Efficient Screening Method Research Articles

Fluorescence polarization immunoassay based on fragmentary hapten for rapid and sensitive screening of polymyxins in human serum

Polymyxin B (PMB) and polymyxin E (PME, or named colistin) are old polypeptide antibiotics and recently reintroduced as the last-line therapy for serious infections caused by multidrug-resistant Gram-negative pathogens. A rapid and reliable screening method for PMB and PME in human serum is urgently required due to their narrow therapeutic windows and individual differences. To develop fluorescence polarization immunoassay (FPIA) with high sensitivity and uniform specificity for PMB and PME, one novel fragmentary polymyxins hapten, named H1, was designed only containing the fatty acid acylated tripeptides and intendedly introducing sulfhydryl group for selective conjugation purpose. The H1 induced significantly higher titer and affinity antibody response than the PMB as hapten and produced five monoclonal antibodies (mAbs) with IC50 of 1.8–6.6 ng/mL for PMB and cross-reactions of 75.0–105.0 % for PME. To investigate the influence of tracer structure on the FPIA sensitivity, 12 tracers differing in spacer arm lengths, fluorophore types and hapten structures were fully paired with five mAbs, showing a safety length of spacer arm ranged from 4 to 17 Å. Under optimal conditions, the IC50 of FPIA was 3.7 ng/mL in buffer with a limit of detection of 1.4 μg/L in human serum for PMB and PME. The recoveries in spiked human serum were 73.8–91.8 % with CVs less than 12.8 % and whole assay time less than 25 min including sample preparation. These results showed that the FPIA was an efficient, accurate, and sensitive method for rapid screening of polymyxins and suitable for therapeutic drug monitoring.

Designing Hierarchically Porous Single Atoms of Fe-N5 Catalytic Sites with High Oxidase-like Activity for Sensitive Detection of Organophosphorus Pesticides.

Recently, single-atom catalysts (SACs) have been used to construct biosensors for the determination of organophosphorus pesticides (OPs). However, most nanozymes including SACs are peroxidase-like enzymes and require highly toxic and unstable hydrogen peroxide (H2O2) as a co-reactant to generate reactive oxygen species. Inspired by the heme site of cytochrome c oxidases (Ccos), the construction of Fe-N5-coordinated SACs by introducing axial N ligands is expected to bind O2 to generate active metal-oxygen intermediates. Herein, a SAC with an Fe-N5 active center confined by hierarchically porous carbon nanoframes (Fe SAs/N5-pC-4) was prepared by a polymerization-pyrolysis-evaporation-etching strategy, and its underlying enzyme-like mechanism was uncovered through experiments and density functional theory calculations. The 100% metal atom utilization, increased accessible active sites, accelerated mass transfer, excellent hydrophilicity, and an electron-driven mechanism of axial N endow the SAC with enhanced oxidase-like activity. Notably, its catalytic rate constant (0.398 s-1) is 569 times greater than that of the commercial Pt/C catalyst. Similar to the catalytic mechanism of Ccos, O2 can be converted into reactive oxygen species, avoiding the use of co-reactant H2O2 effectively. In addition, based on the inhibitory effect of thiols on the active site of Fe SAs/N5-pC-4, a biosensor was constructed and applied to the colorimetric analysis of OPs. This provides a facile, cost-effective method for efficient OP screening at sites to help control their contamination.

Cytological and histological examination of the endometrium based on menstruation age groups

Introduction: Abnormal uterine bleeding (AUB) is one of the most common symptoms of endometrial cancer that necessitates endometrial biopsy. Objectives: This study aimed to compare the cytological and histological findings in endometrial specimens among women with AUB in three different menstrual groups (pre-menopause, perimenopause, and post-menopause) and to evaluate the statistical accuracy of cytology compared to histopathology (gold standard). Patients and Methods: This descriptive study was conducted on 78 women with AUB admitted to the gynecology and obstetrics unit of Shahid Beheshti hospital, Isfahan, Iran from 2018 to 2020. Patients were divided into three groups (pre-menopause, perimenopause, and post-menopause) based on their menstrual condition. Samples were obtained using endometrial curettage and cytobrush and were analyzed by two surgical pathologists. The results were described in four categories: negative, atypical endometrial cells of undetermined significance (AEC-US), atypical endometrial cells encompassing the spectrum of precursors to the malignant endometrial tumor (AEC-PEMT), and positive. Results: Out of 78 samples, 36 (46.2%) were reported negative for epithelial abnormality, 15 (19.2%) were AEC-US, 9 (11.5%) were AEC-PEMT, and 18 (23.1%) were positive for epithelial lesions. There was a significant association between cytological findings and menstrual groups (P = 0.004). Positive results were more frequently reported in the post-menopausal group, whereas negative results were most common in the pre-menopause group. The sensitivity and specificity of cytological evaluation in the pre-menopause, perimenopause and post-menopause groups were 100% and 95% (accuracy: 100%, 95% CI: 0.94-1), 77% and 53%, (accuracy: 62%, 95% CI: 0.40-0.85), and 100% and 34% (accuracy: 41%, 95% CI: 0.20-0.61), respectively. Conclusion: Cytological examination of the endometrium demonstrated high sensitivity in the pre-menopausal and post-menopausal women with AUB and can therefore be used as an efficient and valuable method of screening for endometrial neoplasia.

Visualization of estimated prevalence of CES-D positivity accounting for background factors and AIS scores

Development of methods for population screening is necessary to improve the efficiency of secondary prevention of diseases. Until now, a common cutoff has been used for all people in the data set. However, if big data for health information can be used to modify individual cutoffs according to background factors, it may avoid wasting medical resources. Here we show that the estimated prevalence of the Center for Epidemiologic Studies Depression Scale positivity can be visualized by a heatmap using background factors from epidemiological big data and scores from the Athens Insomnia Scale. We also show that cutoffs based on the estimated prevalence can be used to decrease the number of people screened without decreasing the number of prevalent cases detected. Since this method can be applied to the screening of different outcomes, we believe our work can contribute to the development of efficient screening methods for various diseases.

Activity assay based on the immobilized enzyme kallikrein and mass spectrometry

Deregulated activity and expression of human kallikreins (KLKs) may be involved in various pathologies, so these enzymes are an attractive biological target for identifying molecules that can modulate KLK activity. This identification involves applying fast and efficient screening methods. This work describes an off-line assay with mass spectrometry (MS) detection that uses KLK immobilized on Sepharose-NHS as a micro-column configuration (IMER-KLK-Sepharose-NHS). The mass spectrometry used has an ion trap analyzer and electrospray ionization (EIS). The HPLC-MS method for quantifying KLK activity was developed. The enzymatic assay conditions were optimized, and the IMER-KLK-Sepharose-NHS kinetic parameter (KMapp = 15.48 ± 3 μmol L−1) was evaluated. Finally, the method was validated by using leupeptin as a reference inhibitor (IC50 = 0.85 ± 0.10 μmol L−1). The developed method was able to identify the reference inhibitor and can be an alternative for screening KLK inhibitors.

Lipophilic poly(glycolide) blocks in morpholin-2-one-based CARTs for plasmid DNA delivery: Polymer regioregularity, sequence of lipophilic/polyamine blocks, and nanoparticle stability as factors of transfection efficiency

In the present work, monosubstituted glycolides (G2–G4) derived from the (L)-tyrosine-based glycolide G1 and containing –CH2C6H4OR substituents (R = decanoyl, oleoyl, C(O)CH(n-C6H13)(n-C8H17)) have been used to design biodegradable and biocompatible lipophilic blocks in N-BOC-morpholin-2-one (M1)-based Charge-Altering Releasable Transporters (CARTs). Copolymers of dodecyl carbonate C1 (the ‘benchmark’ lipophilic comonomer) and M1, having direct (P1) or inverse (P2) block sequences; copolymers of G2–G4 and M1, having inverse block sequences and irregular, head-to-head in places, microstructures of the poly(glycolide) blocks (P3–P7); and copolymers of G2 and M1, having solely alternating head-to-tail microstructures of the poly(glycolide) blocks and direct (P8) or inverse (P9) block sequences, have been synthesized. DLS studies have shown that, after deprotection, the copolymers P1′–P9′ form liposomal particles in aqueous medium, differing considerably in their colloidal behavior, which depended on the length of the polyamine fragment and the nature of the lipophilic block. The ability of P1′–P9′ to deliver the plasmid pEGFP-N1 into cultured HEK293T cells (transfection efficiency) has been assessed at N/P ratios of the corresponding polyplexes in the range 1:1–50:1 by flow cytometry and further confirmed by fluorescence microscopy. Transfection experiments have demonstrated that the CARTs P1′, P4′, and P6′, containing ten (2-oxyethyl)glycine fragments, are about four times more efficient in delivering pDNA than commercial agents (Lipofectamine™ 3000, PEI MAX, X-tremeGENE™ 9, and GenJet™ In Vitro DNA Transfection Reagent). A correlation between the results of those studies and DLS / ζ-potential measurements has been established for P1′ and P4′: the transfection is efficient, if N/P ratios are equal to those favoring ζ-potential inversion at physiological pH. We conclude that glycolide-based CARTs show significant promise for further development as non-viral NA delivery vehicles and that DLS should be regarded as an efficient method of primary screening of amphiphilic copolymers, as well as copolymer-NA complexes and formulations thereof.

An efficient and novel method to screen Botrytis cinerea resistance genes based on TRV-induced gene silencing with lily petal discs

Lily (Lilium spp.) is an important ornamental crop because of its colorful flowers and fragrant smell. However, gray mold often occurs on lily flowers and leaves, which causes extensive economic losses. Most lily cultivars have poor tolerance to gray mold and are difficult to resist Botrytis cinerea, so it is urgent to identify resistance genes for developing resistant cultivars. So far, only a few B. cinerea resistant genes in lilies have been identified. This study aims to build an efficient method to screen B. cinerea resistance genes in lily. The resistance determination of detached lily discs revealed that the petal discs were more sensitive to B. cinerea than the leaves, and ‘Sorbonne’ was a suitable experimental material with B. cinerea pathogen susceptibility for investigation. In addition, it was also found that the petal discs from the completely opened ‘Sorbonne’ flowers were inoculated with B. cinerea spore suspension at 1 × 105 concentrations, which was more appropriate for symptom observation. Based on these results, we developed an Agrobacterium-mediated tobacco rattle virus (TRV)-induced gene silencing (VIGS) method on detached ‘Sorbonne’ petal discs for screening B. cinerea-resistance genes. The bacteria solution at an OD600 of 1.5 got the highest VIGS efficiency at 9 days post infection. Three differently expressed genes, LlWRKY3, LlWRKY7, and LlWRKY10, which came from the B. cinerea-infection transcriptome, were identified with this TRV-based VIGS method. Silencing of them in petal discs caused them to be more sensitive to B. cinerea. In the current study, the VIGS method based on TRV was successfully developed on detached ‘Sorbonne’ petal discs. Target genes of petal discs that were vacuum infiltrated with Agrobacterium bacteria solution at an OD600 of 1.5 were silenced well after 9 days. Compared with traditional method of screening resistant gene, the TRV-VIGS method with petal discs was a convenient and efficient method for screening B. cinerea resistant genes, and have important value in breeding of lily with high resistance to gray mold.

Agar Plate‐Based Screening Approach for the Identification of Enzyme‐Catalyzed Oxidations

AbstractBiocatalytic oxidation reactions are in high demand. Among the applied enzymes, the heme‐thiolate enzyme subfamily of unspecific peroxygenases (UPOs), which relies on hydrogen peroxide as cosubstrate and oxidant, has generated great interest. Almost two decades after their first description, databases provide thousands of putative UPO sequences, but only a few enzymes have been characterized. To address this gap, efficient screening methods for the identification of novel UPOs are necessary. Here, a new screening strategy is presented based on solid cultivation of wild‐type fungi. The identification of a promising candidate strain highlights the applicability of this approach.

Predicting the prevalence of lung cancer using feature transformation techniques

Healthcare sector is one of the most important sectors of any country as a big part of the country’s economy is associated with it. The research is about to contribute to the health sector by minimizing the expenses of a lung cancer diagnosis. The study tries to devise an efficient method for initial screening of the patients with symptoms through their demographic and clinical data. The study seeks appropriate feature transformation techniques from dimensionality reduction techniques in combination with an apposite regression model that can perform this task robustly using the lung cancer dataset for early carcinoma diagnosis. To equip the health sector with state-of-the-art technology and for the betterment of humankind, the most beneficial tool of today is machine learning. Lungs play a vivacious role in the human body, oxygen is circulated in the body, and the air is taken from the atmosphere and sends it into the bloodstream. Several people decease every year because of lung carcinoma as lung cancer is normally identified at the latter phase due to lack of awareness. It stays unidentified because people have pneumonia often, which converts into lung cancer later. The projected research seeks to enable health professionals to the rationalization of primary diagnosis and treatment of lung carcinoma in developing countries. The proposed methodology has selected the optimized combination of regression-based machine learning technique and feature transformation technique for the available patterns based on demographic and clinical features of lung cancer patients. Based on the gathered results during training (RMSE = 0.1324, R2-Score = 0.7428) and testing (RMSE = 0.1273, R2-Score = 0.7405) it can be concluded that the fast independent component analysis and elastic net regression technique provided optimized results and outperform the other aspirant techniques.

High detection rate for disease-causing variants in a cohort of 30 Iranian pediatric steroid resistant nephrotic syndrome cases.

BackgroundSteroid resistant nephrotic syndrome (SRNS) represents a significant renal disease burden in childhood and adolescence. In contrast to steroid sensitive nephrotic syndrome (SSNS), renal outcomes are significantly poorer in SRNS. Over the past decade, extensive genetic heterogeneity has become evident while disease-causing variants are still only identified in 30% of cases in previously reported studies with proportion and type of variants identified differing depending on the age of onset and ethnical background of probands. A genetic diagnosis however can have implications regarding clinical management, including kidney transplantation, extrarenal disease manifestations, and, in some cases, even causal therapy. Genetic diagnostics therefore play an important role for the clinical care of SRNS affected individuals.Methodology and resultsHere, we performed NPHS2 Sanger sequencing and subsequent exome sequencing in 30 consanguineous Iranian families with a child affected by SRNS with a mean age of onset of 16 months. We identified disease-causing variants and one variant of uncertain significance in 22 families (73%), including variants in NPHS1 (30%), followed by NPHS2 (20%), WT1 (7%) as well as in NUP205, COQ6, ARHGDIA, SGPL1, and NPHP1 in single cases. Eight of these variants have not previously been reported as disease-causing, including four NPHS1 variants and one variant in NPHS2, ARHGDIA, SGPL1, and NPHP1 each.ConclusionIn line with previous studies in non-Iranian subjects, we most frequently identified disease-causing variants in NPHS1 and NPHS2. While Sanger sequencing of NPHS2 can be considered as first diagnostic step in non-congenital cases, the genetic heterogeneity underlying SRNS renders next-generation sequencing based diagnostics as the most efficient genetic screening method. In accordance with the mainly autosomal recessive inheritance pattern, diagnostic yield can be significantly higher in consanguineous than in outbred populations.

Quantitative detection of malachite green in sediment by a time-resolved immunofluorescence method combined with a portable 3D printing equipment platform

Rapid detection technology of aquaculture fishery drug residues is needed to supplement large-scale instrument methods. To do this, the time-resolved fluorescence immunoassay (TRFIA) method and portable three-dimensional (3D) printing equipment platform were used, in combination with smartphones, to detect malachite green (MG) in pond sediments. The TRFIA was coupled to MG monoclonal antibodies (mAb) through lanthanide metal microspheres europium (Eu3+). The labeled antibody produced competitive immunity in the immune reaction system, and the specific fluorescence intensity in the product was determined by a portable 3D printing equipment platform to achieve quantitative analysis. To test this method, leucomalachite green (LMG) was converted to MG by oxidation of dicyanoquinone (DDQ), and a qualitative analysis was achieved. Methodological evaluation results were satisfactory, recoveries were 83 %–104 %, the limit of detection (LOD) was 0.3 ng/g, the limit of quantitation (LOQ) was 0.7 ng/g, and the coefficient of variation was 1.3 %–7.3 %. The linear equation y = −0.1496x + 0.5585 was in the range of 0–10 ng/g. The linear regression correlation coefficient was 99.2 %. The TRFIA was confirmed and positive samples were measured. Results were consistent with the standard method, which demonstrated that the TRFIA was feasible and that the detection results were reliable. Compared with the national standard method, the TRFIA saves time, is more convenient, and has high sensitivity. It provides an efficient technical method for the rapid screening of MG in the sediments of aquaculture environments.

Rapid screening of 15 highly toxic pesticide residues in Angelica sinensis decoctions by thermal desorption electrospray ionization mass spectrometry

ABSTRACT A decoction is the most common form of Chinese herbal medicine dosing. An Angelica sinensis decoction was qualitatively and semi-quantitatively analyzed for 15 highly toxic pesticide residues by thermal desorption electrospray ionization mass spectrometry (TD-ESI/MS) with a triple quadrupole mass spectrometer in multiple reaction monitoring mode without pretreatment. The instrument response was optimal and sensitivity was highest with the following parameters: thermal desorption temperature, 300 °C; 0.1% formic acid aqueous solution-methanol (1:1,V:V) solvent; syringe pump flow rate, 150 μL/h; and sampler length, 5 cm. Good linearity was attained within the appropriate linear range with the detection limits ranging from 0.01–0.05 mg/L. The same positive sample was verified for QuEChERS pretreatment coupled with liquid chromatography tandem mass spectrometry, showing good agreement with TD-ESI/MS. The newly developed method was demonstrated to be a rapid, green, and efficient analytical method for screening 15 highly toxic pesticide residues in Angelica sinensis decoctions.

Triple immunochromatographic test system for detection of priority aquatic toxins in water and fish.

Aquatic toxins are a group of toxic compounds produced by several types of freshwater and marine algae and cyanobacteria and transported through the food chains of water bodies. Potential contamination of aquaculture products (raw and processed fish and seafood) with aquatic toxins requires the use of efficient screening methods for their control. In this study, a multiplex immunochromatographic test system for the simultaneous detection of three aquatic toxins-phycotoxins domoic acid (DA) and okadaic acid (OA), and cyanotoxin microcystin-LR (MC-LR)-is for the first time developed. For this, a competitive indirect immunochromatographic analysis (ICA) based on gold-labeled secondary antibodies was carried out. The LODs/cutoffs/working ranges of the ICA were 0.05/0.3/0.07-0.29, 1.3/100/3.2-58.2, and 0.1/2.0/0.2-1.1ng/mL for MC-LR, DA, and OA, respectively. The assay duration was 18min. The developed test system was used to analyze water samples from natural sources (salt and fresh water) and fish samples. For sample preparation of water, simple dilution with a buffer was proposed; for fish samples, methanol-water extraction was utilized. It was demonstrated that the triple LFIA specifically detected target aquatic toxins with recoveries of 85.0-121.5%. The developed multiplex LFIA can be considered a promising analytical solution for the rapid, easy, and sensitive control of water and food safety.

How to simply and efficiently screen microbial strains capable of anaerobic biosynthesis of biosurfactants: Method establishment, influencing factors and application example evaluation.

Microbial resources capable of anaerobic biosynthesis of biosurfactants are increasingly interested for their application in oxygen-deficient environments, such as in-situ microbial enhanced oil recovery and anaerobic bioremediation. How to simply and efficiently screen microbial strains capable of anaerobic biosynthesis of biosurfactants need be further studied in depth. In this study, an efficient and simple screening method was established based on the oil displacement characteristic of biosurfactants combined with the anaerobic culture technology using microplate assays. Strains whose anaerobic culture in microwells can form oil displacement circles with diameters larger than 10 mm were screened for scale-up culture in anaerobic tubes. The screened strains which can reduce the surface tension of anaerobic culture to lower than 45 mN/m were verified as positive strains. Using this screening method, eight positive strains and thirteen positive strains were screened from oil reservoir produced water and oily sludge, respectively. Through phylogenetic analysis, some screened strains were identified as Pseudomonas sp., Bacillus sp., and Enterobacter sp. This study also found that more microbial strains might be isolated after enrichment culture of environmental samples, whereas more microbial species would be isolated without enrichment. Suspension of environmental samples prepared with distilled water or normal saline had no significant effect. The established screening method is highly targeted and efficient for microbial strains capable of anaerobic biosynthesis of biosurfactants. The diameter of oil displacement circle is a reliable screening indicator. This study will contribute to explore more microbial resources which can anaerobically biosynthesize biosurfactants.

Genetic Variability Assessment of Tropical Indica Rice (Oryza sativa L.) Seedlings for Drought Stress Tolerance.

Drought stress is one of the most devastating abiotic factors limiting plant growth and development. Devising an efficient and rapid screening method at the seedling stage is vital in identifying genotypes best suited under drought conditions. An experiment was conducted to assess 74 rice genotypes for drought tolerance using specially designed mini-hoop structures. Two treatments were imposed on rice seedlings, including 100% moisture and a 50% moisture regime. Several shoot morpho-physiological traits and root traits were measured and analyzed. The genotypes exhibited a wide range of variability for the measured traits, with the leaf area showing the most significant variation, followed by plant height, tiller number, and shoot dry weight. In contrast, the drought did not significantly affect most root traits. The germplasm was classified into different categories using cumulative drought stress response indices (CDSRI); 19 genotypes (26%) were identified as drought sensitive, and 33 (45%), 15 (20%), and 7 (9%) were determined as low, moderately, and highly drought-tolerant, respectively. Genotypes IR86638 and IR49830 were the most and least drought-tolerant, respectively. Overall, a poor correlation was observed between CDSRI, total shoot traits (R2 = 0.36), and physiological parameters (R2 = 0.10). A strong linear correlation was found between CDSRI and root traits (R2 = 0.81), suggesting that root traits are more crucial and better descriptors in screening for drought tolerance. This study can help rice breeders and scientists to accelerate breeding by adopting a mini-hoop rapid screening method. The tolerant genotypes could serve as appropriate donor parents, progenies, and potential genotypes for developing drought-tolerant commercial cultivars.

A novel method for efficient screening and annotation of important pathway-associated metabolites based on the modified metabolome and probe molecules

植物次生代谢物在抵御生物/非生物胁迫、生物间互作以及信息传递等方面发挥重要作用,次生代谢途径解析对植物分子育种、天然产物合成等方面具有重要意义。液相色谱-高分辨串联质谱(LC-HRMS/MS)为次生代谢物鉴定及途径表征提供了技术手段。非靶向LC-HRMS/MS方法可获得丰富的质谱信号,包括一级质谱和二级质谱(MS, MS/MS),但受质谱数据库规模以及次生代谢物复杂性的制约,次生代谢物注释十分困难。该研究以玉米叶片中苯丙烷途径代谢物为例,发展用于非靶向代谢组数据中重要途径代谢物的高效筛选和注释新方法。首先,利用公共代谢途径数据库及文献获取参与苯丙烷代谢途径的61种修饰反应类型,进而从非靶向实验数据中筛选出修饰代谢组。其次,获取开源串联质谱数据中的苯丙烷类化合物作为探针分子,构建探针分子质谱数据库。将探针分子与修饰代谢组共建分子网络,锁定目标途径代谢物并注释结构。该方法在正、负离子模式下分别筛选出玉米叶片中392个和417个苯丙烷途径候选代谢物,去冗余后共注释出129个代谢物,涉及苯丙烷代谢的主要分支途径,如黄酮途径的8个类黄酮、19个氧苷类黄酮和32个碳苷类黄酮,31个羟基肉桂酸途径代谢物以及22个木脂素途径代谢物;其中26个在PubChem和SciFinder数据库中未见收录。该研究利用探针分子结合修饰组可快速锁定途径代谢物,且有助于快速、准确的网络传播注释,可显著提高目标途径代谢物筛选与注释效率,为植物次生代谢途径的深入解析提供分析手段。

Rapid Screening of Bimetallic Electrocatalysts Using Single Nanoparticle Collision Electrochemistry.

The pace of nanomaterial discovery for high-performance electrocatalysts could be accelerated by the development of efficient screening methods. However, conventional electrochemical characterization via drop-casting is inherently inaccurate and time-consuming, as such ensemble measurements are serially performed through nanocatalyst synthesis, morphological characterization, and performance testing. Herein, we propose a rapid electrochemical screening method for bimetallic electrocatalysts that combines nanoparticle (NP) preparation and performance testing at the single NP level, thus avoiding any inhomogeneous averaging contribution. We employed single NP collision electrochemistry to realize in situ electrodeposition of a precisely tunable Pt shell onto individual parent NPs, followed by instantaneous electrocatalytic measurement of the newborn bimetallic core-shell NPs. We demonstrated the utility of this approach by screening bimetallic Au-Pt NPs and Ag-Pt NPs, thereby exhibiting promising electrocatalytic activity at optimal atomic ratios for methanol oxidation and oxygen reduction reactions, respectively. This work provides a new insight for the rapid screening of other bimetallic electrocatalysts.

Identifying potential ligands specifically binding to beta1-adrenoceptor from Radix Aconiti Lateralis Praeparata extract by affinity chromatographic method

As expressed predominantly in cardiac tissue, beta1-adrenoceptor (β1-AR) is broadly accepted as one of the main targets for drugs against cardiovascular ailments. However, the discovery of β1-AR ligand is gravely challenged due to the lack of efficient screening method. This work developed a general strategy for pursuing β1-AR ligands from the herbal extract by immobilizing haloalkane dehalogenase (Halo)-tagged β1-AR onto microspheres coated with 6-chlorohexanoic acid, and applying the immobilized β1-AR in the analysis of ligand-receptor interaction. The morphology was characterized by scanning electron microscope (SEM) and X-ray photoelectron spectroscopy (XPS). The chromatographic specificity of the immobilized receptor column was evaluated by determining the association constants of atenolol, esmolol and metoprolol using stepwise frontal analysis plus injection amount-dependent method. The potential ligands binding to β1-AR was screened by collecting the peak with retention time longer than the void time, and identified the collection by reverse phase liquid chromatography coupled with tandem mass spectrometry. The association constants of the three drugs to β1-AR were (3.33 ± 0.29)× 106 M−1, (2.33 ± 0.23)× 106 M−1 and (2.06 ± 0.03)× 106 M−1, indicating a desired specificity of the immobilized receptor for recognizing its ligands. Molecular docking showed that van der Waals, hydrogen bonds, and hydrophobic interactions were the principal interaction forces for the receptor-drug complexes. Benzoylmesaconine was screened as the potential ligand of β1-AR in Radix Aconiti Lateralis Praeparata extract. The association constant of the ligand was (1.06 ± 0.02)× 105 M−1, hinting structural modification may be required before clinical application. The immobilized β1-AR is possible to provide a rapid method for screening potential ligands in herbal extract.

Cytohistological findings in HPV‐negative cases from HPV primary screening programme: quality assurance study

To evaluate the risk of false HPV-negative results and possible related morphological abnormalities in HPV primary cervical cancer screening. Out of 53,661 HPV-negative cases, 5469 (10.2%) randomly selected cytology slides were evaluated as a part of the quality assurance protocol. The Bethesda category negative for intraepithelial lesion or malignancy (NILM) in the HPV-negative cases given was present in 95.4%. Due to cytology other than NILM, 0.4% of cases were referred to colposcopy and 4.2% to the follow-up in one year. In the follow-up HPV negativity and NILM cytology was present in 88.3% of attended women. Cases other than HPV negative and NILM were referred to colposcopy. One biopsy-proven histological HSIL was found in the first round and one in follow-up screening. More comprehensive genotyping of HSIL cases revealed genotypes 69 and 11. Only two HPV test negative cases with histological HSIL were revealed forming 0.04% of quality control group. In both cases, HPV genotype not included in screening tests was found. According to the results, the primary HPV test with cytology triage is an efficient and specific method for cervical cancer screening despite of the fact that some non-high-risk genotypes are missed.

Evaluation of infrared thermography with a portable camera as a diagnostic tool for peripheral arterial disease of the lower limbs compared with color Doppler ultrasonography.

IntroductionWe aimed to evaluate whether infrared thermography (IRT) with a portable camera is a useful tool for diagnosing or screening peripheral arterial disease of the lower limbs (PAD-LL) when compared with the traditional method of color Doppler ultrasonography.Material and methodsThe study enrolled 90 volunteers: 45 who were diagnosed with PAD-LL (PAD-LL group) and 45 who did not have a diagnosis of PAD-LL (control group). The diagnosis was made using color Doppler ultrasonography, and the results were compared with those of IRT.ResultsThe IRT-based procedure evaluated in this study had a sensitivity of 97.62% and a specificity of 91.67% for PAD-LL diagnosis compared to color Doppler ultrasonography. The method was limited for diagnosing PAD-LL manifesting above the knees (suprapopliteal PAD-LL). Our results also suggest that the ankle-brachial index is an important predictor of PAD-LL, with a sensitivity of 91.17% and a specificity of 75% at a value of ≤ 0.9. Current or previous smoking habits, higher body mass index, and the presence of diabetes mellitus were significantly elevated in the PAD-LL group.ConclusionsOur results indicate that IRT is an efficient and low-cost method for screening and diagnosing PAD-LL, particularly infrapopliteal PAD-LL manifesting below the knees. However, further studies are required to establish the validity of this technique.