Two-step Sample Research Articles

A candidate reference measurement procedure for quantification of glycocholic acid in human serum based on isotope dilution liquid chromatography-tandem mass spectrometry

Accurate measurement of serum glycocholic acid (GCA) is crucial for evaluating the activity of chronic hepatitis. Moreover, GCA is a novel identified biomarker for hepatocellular carcinoma. Although some laboratories have used the liquid chromatography-tandem mass spectrometry (LC–MS/MS) method to measure GCA in recent years, the problem of potential interference of GCA analogues has not been solved well yet. Neither reference measurement procedures nor reference materials for GCA have been listed in the Joint Committee for Traceability in Laboratory Medicine (JCTLM) database. For standardization of GCA, it is urgent to establish a candidate measurement procedure for GCA. In this study, a candidate reference measurement procedure for the quantification of GCA in human serum based on isotope dilution liquid chromatography-tandem mass spectrometry (ID-LC–MS/MS) by a two-step sample pretreatment of protein precipitation and MAX solid-phase extraction was developed and validated. GCA can be completely separated from its structural analogues with gradient elution in 9 min compared with short time gradients published in previous literature by Huang’s group. Method validation indicated perfect quantitation precision with intra-day and inter-day values that were ≤1.30% and ≤1.80%, respectively. The method showed excellent linearity with high regression coefficients (R2 > 0.999) over a range of 0.92 ng/g–38.38 μg/g and perfect recoveries at three spiked levels (99.87–100.43%). No interference, matrix effect, and carryover were observed. Moreover, the cRMP was successfully applied to measure GCA in serum samples and compared with two immunoassays in a clinical laboratory. As a candidate reference method, this method can promote a GCA standardization program.Graphical

Towards elevated perfluorooctanoic acid (PFOA) enrichment in water: Sequential liquid-liquid extraction pretreatment for ion chromatography detection

The widespread detection of perfluorooctanoic acid (PFOA) in the environment has raised significant concerns. The standard PFOA analytical method relies on expensive solid-phase extraction (SPE) and liquid chromatography tandem mass spectrometry (LC-MS/MS) instruments, making routine use prohibitive. We herein proposed a cost-effective yet novel enrichment method for determining PFOA at ng L−1 level. This method entailed a two-step sample preparation process: firstly, PFOA was extracted and enriched using a forward-extraction under acidic conditions, followed by a backward-extraction and enrichment step utilizing alkaline water. The enriched samples were subsequently subjected to a common ion chromatography (IC). Results reveal that maintaining a forward-extraction pH below its pKa value (2.8) is essential, as protonated PFOA proves effective in enhancing the enrichment factor (EF). The challenge lied in driving PFOA from forward-extractant to aqueous backward-extractant due to the decreased hydrophobicity of deprotonated PFOA (log Kow2 = 1.0). In addition, we found that evaporating forward-extractant with alkaline backward-extractant (containing 5% methanol) reduced potential analytical uncertainties associated with PFOA evaporation and sorption. Under optimal conditions, the method achieved a detection limit of 9.2 ng L−1 and an impressive EF value of 719. Comparison with SPE-LC-MS/MS confirmed the proposed method as a promising alternative for PFOA determination. Although initially targeted for PFOA, the novel methodology is likely applicable to preconcentration of other poly-fluoroalkyl substances.

A comparative study of TiO2 nanostructures prepared by one-step and two-step hydrothermal method: Physical and photoelectrochemical properties

Anatase TiO2 films grown on Ti foils presenting nanosheet or nanowire morphologies were fabricated by one-step and two-step hydrothermal methods with different durations. Herein, the periodic patterns on the surface of Ti foils caused by the TiO2 films peeling off promoted the growth of the two-step samples. Besides, the bandgaps of two-step samples were lower than those of one-step samples and the lowest bangap of 3.0 eV was obtained from the two-step sample with 4 h treatment. Meanwhile, this 4 h sample with the longest lifetime (79.5 ms) showing the optimized photocurrent density of 352.8 μA/cm2 in the two-step samples was superior to the optimum one (4 h) in one-step samples with the density of 268.3 μA/cm2. Thus, relatively short hydrothermal treatment is benefit to the photoelectrochemical performance whether for one-step or two-step samples. Besides, the periodic patterns increased the density of second-step TiO2 resulting in the narrowed bandgap and superior photocurrent density of two-step samples.

ESG performance and corporate fraudulence: Evidence from China

By adopting a large sample of Chinese public listed firms from 2014 to 2021, we examine whether firms' ESG performance inhibits corporate fraud. Using panel data regression techniques, we find that high ESG performance mitigates corporate fraudulence. After conducting a series of robustness tests, including propensity score matching (PSM) model, Heckman two-step sample selection model, instrumental variable (IV) and 2SLS regression model, and firm fixed effect model, the results remain unchanged. Further analyses suggest that this negative relationship is more pronounced in non-state-owned enterprise (non-SOE) firms and firms that voluntarily disclose ESG information. The mechanism analysis suggests that high-quality ESG engagement improves firms' governance performance and inhibits managerial myopia, which thereby mitigates corporate fraud. Overall, our findings provide incremental evidence on the role of ESG in filling institutional voids in emerging economies. Our findings also provide significant policy implications to regulators and policy makers who seek to promote fair information disclosure and mitigate corporate fraud risk.

Improving the diagnosis of active tuberculosis: a novel approach using magnetic particle-based chemiluminescence LAM assay

ObjectivesTuberculosis (TB) is a significant global health concern, given its high rates of morbidity and mortality. The diagnosis using urine lipoarabinomannan (LAM) primarily benefits HIV co-infected TB patients with low CD4 counts. The focus of this study was to develop an ultra-sensitive LAM assay intended for diagnosing tuberculosis across a wider spectrum of TB patients.Design & MethodsTo heighten the sensitivity of the LAM assay, we employed high-affinity rabbit monoclonal antibodies and selected a highly sensitive chemiluminescence LAM assay (CLIA-LAM) for development. The clinical diagnostic criteria for active TB (ATB) were used as a control. A two-step sample collection process was implemented, with the cutoff determined initially through a ROC curve. Subsequently, additional clinical samples were utilized for the validation of the assay.ResultsIn the assay validation phase, a total of 87 confirmed active TB patients, 19 latent TB infection (LTBI) patients, and 104 healthy control samples were included. Applying a cutoff of 1.043 (pg/mL), the CLIA-LAM assay demonstrated a sensitivity of 55.2% [95%CI (44.13%~65.85%)], and a specificity of 100% [95%CI (96.52%~100.00%)], validated against clinical diagnostic results using the Mann-Whitney U test. Among 11 hematogenous disseminated TB patients, the positive rate was 81.8%. Importantly, the CLIA-LAM assay consistently yielded negative results in the 19 LTBI patients.ConclusionOverall, the combination of high-affinity antibodies and the CLIA method significantly improved the sensitivity and specificity of the LAM assay. It can be used for the diagnosis of active TB, particularly hematogenous disseminated TB.

Returnee directors and green innovation

This study examines the effect of returnee directors, defined as local directors who have studied or worked in foreign countries, on a firm’s green innovation. Utilising robust econometric modelling, we examine a large sample of A-share nonfinancial firms listed on the Shenzhen and Shanghai stock exchanges. Our findings reveal a positive and significant relationship between returnee directors and green innovation. Interestingly, this association remains consistent across both environmentally sensitive and non-sensitive industries. However, it becomes more pronounced when returnee directors are non-executive, function within state-owned enterprises, number more than one on the board, or operate in firms with poor internal and external corporate governance environments. Furthermore, we demonstrate that returnee directors, armed with foreign education or work experience, increase green innovation. The results withstand a series of robustness checks, including the Heckman two-step sample selection model and propensity score matching. Overall, the results suggest that returnee directors drive green innovation.

Hypothesis testing for mean vector and covariance matrix of multi-populations under a two-step monotone incomplete sample in large sample and dimension

In this study, we focus on the critical issue of analyzing data sets with missing data. Statistically processing such data sets, particularly those with general missing data, is challenging to express in explicit formulae, and often requires computational algorithms to solve. We specifically address monotone missing data, which are the simplest form of data sets with missing data. We conduct hypothesis tests to determine the equivalence of mean vectors and covariance matrices across different populations. Furthermore, we derive the properties of likelihood ratio test statistics in scenarios involving large samples and large dimensions.

Spatial differencing for sample selection models with ‘site-specific’ unobserved local effects

Summary This paper proposes an estimator which combines spatial differencing with a two-step sample selection estimator. We derive identification, estimation, and inference results from ‘site-specific’ unobserved effects. These effects operate at a spatial scale that cannot be captured by administrative borders. Therefore, we use spatial differencing. We show that under justifiable assumptions, the estimator is consistent and asymptotically normal. A Monte Carlo experiment illustrates the small sample properties of our estimator. We apply our procedure to the estimation of a female wage offer equation in the United States and the results show the relevance of spatial differencing to account for ‘site-specific’ unobserved effects.

Intra-subject enveloped multilayer fuzzy sample compression for speech diagnosis of Parkinson's disease.

Machine learning-based Parkinson's disease (PD) speech diagnosis is a current research hotspot. However, existing methods use each corpus sample as the base unit for modeling. Since different corpus samples within the same subject have different sensitive speech features, it is difficult to obtain unified and stable sensitive speech features (diagnostic markers) that reflect the pathology of the whole subject. Therefore, this study aims at compressing the corpus samples within the subject to facilitate the search for diagnostic markers with high diagnostic accuracy. A two-step sample compression module (TSCM) can solve the problem above. It includes two major parts: sample pruning module (SPM) and sample fuzzy clustering mechanism (SFCMD). Based on stacking multiple TSCMs, a multilayer sample compression module (MSCM) is formed to obtain multilayer compression samples. After that, simultaneous sample/feature selection mechanism (SS/FSM) is designed for feature selection. Based on the multilayer compression samples processed by MSCM and SS/FSM, a novel ensemble learning algorithm (EMSFE) is designed with sparse fusion ensemble learning mechanism (SFELM). The proposed EMSFE is validated by visualization of extracted features and performance comparison with related algorithms. The experimental results show that the proposed algorithm can effectively extract the stable diagnostic markers by compressing the corpus samples within the subject. Furthermore, based on LOSO cross validation, the proposed algorithm with extreme learning machine (ELM) classifier can achieve the accuracy of 92.5%, 93.75% and 91.67% on three datasets, respectively. The proposed EMSFE can extract unified and stable sensitive features that accurately reflect the overall pathology of the subject, which can better meet the requirements of clinical applications.The code and datasets can be found in: https://github.com/wywwwww/EMSFE-supplementary-material.git.

Multiplex Digital Immunoassays with Reduced Pre-partition Reaction via Massively Parallel Reagent Delivery on a Bead-Based SlipChip.

The emergence of digital immunoassays has advanced the sensitivity of protein analysis to ultrahigh sensitivity at the attomolar level. However, the background signal generated by the premixing of immunocomplexes and fluorogenic substrates can limit the precise quantification, especially in multiplexed assays. Herein, a bead-based SlipChip (bb-SlipChip) microfluidic device capable of massively parallel two-step sample loading is presented. The background signal can be suppressed through a two-step loading mechanism. Specifically, encapsulate the beads into the microwells first and then, through a slipping process, deliver the fluorogenic substrate in parallel into 281,200 microwells of 68 fL to perform the digital immunoassay. The quantification capability is demonstrated with a duplex assay of IL-6 and IL-10, achieving a limit of detection of 5.2 and 15.3 fg/mL, which is approximately 2-3 times improved compared to a commercial Simoa system. The bb-SlipChip provides a robust and universal method for digital immunoassay and can be extended to higher multiplexed detection as well as other biomedical applications involving microbeads.

Corporate key labs: Breakthrough or white elephant?

Based on manually collected data from Chinese listed firms' key laboratories at the state and provincial levels, this study shows that key laboratories improve firms' innovation output. Corporations with key laboratories at the state or provincial level produce more patents and citations than their counterparts. A series of endogenous treatment effects, including the Heckman two-step sample selection model, instrumental variable estimation, policy shock analysis, and propensity score matching, indicate that this study's main conclusion is robust and consistent. We also observe that key laboratories' beneficial impact on innovation output becomes more prominent for firms belonging to high-tech industries, those led by an inventor or scientist CEO, and those located in cities that enforce the protection of intellectual property. Further, key laboratories foster innovation mainly by developing scientific research capacity, increasing human capital, and improving R&D subsidies. Our findings demonstrate that key laboratories can benefit firms, their stakeholders, and the public in an emerging market such as China.

Substantial coercivity enhancement in Dy-free Nd-Fe-B sintered magnet by Dy grain boundary diffusion

To date, the highest coercivity reported for Dy-free Nd–Fe–B sintered magnets under the Dy grain boundary diffusion (GBD) process is ∼2.4 T. Therefore, exploring various approaches for achieving higher coercivity is beneficial for eliminating the dependence on the scarce and expensive heavy rare-earth elements, Dy and Tb. In this study, we report record-breaking high coercivity by separately diffusing Dy70Cu30 and then Pr68Cu32 eutectic alloys through grain boundaries in fine-grained Dy-free sintered magnets. The two-step GBD-processed sample containing a low Dy content (only 0.9 wt%) exhibited a high coercivity of 2.8 T and an excellent temperature coefficient of coercivity of β = -0.447%/ °C. The second GBD process facilitated Dy infiltration from the bulk surface toward the center by chemically induced liquid film migration to form a uniform Dy-rich shell throughout the bulk. Pr was also enriched in the Dy-rich shell near the surface of Nd2Fe14B grains in the two-step GBD-processed sample. However, the Pr-enrichment in the shell did not cause the degradation of the coercivity at high temperatures. This unique shell structure strengthened the anisotropy field to achieve 2.8 T coercivity without alloying Dy to the initial sintered magnet.

Optimizing the magnetic and giant magneto-impedance (GMI) properties of glass-coated amorphous Co-based wires via single- and two-step direct current annealing

The effects of both single- and two-step direct current (DC) annealing procedures on the magnetic properties (including the giant magneto-impedance (GMI)) of glass-coated Co66Fe6Si13B15 amorphous wires were evaluated. For both methods, the GMI ratio initially increased with the current intensity and reached a peak value before declining at higher current intensities. A maximum improvement of the GMI ratio of about five times was obtained for the two-step DC-annealed sample, while that for single-step DC-annealed samples was only 2.44 times. A higher GMI sensitivity was also observed for the two-step DC-annealed wires. The better GMI performance of the two-step DC annealed samples was attributed to the more homogeneously distributed precipitation in the matrix and the ordered circumferential magnetic domains on the surface. The results obtained further imply that DC annealing at a higher current intensity but over a shorter treatment time is a more efficient and energy-saving method for enhancing the GMI performance of Co-based amorphous wires for potential industrial applications.

Do geographically nearby major customers mitigate suppliers’ stock price crash risk?

This study examines the impact of geographically nearby major customers on suppliers' stock price crash risk. Using a sample of Chinese A-share listed firms and their top five (major) customers during the period 2008–2019, we find a significantly negative association. This association is robust in a series of robustness checks, including the use of instrumental variables estimations, propensity score matching procedure, and Heckman two-step sample selection model. The mitigating effect of supplier−customer proximity on crash risk is more pronounced for suppliers with lower corporate transparency and greater operational uncertainty. Finally, we identify two possible mechanisms through which geographically nearby major customers reduce suppliers’ crash risk: fewer financial restatements and higher accounting conservatism of suppliers. The findings of this study indicate that listed firms may choose geographically nearby customers to reduce crash risk.

Size and characteristics of family caregiving for people with serious illness: A population-based survey.

Family caregivers play a vital role in care for people with serious illness. Reliable population-level information on family caregiving is scarce. We describe the socio-demographic and family caregiving characteristics and experiences of family caregivers of people with serious illness in the adult population. We performed a secondary analysis of the cross-sectional population-based 19th Social-Cultural Changes survey. A random sample of 2,581 Dutch-speaking people aged 18-95, living in Flanders or Brussels, were contacted for participation in the survey between March and July 2014 using a stratified two-step sample. Differences between groups are described using Pearson chi-square tests and analysis of variance. Response rate was 58.7% (1,515/2,581). Over a 12-month period, 7.6% of respondents provided family care for someone with a serious illness (n = 114). They were most often aged 55-74 (36.0%), women (57.9%), worked full-time (42.3%); 31.8% provided at least 10 h of family care each week. Family caregivers of people with serious illness, compared with family caregivers of people with other conditions, provided more medical and nursing care (33.3% vs. 22.5%, p = 0.027), and experienced a higher burden of family caregiving (p = 0.038) but a similarly high meaningfulness of family caregiving. A considerable part of the adult working population provides family care for someone with serious illness. While family caregiving for someone with serious illness shows similarities with family caregiving for people with other conditions in terms of caregiver characteristics and the impact of caregiving on work-life balance and the meaning derived from it, it is also associated with increased burden.

Influence of Cu distribution on the coercivity of Sm(CobalFe0.25Cu0.06Zr0.02)7.5 magnets during microwave aging process

In this study, we investigated the magnetic properties, microstructures, and Cu distribution on the mechanism of coercivity enhancement of Sm(CobalFe0.25Cu0.06Zr0.02)7.5 magnets during two-step and multistage microwave aging treatments. The intrinsic coercivity, Hcj, and maximum energy product, (BH)max, increase from 15.9 to 24.4 kOe and from 24.3 to 26.5 MGOe, respectively, upon transitioning from two-step to multistage aging. The aged alloys possess a cellular microstructure comprising rhombohedral 2:17 R cell phase and hexagonal 1:5 cell boundary phase after a short microwave aging time of 3 h. The multistage aged alloy exhibits a larger cell size, thicker cell walls, and higher Cu concentration in the 1:5 phase than the two-step aged sample. Elemental mapping reveals Cu atom enrichment at the cell boundary centers and at the 2:17 R/1:5 triple junction of the cell boundary phase, but depletion at the cell boundary edges during multistage aging. First-principles calculations indicate that Cu atoms occupy the Co(2c) sites of a SmCo5 unit cell and weaken not only the strong Co(2c)–Co(3g) direct exchange-coupling interaction, but also the exchange-coupling interaction between Sm 4f and Co 3d electrons, giving rise to low 1:5 phase exchange constant values. Furthermore, the enrichment of Cu atoms at the center of the cell wall and at the 2:17 R/1:5 triple junction of the cell boundary phase produces a gradual increase in the domain wall energy difference as a function of the edge-to-center distance in the cell boundary phase. This enrichment enhances the domain wall pinning strength and increases coercivity. A high concentration and optimal distribution of Cu are critical for achieving a high Hcj. The results of this study provide theoretical guidance to future studies on the mechanism controlling the coercivity of Sm2Co17-based permanent magnets.

The determinants of export behaviour: a study of food processing industry in India

The study examines the factors that determine the export behaviour of food processing firms operating in India based on data from the Centre for Monitoring Indian Economy (CMIE) Prowess Database for 2011-2016. It examines the effect of three technological variables on the export behaviour of firms: information and communication technology (ICT) investment, in-house research and development (R&D), and the import of embodied technologies. Further, it tries to understand if the technological activities of affiliated firms have any effect on their export performance. Using the Heckman two-step sample selection analysis, it reveals that technology investments are important determinants for the export behaviour of firms. Other control variables like firm size, firm age, and the capital-labour ratio were also found to be important in determining the export behaviour of food processing firms. The study gives policy suggestions for improving the global competitiveness of firms in this industry.

Effect of two-step cryorolling and aging on mechanical and electrical properties of a Cu–Cr–Ni–Si alloy for lead frames applications

Abstract High strength and high electrical conductivity Cu–Cr-based alloys are important candidate materials for preparing lead frames, railway contact wires and connects. However, high strength and high electrical conductivity are mutually exclusive in nature. To optimize the mechanical and electrical properties of a Cu–Cr–Ni–Si alloy, the two-step cryorolling and aging (CRA) was carried out in this study. Here, by the analysis of microstructure evolution, precipitation behavior, and corresponding strengthening mechanisms of the alloy at different rolling and aging conditions, we find that the yield strength and electrical conductivity for the two-step CRA sample are improved by 91.3 MPa and 4.9 %IACS (International Annealed Copper Standard) than those of the sample subjected to single-step room temperature rolling and aging (RTRA). The higher yield strength for the two-step CRA sample is mainly determined by the precipitation strengthening contributed from Ni2Si and Cr precipitates, rather than the twin boundary strengthening and dislocation strengthening. In fact, the dislocation density of the two-step CRA sample is lower than that of the single-step RTRA sample because of the faster drop rate of recovery, which weakens the effect of dislocation strengthening. The combination of the first-step aging and second-step CR process for the two-step CRA sample contributes to the formation of more precipitates from the matrix, thereby significantly increasing the precipitation strengthening and decreasing the impurity scattering in the final aging. The results provide a new preparation process and a reliable theoretical approach for optimizing the mechanical and electrical properties of the Cu–Cr alloys.

Health, Sexual Activity, and Sexual Satisfaction.

Physical and mental health disorders can markedly affect sexual activity and sexual satisfaction. Until now, there has been a lack of representative data from the German population that could serve as a basis for demonstrating specific links between sexuality and health. In the German Health and Sexuality Survey (GeSiD), 4955 men and women in a two-step stratified random sample drawn from local resident registration offices across Germany were surveyed by questionnaire. The response rate was 30.2%. The questions involved, among other things, the respondents' selfassessment of sexual activity and sexual satisfaction, general health status, and any chronic illnesses or disabilities. The percentage of respondents who reported having been sexually active with one partner in the past four weeks was highest among men aged 36 to 45 and women aged 26 to 35. In male respondents, the likelyhood of having been sexually active in the past four weeks decreased from 79.1% to 59.0% and 30.1%, respec - tively, if they described their own health status as fair or bad compared with those in very good health. In female respondents the decrease was from 72.5 % to 48.0 % and 32.4%, respectively. A comparable association was found with self-reported chronic illnesses and disabilities that were perceived as impairing sexuality. Sexual satisfaction was present to a similar extent in all age groups. Self-reported physical and mental illnesses are often associated with marked impairment of sexuality. Even illnesses without any immediately evident relation to sexuality often affect aspects of sexual health. It is, therefore, advisable for physicians to address their patients' sexuality routinely in everyday clinical practice.

Depth profiling of surface damage in n-type GaN induced by inductively coupled plasma reactive ion etching using photo-electrochemical techniques

Depth profiling of the dry-etching damage in n-type GaN induced by inductively coupled plasma reactive ion etching was carried out by cyclical electrochemical impedance spectroscopy (EIS) measurements and damage-free photo-electrochemical (PEC) etching. The GaN samples were dry-etched under different etching bias power (Pbias) conditions, and PEC etchings were conducted in increments of 10 nm after EIS measurements. The damage depth was determined to be less than 50 nm for the sample corresponding to Pbias = 30 W and was less than 10 nm for the two-step etching sample corresponding to Pbias = 30 W + 5 W.