Total Coefficient Of Variation Research Articles

Development of a quantitative thyroid-stimulating hormone assay system for a benchtop digital ELISA desktop analyzer.

Regular checkups for thyroid-stimulating hormone (TSH) levels are essential for the diagnosis of thyroid disease. The enzyme-linked immunosorbent assay (ELISA) technique is a standard method for detecting TSH in the serum or plasma of hospitalized patients. A recently developed next-generation ELISA, the digital immunoassay (d-IA), has facilitated detection of molecules with ultra-high-sensitivity. In this study, we developed a TSH assay system using the d-IA platform. By utilizing the ultrasensitivity of d-IA, we were able to use a sample volume of as little as 5µL for each assay (the dead volume was 5µL). The limits of blank, detection, and quantification (i.e., functional sensitivity), were 0.000346, 0.001953, and 0.002280μIU/mL, respectively, and the precision of the total coefficient of variation did not exceed 10%. The correlation between serum and plasma levels indicated good agreement. Thus, our system successfully measured TSH using d-IA with a small sample volume and equal functional sensitivity to the current third generation like ARCHITECT TSH assay, which has a functional sensitivity of 0.0038μIU/mL.

Simultaneous quantitation of multiple myeloma related dietary metabolites in serum using HILIC-LC-MS/MS.

Recent studies from targeted and untargeted metabolomics have consistently revealed that diet-related metabolites, including carnitine (C0), several species of acylcarnitines (AcyCNs), amino acids, ceramides, and lysophosphatidylcholines (LPCs) may serve as potential multiple myeloma (MM) biomarkers. However, most of these approaches had some intrinsic limitations, namely low reproducibility and compromising the accuracy of the results. This study developed and validated a precise, efficient, and reliable liquid chromatography tandem mass spectrometric (LC-MS/MS) method for measuring these 28 metabolic risk factors in human serum. This method employed isopropanol to extract the metabolites from serum, gradient elution on a hydrophilic interaction liquid chromatographic column (HILIC) for chromatographic separation, and multiple reaction monitor (MRM) mode with positive electrospray ionization (ESI) for mass spectrometric detection. The correlation coefficients of linear response for this method were more than 0.9984. Analytical recoveries ranged from 91.3 to 106.3%, averaging 99.5%. The intra-run and total coefficients of variation were 1.1-5.9% and 2.0-9.6%, respectively. We have simultaneously determined the serological levels of C0, several subclasses of AcyCNs, amino acids, ceramides, and LPCs within 15 min for the first time. The established LC-MS/MS method was accurate, sensitive, efficient, and could be valuable in providing insights into the association between diet patterns and MM disease and added value in further clinical research.

Reply to commentary on "Is pre-heat necessary for the measurement of 8-oxo-7,8-dihydroguanosine and 8-oxo-7,8-dihydro-2'-deoxyguanosine in urine samples".

We thoroughly read the commentary from Hu et al. to our recently published article1 and admire their work. To make our point clearer and better clarify the necessity of pre-heat for the accurate measurement of 8- oxo- 7,8-dihydroguanosine (8-oxoG) and 8- oxo- 7,8- dihydro- 2′ - deoxyguanosine (8-oxodG) in urine samples, we give some more detailed instructions, which are shown as follows. (1) The conclusion “it is not always necessary to pre-heat the frozen urine samples to release 8-oxoG and 8-oxodG from precipitates” is only applicable for some certain situations, such as thorough mix and dilution (e.g., at least five times).2 As we emphasized in the article,1 this necessary is closely associated with different pretreatment and determination methods. Our results showed that 11-times dilution can completely release 8-oxoG and 8-oxodG from precipitations. Thus, whether the process of heating urine samples for the release of 8-oxoG and 8-oxodG is necessary and the specific details should be evaluated under different pretreatment and determination methods. (2) Thorough mix of the precipitations in urine samples after thawing is very important, which could guarantee that each aliquot (10 μL in this study) has a homogeneous distribution of precipitations in suspension. We would thoroughly successively mix all urine samples after thawing without discarding precipitations, and specially vortex it for several seconds before we draw every sample for further dilution. We have established and applied the LC–MS/MS method for the determination of 8-oxoG and 8-oxodG in urine samples for several years,3 and more than two researchers in our laboratory have validated the unnecessary to pre-heat the frozen urine samples to release 8-oxoG and 8-oxodG from precipitations with this pretreatment methods. Moreover, the verification results including precision were deemed good with the total coefficient of variation <4.2%.4 (3) To obtain more accurate measurements, the fresh urine samples without precipitations are first recommended. In this study1 and previous reports,3, 4 we all collected fresh urine samples, which are drawn at the same day; thus, there were no apparent precipitations in the urine samples before aliquoted and frozen. After storing at −20°C or − 80°C, most of them would form precipitations. Since diluting the urine 11-times with ddH2O containing 1% formic acid could completely release 8-oxoG and 8-oxodG from urinary precipitations, it is very important to thoroughly mix the samples and draw the needed volume with homogeneous distribution of precipitations as soon as possible. If the operators could not guarantee their performance and heater instruments are available, they should pre-heat the frozen urine samples to completely release the 8-oxoG and 8-oxodG from precipitations. This work was funded by research grants from the National High Level Hospital Clinical Research Funding (2022-PUMCH-B-073) and National High Level Hospital Clinical Research Funding (2022-PUMCH-A-138). The authors declare that they have no competing interests.

Coordinated variation in root and leaf functional traits of Hippophae rhamnoides treated at different stump heights in feldspathic sandstone areas of Inner Mongolia.

This study was aimed to clarify the effects of stumping on root and leaf traits as well as the tradeoffs and synergies of decaying Hippophae rhamnoides in feldspathic sandstone areas, and to select the optimal stump height that contributed to the recovery and growth of H. rhamnoides. variations and coordination between leaf traits and fine root traits of H. rhamnoides were studied at different stump heights (0, 10, 15, 20cm, and no stumping) in feldspathic sandstone areas. All functional traits of the leaves and roots, except the leaf C content (LC) and the fine root C content (FRC), were significantly different among different stump heights. The total variation coefficient was the largest in the specific leaf area (SLA), which is therefore the most sensitive trait. Compared to non-stumping, SLA, leaf N content (LN), specific root length (SRL) and fine root N content (FRN) all improved significantly at stump height of 15cm, but leaf tissue density (LTD), leaf dry matter content (LDMC), leaf carbon to nitrogen ratio (LC : LN), fine root tissue density (FRTD), fine root dry matter content (FRDMC) and fine root carbon to nitrogen ratio (FRC : FRN) all decreased significantly. The leaf traits of H. rhamnoides at different stump heights follow the leaf economic spectrum, and the fine roots show a similar trait syndrome to the leaves. SLA and LN are positively correlated with SRL and FRN and negatively with FRTD and FRC : FRN. LDMC and LC : LN are positively correlated with FRTD and FRC : FRN, and negatively correlated SRL and RN. The stumped H. rhamnoides changes to the 'rapid investment-return type' resource trade-offs strategy, and the growth rate is maximized at the stump height of 15cm. Our findings are critical to the prevention and control of vegetation recovery and soil erosion in feldspathic sandstone areas.

Partial validation of the Vcheck canine pancreatic lipase assay.

Measurement of canine pancreatic lipase immunoreactivity (cPLI) is used for diagnosing pancreatitis in dogs. Because pancreatitis can be a life-threatening disease with severe complications, an in-house cPLI test would be valuable to obtain rapid test results. The aim of this study was to evaluate a point-of-care cPLI test, Vcheck cPL. Precision, determined according to EP15, and linearity under dilution were determined and judged against preset quality goals. Results from the Vcheck cPL were compared with a previously validated cPLI ELISA, Spec cPL. In a retrospective study, cPLI results from dogs with and without acute pancreatitis, as determined by pancreatic ultrasound examination, were investigated to assess the performance of the assay in a clinical setting. Statistical analysis included the Mann-Whitney test, Chi-square test, and Passing-Bablok regression analysis with a significance level of 0.05. Precision of the assay was acceptable, with intra-, inter-, and total coefficients of variation (CV%) less than 12.1%, 6.4%, and 12.1%, respectively. Results from the linearity study indicated that the method was acceptably linear at lower concentrations but not in the high-concentration range. The method comparison study revealed that Vcheck generally measured higher concentrations compared with Spec cPL, and that the methods should not be used interchangeably. Dogs with acute pancreatitis had significantly higher cPLI concentrations compared with dogs without pancreatitis (P < 0.01), but there was a marked overlap in cPL concentrations between the two groups.

Development and application of a high accuracy method for measuring Pb in blood

BackgroundLead (Pb) is an environmental pollutant, and human exposure is assessed by measuring blood Pb concentrations. Today, the use of Pb-based products is restricted, but the health effects of low Pb concentrations require investigation. For this, a high-accuracy method is needed. Here, we report a new inductively coupled plasma mass spectrometry (ICP-MS) method for quantifying Pb in blood. MethodsBlood samples and calibrators were gravimetrically spiked with bismuth as an internal standard, digested with ultrapure nitric acid, and diluted 100 times. The calibrators were made from high-purity reference materials supplied by the National Institute of Standards and Technology, and the sample concentration was measured using the calibration bracketing method. The analytical performance and application of the new method were investigated. ResultsThe method had good specificity and a negligible matrix effect. The intra- and inter-assay coefficients of variation (CVs) were <1.34 % and <1.88 %, respectively, and the total CV was <1.74 %. The range of recovery was 98.9 %–99.4 %. SRM955d, BCR-635, and BCR-636 were within the certified intervals, with mean relative bias values of 0.93 %, −0.13 %, and −0.56 %, respectively. The limit of quantification was 1.1 µg/l. The method was used to assign target values to external quality assessment samples and to assess routine methods, showing good results at high concentrations. However, at low concentrations, two laboratories did not pass. ConclusionsThis ICP-MS method is a simple, accurate method and can be a candidate reference method for blood Pb measurement. But more research will be needed to verify this.

Reference change value of global longitudinal strain in clinical practice: A test-rest quality implementation project.

Reference change value (RCV) is used to assess the significance of the difference between two measurements after accounting for pre-analytic, analytic, and within-subject variability. The objective of the current study was to define the RCV for global longitudinal strain (GLS) using different semi-automated software in standard clinical practice. Using a test-retest study design, we quantified the median coefficient of variation (CV) for GLS using AutoStrain and Automated Cardiac Motion Quantification (aCMQ) by Philips. Triplane left-ventricular ejection fraction (LVEF) was measured for comparison. Multivariable regression analysis was performed to determine factors influencing test-retest CV including image quality and the presence of segmental wall motion abnormalities (WMA). RCV was reported using a standard formula assuming two standard deviations for repeated measurements; results were also translated into Bayesian probability. Total measurement variation was described in terms of its three different components: pre-analytic (acquisition), analytic (measuring variation), and within-subject (biological) variation. Of the 44 individuals who were screened, 41 had adequate quality for strain quantification. The mean age of the cohort was 56.4±16.8 years, 41% female, LVEF was 55.8±9.8% and the median and interquartile range for LV GLS was -17.2 [-19.3 to -14.8]%. Autostrain was more time efficient (80% less analysis time) and had a lower total median CV than aCMQ (CV=7.4%vs. 17.6%, p <.001). The total CV was higher in patients with WMA (6.4%vs. 13.2%, p=.035). In non-segmental disease, the CV translates to a RCV of 15% (corresponding to a probability of real change of 80%). Assuming a within-subject variability of 4.0%, the component analysis identified that inter-reader variability accounts for 3.7% of the CV, while acquisition variability accounts for 4.0%. Using test-retest analysis and CVs, we find that an RCV of 15% for GLS represents an optimistic estimate in routine clinical practice. Based on our results, a higher RCV of 17%-21% is needed in order to provide a high probability of clinically meaningful change in GLS in all comers. The methodology presented here for determining measurement reproducibility and RCVs is easily translatable into clinical practice for any imaging parameter.

A robust LC-MS/MS method to measure 8-oxoGuo, 8-oxodG, and NMN in human serum and urine

ObjectiveTo establish and validate a robust LC-MS/MS method for simultaneously measuring 8-oxoGuo, 8-oxodG, and NMN in serum and urine to evaluate the oxidative stress status. MethodsA Waters TQ-XS triple quadrupole mass spectrometer system coupled with an Acquity UPLC Primer HSS T3 column was chosen. The clinical performance was verified according to the CLSI C62-A and EP-15 guidelines. Furthermore, matched serum and urine samples from 22 apparently healthy check-ups, 20 patients with atherosclerosis, and 18 individuals with dementia were evaluated. ResultsThe recovery for serum 8-oxoGuo, urine 8-oxoGuo, serum 8-oxodG, urine 8-oxodG, serum NMN, and urine NMN was 88.8–112.4%, 102.4–114.1%, 88.5–107.7%, 94.9–102.6%, 98.4–108.9%, and 88.5–108.6%, respectively. Based on the inter-assay results, total coefficient of variation, matrix effect, and carryover, the LC-MS/MS method was deemed robust. The limit of quantification was 0.017, 0.018, and 0.150 nmol/L for 8-oxoGuo, 8-oxodG, and NMN, respectively, which are suitable for accurate measurements in human serum and urine samples. Higher 8-oxoGuo and 8-oxodG levels and lower NMN levels, indicative of significantly higher oxidative stress status, were found in patients with dementia compared to healthy subjects. ConclusionWe established and validated a robust LC-MS/MS method to simultaneously measure 8-oxoGuo, 8-oxodG, and NMN in serum and urine.

Quantification of the anti-neoplastic polyacetylene falcarinol from carrots in human serum by LC-MS/MS

Falcarinol is a polyacetylene which is found in carrots and known to have anti-neoplastic properties in rodents. Research in the bioactivity of falcarinol in humans require methods for quantification of falcarinol in human serum. Here we report the development of an LC-MS/MS method and its use to measure serum falcarinol concentrations in humans following intake of a carrot product. Falcarinol was measured by LC-MS/MS using the m/z 268 to m/z 182 mass transition. Six calibrator levels (0.2–20 ng/mL) and 3 control levels (0.4, 2 and 8 ng/mL) were prepared by addition of falcarinol to human serum pools. Linearity of the developed method was good with a mean R2 of 0.9942. Within-day, between-day and total coefficients of variation were 6.9–13.1%, 4.1–5.0% and 8.1–14.0%, respectively. The limits of detection and quantitation were 0.1 and 0.2 ng/mL, respectively, matrix effects 84.2%, recovery 101.4–105.4% and carry-over −0.24–0.07%. Serum falcarinol concentrations were measured in 18 healthy volunteers prior to and at 9 time-points following intake of a carrot product. Falcarinol concentrations peaked at the 1-hour time-point after intake in 15 out of 18 volunteers and declined according to a single exponential decay function with an aggregate t½ of 1.5 h.In conclusion, an LC-MS/MS method for quantification of falcarinol in human serum with acceptable performance was developed and used to measure falcarinol concentrations following intake of a carrot product.

Real world hypoglycaemia related to glucose variability and Flash glucose scan frequency assessed from global FreeStyle Libre data.

Flash glucose monitoring provides a range of glucose metrics. In the current study, we aim to identify those that indicate that glycaemic targets can be consistently met and contrast the total (t-CV) and within-day coefficient of variation (wd-CV) to guide the assessment of glucose variability and hypoglycaemia exposure. De-identified data from Flash readers were collected. The readers were sorted into 10 equally sized groups of scan frequency followed by quartiles of estimated A1c (eA1c). A similar grouping was performed for the total coefficient of variation (t-CV) and within-day coefficient of variation (wd-CV). In addition, analysis of the association of time below 54 mg/dl and glucose variability measured by t-CV and wd-CV was performed. The dataset included 1002 946 readers. Readers sorted by 10 equal groups of scan rate and quartiles by eA1c, t-CV and wd-CV represented 25 074 readers per group. The association of lower eA1c with higher time in range and reduced time above range was clear. The correlation of eA1c quartiles and time below range was not consistent. An association between glucose variability and hypoglycaemia was found. Both wd-CV and t-CV were associated with time below range. For achieving the consensus target of <1% time below 54 mg/dl, the associated wd-CV and t-CV values were 33.5% and 39.5%, respectively. The type of CV reported by the different continuous glucose monitoring systems should be acknowledged. CV <36% might not be adequate to ensure low hypoglycaemia exposure. To our knowledge, the majority of continuous glucose monitoring reports the t-CV. Appropriate thresholds should be used to identify patients that would probably meet time below range targets (t-CV <40% or wd-CV <34%).

Comparison of four different immunoassays and a rapid isotope-dilution liquid chromatography-tandem mass spectrometry assay for serum folate

Accurate measurement of serum folate is essential for the diagnosis and management of various disorders. This study aims to investigate the between-method differences of four immunoassays and a rapid isotope-dilution liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS) method. Roche Cobas (USA), Abbott Alinity i2000 (USA), Beckman Coulter Access (USA), Mindray CL-6000i (China), and the ID-LC-MS/MS method were compared using 46 human serum samples. The results were analysed by Passing-Bablok regressions and Bland-Altman plots. A bias of 13.31% based on biological variation was used as the bias criterion. All the within-run and total coefficients of variation (CVs) met the specification. The folate concentrations determined by all the assays were significantly different (p=0.0028). All assays had correlation coefficients over 0.97 with each other. The 95% confidence intervals (CIs) for the slope seldom contained 1 and few 95% CIs for the intercept contained 0 in the regression equations. Compared to ID-LC-MS/MS, the biases of all assays ranged from-20.91 to13.56nmol/L, and the mean relative biases ranged from-9.85 to 40.33%. The predicted mean relative biases at the medical decision levels rarely met the criterion. Assays for serum folate had good correlations with each other but lacked good agreement. The accuracy and consistency of assays for serum folate should be measured and assessed routinely. Standardization work to improve the accuracy of serum folate assays, such as the extension of traceability to reference methods or materials, calibration standardization efforts, and assay-adjusted cut-offs should be promoted.

Evaluation of meibomian gland morphology and anterior segment parameters by Sirius topography systems in polycystic ovary syndrome.

To compare findings in specular microscopy, corneal topography, and noncontact meibography in polycystic ovary syndrome (PCOS) patients with healthy controls. A total of 40 women with PCOS and 32 healthy controls were enrolled in the study. Schirmer's test, Ocular Surface Disease Index (OSDI), noninvasive tear break-up time (NITBUT), the mean keratometry (Km), maximum keratometry (Kmax), central (CCT), thinnest (TCT) and apical (ACT) corneal thicknesses, meibomian gland (MG) loss, meiboscores, morphology of MGs, endothelial cell density (ECD), coefficient of variation (CV), and percentage of hexagonal cells (PHEX) were analyzed. Correlations between anti-Mullerian hormone (AMH) and sex hormones and the findings of PCOS patients were evaluated. Mean OSDI score, intraocular pressure, Km and Kmax values, the mean MG loss of upper and lower eyelids, lower eyelid meiboscore, and degree of morphological abnormalities of MGs were higher in cases with PCOS than healthy controls. There were no significant differences between groups in Schirmer results, first and average NITBUT, mean values of TCT, ACT, CCT, ECD, CV, and PHEX (P > 0.05, for all). There were correlations between plasma AMH level and Kmax, back Km and PHEX, and between estradiol (E2) and PHEX; there were negative correlations between E2 and total MG loss and CV and between total testosterone and ACT. Loss and morphological deterioration of the MGs are observed in PCOS patients, even if the tear parameters are not impaired yet. In eyes with PCOS, keratometry values become steeper in proportion to AMH levels. The PCOS patients should be followed carefully for the development of corneal ectasia.

Multisite reproducibility and test-retest reliability of the T1w/T2w-ratio: A comparison of processing methods

BackgroundThe ratio of T1-weighted (T1w) and T2-weighted (T2w) magnetic resonance imaging (MRI) images is often used as a proxy measure of cortical myelin. However, the T1w/T2w-ratio is based on signal intensities that are inherently non-quantitative and known to be affected by extrinsic factors. To account for this a variety of processing methods have been proposed, but a systematic evaluation of their efficacy is lacking. Given the dependence of the T1w/T2w-ratio on scanner hardware and T1w and T2w protocols, it is important to ensure that processing pipelines perform well also across different sites. MethodsWe assessed a variety of processing methods for computing cortical T1w/T2w-ratio maps, including correction methods for nonlinear field inhomogeneities, local outliers, and partial volume effects as well as intensity normalisation. These were implemented in 33 processing pipelines which were applied to four test-retest datasets, with a total of 170 pairs of T1w and T2w images acquired on four different MRI scanners. We assessed processing pipelines across datasets in terms of their reproducibility of expected regional distributions of cortical myelin, lateral intensity biases, and test-retest reliability regionally and across the cortex. Regional distributions were compared both qualitatively with histology and quantitatively with two reference datasets, YA-BC and YA-B1+, from the Human Connectome Project. ResultsReproducibility of raw T1w/T2w-ratio distributions was overall high with the exception of one dataset. For this dataset, Spearman rank correlations increased from 0.27 to 0.70 after N3 bias correction relative to the YA-BC reference and from -0.04 to 0.66 after N4ITK bias correction relative to the YA-B1+ reference. Partial volume and outlier corrections had only marginal effects on the reproducibility of T1w/T2w-ratio maps and test-retest reliability. Before intensity normalisation, we found large coefficients of variation (CVs) and low intraclass correlation coefficients (ICCs), with total whole-cortex CV of 10.13% and whole-cortex ICC of 0.58 for the raw T1w/T2w-ratio. Intensity normalisation with WhiteStripe, RAVEL, and Z-Score improved total whole-cortex CVs to 5.91%, 5.68%, and 5.19% respectively, whereas Z-Score and Least Squares improved whole-cortex ICCs to 0.96 and 0.97 respectively. ConclusionsIn the presence of large intensity nonuniformities, bias field correction is necessary to achieve acceptable correspondence with known distributions of cortical myelin, but it can be detrimental in datasets with less intensity inhomogeneity. Intensity normalisation can improve test-retest reliability and inter-subject comparability. However, both bias field correction and intensity normalisation methods vary greatly in their efficacy and may affect the interpretation of results. The choice of T1w/T2w-ratio processing method must therefore be informed by both scanner and acquisition protocol as well as the given study objective. Our results highlight limitations of the T1w/T2w-ratio, but also suggest concrete ways to enhance its usefulness in future studies.

A standard addition method to quantify serum lithium by inductively coupled plasma mass spectrometry.

Therapeutic monitoring of lithium (Li) is important because of its narrow therapeutic range and therapeutic index. Here, the authors present the evaluation of an accurate method for the determination of lithium in serum. Serum samples were diluted with 0.3% ultrapure nitric acid and were spiked with an internal standard germanium (Ge). The Li/Ge ratio was detected in He mode; we utilized standard addition method to quantify lithium in human serum. The new inductively coupled plasma mass spectrometry (ICP-MS) assay was characterized for linearity, specificity, imprecision, trueness, accuracy, and comparison. The correlation coefficients (r) of linearity were all > 0.9999. The specificity proved to be good. The total coefficients of variation (CV) were 1.11% and 0.49% for the two serum samples. The mean bias from target values of standard reference materials (SRM 956d) was -0.71% for Level I, -017% for Level II, and 2.20 for Level III. External Quality Assessment Scheme for Reference Laboratories in Laboratory Medicine (RELA) gave satisfied results for the new method. Comparison with the ion-selective electrode routine method got reasonable results. This high accuracy method is an attractive alternative for lithium measurement and can be used as a candidate reference method to improve quality of serum lithium in China.

Occurring the paraquat residues in various Nigerian

Abstract: Because pesticides are ubiquitous and focused on the food chain, they are a substantial source of possible environmental danger to humans and animals. The highest extremely hazardous herbicides to be launched in 60 years ago is paraquat (1,1'-dimethyl-4,4'-bipyridylium dichloride). Although most nations have prohibited or severely limited the use of paraquat (PQ), it is still used in others, particularly in Nigeria. As a result, the present study used a spectrophotometric method to look for PQ in some vegetables, crops, and fruits in Abeokuta that are commonly consumed. The presence and amounts of PQ residues were assessed in 150 samples of various vegetables, crops, and fruits (harvested at Fadama field of the University of Agriculture, Abeokuta). PQ residues were detected in Talinum triangulare, Corchorus olitorius, Amaranthus caudatus, Cratylia argentea, Capsicum frutescens, Lycopersicum esculentum, Raphanus sativus, Zea may, and Dioscorea alata at 0.130.02, 0.27+0.02, 0.06+0.01, 0.10+0.03, 0.15+0.03, 0. Using this technique, no paraquat residues were found in M. paradisicica or C. papaya. At 0.05-1.0 ppm fortification level, the technique yields a mean recovery of over 80% and is reproducible with a total coefficient of variation of 8.0 percent (n=10). All residual levels found were under the PQ tolerance or pesticide maximum limits.

A highly sensitive, simple, and fast gas chromatography-mass spectrometry method for the quantification of serum short-chain fatty acids and their potential features in central obesity.

Short-chain fatty acids (SCFAs) are the main gut microbe metabolites, which have no more than six carbons. SCFAs are an emerging biomarker in metabolic diseases, including central obesity. Commonly, SCFAs are measured in fecal samples, where they are highly abundant, but here they do not reflect direct interactions with related organs. Serum SCFAs are assumed to be more associated with metabolic disease than fecal SCFAs, albeit at very low concentrations. The aim of the present study is to develop a highly sensitive, simple, and fast method for measuring six SCFAs in the serum by gas chromatography-mass spectrometry (GCMS). The serum is mixed with meta-phosphoric acid and 2,2-dimethylbutyric acid, followed by homogenization and centrifugation. Supernatant is then injected into the fused silica capillary column. The method is linear from 0.12-500μmol/L for all SCFAs with an accuracy of 90-117%. The total coefficient of variation for precision ranges from 3.8 to 14.1%. A preliminary study is performed with 32 centrally obese subjects and 17 lean subjects. The mean values of all SCFAs, including acetic, propionic, isobutyric, butyric, isovaleric, and valeric acid, in the centrally obese subjects are significantly higher compared with lean subjects. A significant correlation also exists between all SCFAs, with the waist circumference indicating that serum SCFAs have potential features with respect to metabolic diseases, especially central obesity. The validated GCMS method provides highly sensitive, fast, simple, and reliable SCFA quantitation in the serum and demonstrates the potential features of circulating SCFAs in central obesity.

Analytical and Clinical Performance of a Liquid Chromatography-Tandem Mass Spectrometry Method for Measuring Gastrin Subtypes G34 and G17 in Serum.

Two major forms of gastrin, gastrin-17 (G17) and gastrin-34 (G34), exist in blood. However, conventional immunoassay methods can only quantify total gastrin or G17 alone. Here, we aimed to establish a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify G17 and G34 simultaneously. Serum samples were prepared by anion-exchange solid-phase extraction. The analytical performance of the LC-MS/MS method was validated and the method was compared to chemiluminescence immunoassay (CLIA) and radioimmunoassay (RIA). The G17 and G34 concentrations in 245 serum samples from healthy controls, individuals with gastrinoma, and individuals with other diseases were analyzed. The total runtime of the LC-MS/MS method was 6 min. No substantial matrix effect was observed with internal standard correction. The intraassay coefficients of variation (CVs) for G17 and G34 were 4.0%-14.2% and 4.4%-10.4%, respectively, and total CVs were 5.2%-14.1% and 4.6%-12.4%, respectively. The correlation coefficient between LC-MS/MS and CLIA was 0.87, and between LC-MS/MS and RIA was 0.84. The G17+G34 concentrations for 87.5% of individuals with gastrinoma were higher than the 95th percentile of healthy controls (18.1 pg/mL), whereas the concentrations for individuals with other diseases and gastrinoma overlapped. Based on the Youden indices calculated for G17+G34, G34, and G17, the most specific biomarker was G17 (96.9% clinical specificity at 209.8 pg/mL) for gastrinoma. This method should aid in the diagnosis of diseases associated with increased gastrin concentrations.

Application of genetic screening processor in screening neonatal glucose-6-phosphate dehydrogenase deficiency.

To evaluate the performance of genetic screening processor (GSP analyzer) in neonatal screening for glucose-6-phosphate dehydrogenase (G6PD)deficiency. The accuracy and precision of GSP analyzer was evaluated with the control materials from National Center for Clinical Laboratories and the low and high quality G6PD control kit (fluorescence analysis). GSP analyzer and semi-automatic fluorescence immunoanalyzer (1420 analyzer) were simultaneously used to detect 2622 neonatal screening samples and 41 confirmed samples to analyze the correlation and consistency of the test results; 78 floating samples and 78 non-floating samples were detected to compare the result. A total of 1 100 384 neonatal screening samples from January 2017 to December 2018 and 855 856 neonatal screening samples from January 2019 to December 2020 were detected with 1420 analyzer and GSP analyzer, respectively. Referring to the percentile method and the expert consensus, the new cut-off value of GSP analyzer for G6PD deficiency in screening was established. The relative bias of GSP analyzer in detecting G6PD was 0.71%-4.23%; the intra assay precision was 4.34%-4.91%, the inter assay precision was 0.85%-2.12%, and the total coefficient of variation was 5.44%-5.72%. There was a significant positive correlation between G6PD activity detected by GSP analyzer and 1420 analyzer (=0.740, <0.01). Forty-one clinical confirmed patients were identified by both 1420 analyzer and GSP analyzer (=0.945). The G6PD activity in floating dry blood spots detected by 1420 analyzer was significantly lower than that in non-floating dry blood spots (<0.05), but there was no significant difference in G6PD activity between floating and non-floating dry blood spots detected by GSP analyzer (>0.05). The sensitivities of GSP analyzer and 1420 analyzer in screening G6PD deficiency were both 100.00%, and the specificities were both more than 99.80%. Compared with 1420 analyzer, the positive predictive value, positive rate and prevalence of G6PD deficiency detected by GSP analyzer were increased, and the false positive rate was decreased (all <0.01). The new cut-off value was 26.1 U/dL for male and 29.1 U/dL for female according to the 99.1% percentile of the population. GSP analyzer has better detection performance with high automation, efficiency and throughput, which can be used in large-scale screening for neonatal G6PD deficiency.

Total IgE Variability Is Associated with Future Asthma Exacerbations: A 1-Year Prospective Cohort Study

Few prospective studies have investigated the relationship between IgE variability and risk for asthma exacerbations (AEs). To explore the relationship between IgE variability and AEs. Recruited patients with stable asthma underwent two serum total IgE tests within a month (at screening [baseline IgE] and at 1 month) to obtain the coefficient of variation (CV) of base 10 log-transformed IgE. Patients with IgE CV were divided into IgE CV-high and IgE CV-low cohorts based on the CV median and were observed within 12 months, during which the association between IgE variability and AEs was explored using a negative binomial regression model. The IgE CV levels obtained from 340 patients classified patients into two groups (n= 170 for the IgE CV-high and IgE CV-low groups, respectively) based on the serum total IgE CV median of 2.12% (quartiles 1 and 3: 0.98% and 3.91%, respectively). The IgE CV-high patients exhibited worse asthma control and lung function and more marked airway inflammation, and received more intensive medication use compared with IgE CV-low patients. The IgE CV-high patients exhibited increased rates of moderate-to-severe (adjusted rate ratio= 2.88; 95% confidence interval, 1.65-5.03; P < .001) and severe (adjusted rate ratio= 2.16; 95% confidence interval, 1.08-4.32; P= .029) AEs during the follow-up year compared with IgE CV-low patients. Furthermore, sputum IL-6 partially mediated the associations between IgE CV with moderate-to-severe and severe AEs. Variability in total serum IgE levels is an easily obtained and practical measure for predicting AEs. Future studies are needed to investigate whether IgE variability can be used to guide precision medicine in asthma.

Cover crops decrease maize yield variability in sloping landscapes through increased water during reproductive stages

• Weather and landscape position were the major drivers of simulated maize yield. • Rye cover crop residue increased soil water storage at critical growth stages. • Rye cover crops decreased the total coefficient of variation in yield from 33−26%. • During dry years, rye cover crops increased maize yields on slopes by 24 %. Rolling hill style topography is a common feature of agricultural land throughout the United States. Topographic complexity causes subfield variation in soil resources such as water and nutrients, leading to a mosaic of high- and low-productivity zones that can shift from one year to the next due to weather. Stabilizing yields across these productivity zones using agroecological methods may improve land use efficiency, prevent unnecessary cropland expansion, and reduce the environmental impact of these systems. Here, we hypothesized that cover crops may help to reduce soil water and nutrient losses and increase the stability of subsequent maize yields across time and space. We performed a field study to evaluate the effect of a cereal rye ( Secale Cereale L.) cover crop on maize ( Zea mays L.) yield at three landscape positions (summit, backslope, and toeslope) in Central KY in 2018–2019, and calibrated the DSSAT v4.7.0.001 computer simulation program to test our hypothesis across a thirty-year period. Our field trial showed pronounced variability in maize yield across different landscape positions, ranging from 6.3 Mg ha −1 in the backslope, to 12.2 Mg ha −1 in the toeslope. Model simulations were consistent with results from our field trial and indicated that low yields in the backslope were primarily due to water stress, with >10 % yield reductions in 17 out of 30 simulated years relative to simulations under irrigated conditions where water was not limiting. In contrast, the toeslope and summit positions experienced >10 % yield reductions due to water stress in only 6 of the 30 years. Growing a cereal rye cover crop before maize reduced the frequency of water stress and raised maize yields in the backslope by 6% (500 kg ha -1 ) on average, and 24 % (1235 kg ha -1 ) during dry years. The coefficient of variation across all weather conditions and landscape positions was reduced from 33 % to 26 % when maize followed a rye cover crop compared to fallow. The yield benefits of the cover crop were associated with decreased soil evaporation and runoff that increased water availability during anthesis and late maize reproductive phases. Crop model simulations allowed us to evaluate and parse out the fundamental drivers of the interaction between cover crops and complex topography under different weather scenarios. Overall, our study demonstrates the outsized potential of cover crops to increase and stabilize grain yields in rolling hill landscapes and emphasizes the value of cover crops as a tool for ecological intensification.