dabigatran (D) and its major active metabolite, dabigatran acyl-beta-d-glucuronide (DAG), in human plasma. After optimization and validation, this novel approach was further applied to monitor 5 real samples, and the 25.2-186.8 ng mL-1 D could be observed among those. Based on these results, the novel CE stacking strategy was successfully applied for the analysis of D and DAG in human plasma and could be served as a tool for clinical assays.

Integration of Deep Learning Network and Robot Arm System for Rim Defect Inspection Application.

Automated inspection has proven to be the most effective approach to maintaining quality in industrial-scale manufacturing. This study employed the eye-in-hand architecture in conjunction with deep learning and convolutional neural networks to automate the detection of defects in forged aluminum rims for electric vehicles. RobotStudio software was used to simulate the environment and path trajectory for a camera installed on an ABB robot arm to capture 3D images of the rims. Four types of surface defects were examined: (1) dirt spots, (2) paint stains, (3) scratches, and (4) dents. Generative adversarial network (GAN) and deep convolutional generative adversarial networks (DCGAN) were used to generate additional images to expand the depth of the training dataset. We also developed a graphical user interface and software system to mark patterns associated with defects in the images. The defect detection algorithm based on YOLO algorithms made it possible to obtain results more quickly and with higher mean average precision (mAP) than that of existing methods. Experiment results demonstrated the accuracy and efficiency of the proposed system. Our developed system has been shown to be a helpful rim defective detection system for industrial applications.

Identifiable universal fluorescent multiplex PCR equipped with capillary electrophoresis for genotyping of exons 1 to 5 in human red and green pigment genes.

Congenital red and green color blindness is the most X-linked recessive disorder in humans caused by deletions or gross structural rearrangements of the visual pigment gene array that lead to altered the functions of visual pigments in their retina differ from normal. The incidence is about 7-10% in male and close association of X-linked recessive disorders (such as: hemophilia A, hemophilia B, duchenne muscular dystrophy). However, the traditional genetic analysis methods are time-consuming and low-efficiencies. Therefore, the purpose of the study is to develop a rapid method for genotyping of red and green pigment genes. We describe herein the first method for simultaneous evaluation of ten exons in the red and green pigment genes for genetic analysis. A forward specific primers with identifiable universal fluorescent multiplex PCR (FSIUFM-PCR) method utilized one universal primer (containing two universal non-human sequences) and forward specific primers in the multiplex PCR reaction system for simultaneously fluorescent labeling of eleven gene fragments (ten exons in red and green pigment genes and one internal standard). All the PCR products were analyzed on capillary electrophoresis with short-end injection, which had the advantage of high resolution and rapid separation. Of all 80 detected individuals, 7 subjects with color vision deficiencies (including 3 subjects only had red exons 1-5, 4 subjects had a specific red-green or green-red hybrid gene and 73 subjects with normal color vision). All genotyping results showed good agreement with DNA sequencing data. This method provided a better potential technique for genotyping and identifying of red and green pigment genes. In addition, FSIUFM-PCR method will be useful in many fields, such as diagnosis of diseases, analysis of polymorphisms and quantitative assay.

Stable Luminescent Poly(Allylaminehydrochloride)-Templated Copper Nanoclusters for Selectively Turn-Off Sensing of Deferasirox in ß-Thalassemia Plasma.

Highly stable and facile one-pot copper nanoclusters (Cu NCs) coated with poly(allylamine hydrochloride) (PAH) have been synthesized for selectively sensing deferasirox (DFX) in ß-thalassemia plasma. DFX is an important drug used for treating iron overloading in ß-thalassemia, but needs to be monitored due to certain toxicity. In this study, the PAH-Cu NCs showed highly stable fluorescence with emission wavelengths at 450 nm. The DFX specifically interacted with the copper nanocluster to turn off the fluorescence of the PAH-Cu NCs, and could be selectively quantified through the fluorescence quenching effect. The linear range of DFX in plasma analyzed by PAH-Cu NCs was 1.0-100.0 µg/mL (r = 0.985). The relative standard deviation (RSD) and relative error (RE) were lower than 6.51% and 7.57%, respectively, showing excellent reproducibility of PAH-Cu NCs for sensing DFX in plasma. This method was also successfully applied for an analysis of three clinical plasma samples from ß-thalassemia patients taking DFX. The data presented high similarity with that obtained through a capillary electrophoresis method. According to the results, the PAH-Cu NCs could be used as a tool for clinically sensing DFX in human plasma for clinical surveys.

Effect of Probiotic Supplementation on Newborn Birth Weight for Mother with Gestational Diabetes Mellitus or Overweight/Obesity: A Systematic Review and Meta-Analysis.

High birth weight indicates the future risk of obesity and increased fat mass in childhood. Maternal gestational diabetes mellitus (GDM) or overweight are powerful predictors of high birth weight. Studies on probiotic supplementation during pregnancy have reported its benefits in modulating gut microbiota composition and improving glucose and lipid metabolism in pregnant women. Therefore, probiotic intervention during pregnancy was proposed to interrupt the transmission of obesity from mothers to newborns. Thus, we performed a meta-analysis to investigate the effect of probiotic intervention in pregnant women with GDM or overweight on newborn birth weight. We searched PubMed, EMBASE, Cochrane Library, and Web of Science databases up to 18 December 2019. Randomized controlled trials (RCTs) comparing pregnant women with GDM or overweight who received probiotic intervention during pregnancy with those receiving placebo were eligible for the analysis. Newborn birth weights were pooled to calculate the mean difference with a 95% confidence interval (CI). Two reviewers assessed the trial quality and extracted data independently. Seven RCTs involving 1093 participants were included in the analysis. Compared with the placebo, probiotics had little effect on newborn birth weight of pregnant women with GDM or overweight (mean difference = -10.27, 95% CI = -90.17 to 69.63, p = 0.801). The subgroup analysis revealed that probiotic intake by women with GDM decreased newborn birth weight, whereas probiotic intake by obese pregnant women increased newborn birth weight. Thus, no evidence indicates that probiotic intake by pregnant women with GDM or overweight can control newborn birth weight.

Molecular inversion probe-rolling circle amplification with single-strand poly-T luminescent copper nanoclusters for fluorescent detection of single-nucleotide variant of SMN gene in diagnosis of spinal muscular atrophy.

In this study, a simple fluorescent detection of survival motor neuron gene (SMN) in diagnosis of spinal muscular atrophy (SMA) based on nucleic acid amplification test and the poly-T luminescent copper nanoclusters (CuNCs) was established. SMA is a severely genetic diseases to cause infant death in clinical, and detection of SMN gene is a powerful tool for pre- and postnatal diagnosis of this disease. This study utilized the molecular inversion probe for recognition of nucleotide variant between SMN1 (c.840 C) and SMN2 (c.840 C  >  T) genes, and rolling circle amplification with a universal primer for production of poly-T single-strand DNA. Finally, the fluorescent CuNCs were formed on the poly-T single-strand DNA template with addition of CuSO4 and sodium ascorbate. The fluorescence of CuNCs was only detected in the samples with the presence of SMN1 gene controlling the disease of SMA. After optimization of experimental conditions, this highly efficient method was performed under 50 °C for DNA ligation temperature by using 2U Ampligase, 3 h for rolling circle amplification, and the formation of the CuNCs by mixing 500 µM Cu2+ and 4 mM sodium ascorbate. Additionally, this highly efficient method was successfully applied for 65 clinical DNA samples, including 4 SMA patients, 4 carriers and 57 wild individuals. This label-free detection strategy has the own potential to not only be a general method for detection of SMN1 gene in diagnosis of SMA disease, but also served as a tool for detection of other single nucleotide polymorphisms or nucleotide variants in genetic analysis through designing the different sensing probes.

Microwave assisted synthesis of negative-charge carbon dots with potential antibacterial activity against multi-drug resistant bacteria.

In this research, negative-charge carbon dots (CDs) were synthesized in one-step using a microwave and found to have potential antibacterial ability against multi-drug resistant bacteria. The CDs were synthesized by using citric acid and urea as precursors, and characterized by FT-IR, TEM and fluorescence spectrophotometry. The average size of CDs was about 2.5 nm, and the ζ potential was -11.06 mV. In the following antibacterial activity test, time-killing curve experiments and colony-forming assay were carried out to determine the minimum bactericidal concentration (MBC) and minimum inhibitory concentration (MIC) of the CDs against methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-intermediate Staphylococcus aureus (VISA). The data showed the MBC of the CDs against MRSA is 2.5 mg mL-1, and the MIC of the CDs against MRSA is 0.63 mg mL-1; the MBC of the CDs against VISA is 1.25 mg mL-1, and the MIC of the CDs against VISA is 0.63 mg mL-1. The results demonstrated that the negative-charge CDs have potential against multi-drug resistant Staphylococcus aureus (S. aureus), and may serve as alternatives for therapy in the future.

Nanomaterial-based adsorbents and optical sensors for illicit drug analysis.

The abuse of illicit drugs has been prevalent in recent years and is associated with crime and public health issues. To strengthen public security and fortify public health services with respect to the increasing severity of drug abuse, academic and government institutes have been devoted to constructing relative analytical methods for illicit drugs. To date, the development of sensors has been greatly emphasized due to their features of high sensitivity, prompt detection and flexible manipulation; thus, sensors can serve as alternatives to conventional sophisticated instruments. Recently, the use of nanomaterials has inspired the development of a series of innovative sample pretreatment and detection strategies in the field of analytical chemistry. Herein, this review elaborated the application of nanomaterials in analytical methods, including sample pretreatments, colorimetric sensors and fluorescent sensors. The utilization of nanomaterials in the analytical field provides novel perspectives for the development of detection platforms and facilitates the monitoring of illicit drugs in diverse complex matrices.

The matrix of SDS integrated with linear hydrophilic polymer for resolution of high- and low-molecular weight hyaluronic acids in MEKC.

Hyaluronic acid (HA), a multi-functional material, has a high dispersion in molecular weight, and the functions of HA are determined through the size. Nevertheless, hyaluronic acid mixtures are not easily separated due to their polydispersity. In this study, a capillary electrophoresis strategy was developed for resolution of different molecular-weight HA without enzymatic digestion. Here, hyaluronic acid mixtures with low molecular weight (380 kD; LHA) and high molecular weight (2180 kD; HHA) were successfully resolved by the SDS integrated with low molecular-weight polymer in capillary electrophoresis. By optimizing experimental conditions, the separation of LHA and HHA was completed within 14 min. The optimal conditions were as follows: the running buffer was 25 mM borate buffer (pH 9.75) containing 30 mM SDS and 10% polyethylene glycol (MW: 8000); applied voltage was 20 kV (detector at cathode side) and separation temperature was set at 25 °C. The data of method validation showed that calibration plots were linear (r ≥ 0.9977) over a range of 10-50 µg/mL for LHA, and 40-200 µg/mL for HHA. In the evaluation of precision and accuracy for this method, the RSD and RE values were all less than 4.2%. This fascinating technique was successfully applied to the quality control of cosmetic and pharmaceutical containing different ratios of LHA and HHA, and it was feasible for serving as a tool to quantitatively analyze different sizes of HA for clinical survey.

Specific Gene Capture Combined with Restriction-Fragment Release for Directly Fluorescent Genotyping of Single-Nucleotide Polymorphisms in Diagnosing Spinal Muscular Atrophy.

In this study, a fast and simple fluorescent genotyping strategy, streptavidin magnetic beads combined with biotin-coupled PCR and restriction-fragment release, was developed for determination of nucleotide variants. This method was further applied for analyzing SMN1 gene in diagnosis of spinal muscular atrophy (SMA). After biotin-coupled PCR, the streptavidin magnetic beads would capture the biotin-labeled SMN genetic fragments, and then the restriction enzyme of HPY188I could only digest and release the fluorescent end of SMN1 genetic fragment into the supernatant. Therefore, the SMN1 gene could be easily fluorescently quantified, and SMN2 would not, for diagnosis of SMA. The copy number of the SMN1 gene could be regressed using the relative fluorescent unit versus the known copy number, and the coefficient of correlation is equal to 0.9617 ( r = 0.9617). In this research, a total of 16 blind DNA samples were analyzed, including 6 wild types, 5 carriers, and 5 SMA patients. Importantly, this fast, simple, and highly efficient method is universal for detection of all nucleotides variants by replacing the specific restriction enzyme. This technique has the potency to be served as a tool for fast and accurate diagnosis of genotypes in clinical medicine.

Molecular inversion probes equipped with discontinuous rolling cycle amplification for targeting nucleotide variants: Determining SMN1 and SMN2 genes in diagnosis of spinal muscular atrophy.

The novel techniques of molecular inversion probes (MIPs) combined with discontinuous rolling cycle amplification (DRCA) was developed for determination of the multi-nucleotide variants at single base. The different-length MIPs, a padlock-probe based technology, are designed to simultaneously recognize the identical nucleotide variants. After ligation and DRCA, the different-length genetic products representing the certain genotypes could be simply determined by the short-end capillary electrophoresis (CE) method. By using MIPs-DRCA method, the various gene dosages of SMN1 and SMN2 genes in homologous or heterologous subjects were successfully quantified for diagnosis of spinal muscular atrophy (SMA). The length of the MIP for SMN1 gene was 106 bp, and for SMN2 gene was 86 bp. After method optimization, the MIP products of SMN1 and SMN2 were well separated with the resolution of 1.13 ± 0.17 (n = 3) within 10 min. There were total of 56 DNA blind samples analyzed by this strategy, including 38 wild types, 12 carriers and 6 SMA patients, and the data of gene dosages was corresponding to those analyzed by conformation sensitive CE and denatured high performance liquid chromatography (DHPLC) methods. This MIPs-DRCA method which could be applied to simultaneously genotype multi nucleotide variants at single base, such as K-ras gene, was very feasible for determination of genetic diseases in clinical.

Dialkyl anionic surfactant in field-amplified sample injection and sweeping-micellar electrokinetic chromatography for determination of eight leanness-promoting ß-agonists in animal feeds.

The beta-adrenergic agonists (ß-agonists) working as repartitioning agents that make the carcass leaner and enhance the feeding efficiency in animals have been banned in the European Union, China and Taiwan. Here, traditional anionic surfactants, such as sodium dodecyl sulfate (SDS) were replaced with sodium di-(2-ethylhexyl)-sulfosuccinate (AOT) in field-amplified sample injection and sweeping-micellar electrokinetic chromatography (FASI-sweeping MEKC) for simultaneous analysis of eight ß-agonists in animal feeds. The AOT vesicles provided a better resolution of ß-agonists than micelles of SDS. The detection limits of the eight ß-agonists were above 5ng/mL by using this stacking capillary electrophoresis (CE) method. In comparison of traditional MEKC method (sample injection, 1psi for 5s), the stacking strategy provided 400-2000 fold sensitivity enhancement. After method validation, this method was successfully applied for analyzing four animal feeds, and none ß-agonist was detected. This strategy possessing good resolution of eight ß-agonists was suitable for serving as a tool for routine analysis of animal feeds.

Blood pressure change during phacoemulsification and femtosecond laser-assisted cataract surgery.

AIM: To evaluate blood pressure (BP) changes during phacoemulsification (PC) and femtosecond laser (FSL)-assisted cataract surgery. METHODS: A retrospective chart review was performed for all patients who received traditional phacoemulsification surgery (PC group) and FSL-assisted cataract surgery (FS group) from July 2013 to December 2014. Totally 206 eyes from 133 patients receiving the two types of procedures were included. Patient characteristics (age, gender, and hypertension history), pre- and post-operative BPs were collected. RESULTS: The pro-operative systolic and diastolic BPs (mm Hg) were 124.89±20.48 vs 126.98±16.85, and 71.88±9.81 vs 73.56±10.03, in PC and FS groups, respectively. While the post-operative systolic and diastolic BPs (mm Hg) were 130.13±22.59 vs 134.77±17.52, and 73.41±11.62 vs 78.89±12.2, in PC and FS groups, respectively. Paired-sample t-tests showed obvious systolic and diastolic BP elevations in FS group after surgery (P=0.001 and 0.007) and no reliability in PC group (P=0.094 and 0.359). A linear regression model revealed systolic and diastolic BP elevations, which were related to longer surgical times for FS group (P=0.008 and 0.021). Age, gender, and hypertension history were not correlated with blood pressure elevation in either group. CONCLUSION: BP increases but at a limited level after FSL-assisted cataract surgery compared to traditional phacoemulsification.