Cut-off Limit Research Articles

Биологически активные гидролизаты белков молока и их комплексы включения с циклодектринами

Enzymatic hydrolysis of dairy proteins increases their nutritional and biological value while reducing their allergenic potential. The subsequent complexation of peptides with cyclodextrins (CDs) reduces the bitterness of the hydrolyzed proteins. The research objective was to obtain hydrolysates of whey proteins and their cyclodextrin inclusion complexes with peptides, as well as to describe the peptide composition of the cleaved dairy proteins, biological activity, and sensory profile of the hydrolysates and inclusion complexes. The research featured enzymatic whey protein hydrolysates with an extensive hydrolysis degree and their inclusion complexes with β- and γ-CDs. Dairy proteins were hydrolyzed with alcalase, and the hydrolysates obtained were subjected to micro- and ultrafiltration (cut-off limit – 10 kDa). The peptide composition of the hydrolyzed proteins was determined by the methods of high-performance liquid chromatography and chromatography-mass spectrometry. The antimutagenic activity was evaluated using the Ames test whereas the antibacterial effect was studied with the impedimetric method. The antioxidant activity was detected with fluorimetry and spectrophotometry. The method of competitive enzyme immunoassay was applied to reveal the antigenic properties. The bitterness of the experimental sample s was determined by a sensory evaluation. The research delivered the optimal modes for whey protein cleavage with alcalase that made it possible to achieve efficient micro- and ultrafiltration. The resulting hypoallergenic peptide fractions and their inclusion complexes with β- and γ-CDs possessed antioxidant, antibacterial, and antimutagenic properties. The whey proteolysis and subsequent filtration with/without tindalization demonstrated a 265/589-fold decrease in the residual antigenicity. The fluorimetric method showed a 1.79/1.90-fold increase in the antioxidant activity of the hydrolysate in complexes with β- and γ-CDs. Binding of β-CDs to peptides enhanced their antimicrobial activity against Escherichia coli ATCC 8739 and Staphylococcus aureus ATCC 6538. The hydrolysate samples with β-CDs showed less bitterness. Whey proteolysis with alcalase under optimized conditions and subsequent fractionation resulted in a product with high consumer qualities. Enzymatic hydrolysates of dairy proteins and their CD inclusion complexes were able to substitute native protein components. Their bioactive properties, good taste, and low allergenic potential mean good prospects for the functional food industry.

Development of an immunochromatographic strip for the rapid detection of 11-dehydro-thromboxane B2 in human urine

Urinary 11-dehydro-thromboxane B2 (11-DH-TXB2) is a diagnostic marker for evaluating the effect of aspirin treatment in patients with cardiovascular disease. Therefore, it is of great clinical significance to develop a rapid detection method for 11-DH-TXB2 in urine. In this study, we prepared an ultrasensitive monoclonal antibody (mAb) against 11-DH-TXB2. The mAb belongs to the IgG1 subtype with kappa light chains. In an indirect competitive ELISA, it had a half-maximal inhibitory concentration of 3.63 ng/mL and a high affinity of 5.60 × 109 L/mol for 11-DH-TXB2. We then developed an immunochromatographic strip for point-of-care testing (POCT) of 11-DH-TXB2 in human urine. The strip provides a cut-off value and limit of detection of 500 ng/mL and 31.2 ng/mL, respectively. The result is obtained within 15 min. In summary, the proposed strip is a portable POCT tool for monitoring 11-DH-TXB2 in human urine.

Clearing the Skepticism about Subclinical Hypothyroidism: Is It Beneficial to Treat Patients with Thyroid-Stimulating Hormone >4.5 and <10 mIU/L?

AbstractSubclinical hypothyroidism (SCH) is a heterogeneous clinical condition ranging from asymptomatic to wide variety of clinical manifestations, which are often nonspecific. Being a common laboratory finding, clinicians often face the dilemma of whether to treat or not. Threshold of 10 mIU/L of thyroid-stimulating hormone (TSH) is often used as a cutoff limit to offer treatment. However, still, debate remains on whether to treat less than 10 mIU/L considering special clinical conditions like pregnancy. Whether SCH exists, is screening needed in asymptomatic individuals, is treating asymptomatic cases beneficial or harmful and what threshold level of TSH to be considered for treatment are all potential questions that need to be answered.

Liver Mitochondrial Morphology and Gene Expression as Markers of Liver Reserve: Prognostic Implications for Native Liver Survival in Biliary Atresia

PurposeHepatocyte mitochondrial morphology and gene expression were compared between biliary atresia (BA), infantile cholestasis (IC), and normal liver (NL) as prognostic indicators. MethodsSpecimens of liver at portoenterostomy (PE) for BA, from intrahepatic bile duct paucity patients for IC, and from choledochal cyst or hepatoblastoma patients for NL were collected prospectively (P) beginning in 2021 (P-BA = 11, P-IC = 9, P-NL = 7) and retrospectively (R) from paraffin-embedded tissue going back to 1981 (R-BA = 25, R-IC = 9, R-NL = 4). The P-cohort had transmission electron microscopy (TEM) to image mitochondria, immunoblotting for heat shock protein 60 (HSP60), and quantitative PCR (qPCR) for HSP60 and mitochondrial functional genes. Both cohorts had immunofluorescence for HSP60 quantified as a ratio to albumin-positive hepatocytes (ALB) with HSP60/ALB<1.0 as a cutoff limit using ImageJ. ResultsHSP60 was significantly lower in BA/IC than NL on qPCR (BA: p < 0.01, IC: p < 0.05) and lower in BA than IC/NL on immunoblotting (p < 0.05). HSP60/ALB was significantly lower in BA than NL/IC (p < 0.001). Despite BA subjects being matched for types of BA and ages at PE, HSP60/ALB did not correlate with jaundice clearance (JC; T-Bil<1.2 mg/dL) but was significantly higher in native liver survivors (NLS) after PE compared with liver transplant (LTx) cases (p < 0.05) and significantly lower in LTx cases achieving JC than NLS achieving JC (p < 0.05). TEM showed BA had significantly more mitochondrial inclusion bodies (p < 0.05) and significantly larger cristae (p < 0.01) than IC/NL. qPCR in BA showed significant repression of mitochondrial functional genes for mRNA stabilization and energy facilitation. ConclusionHSP60/ALB correlates with NLS after PE for BA. Level of evidenceII

Detection of cocaine 24 h after administration before nasotracheal intubation.

Cocaine may be applied to decongest the nasal mucosa before nasotracheal intubation, but patients risk a criminal offence if cocaine is detected when patients drive a car shortly after surgery. We aimed to evaluate whether benzoylecgonine levels in saliva exceeded the cut-off point 24 h after administration in patients undergoing nasotracheal intubation and whether cocaine would be detectable above the Danish legal fixed limit in blood samples 1 and 24 h after surgery. We conducted a prospective study following approval from the local research ethics committee and the national medicine agency. Written informed consent was obtained from all patients. We included patients scheduled for surgery under general anaesthesia with nasotracheal intubation. They received 80 mg cocaine as a nasal spray 5 min before induction and nasotracheal intubation. The primary outcome was a dichotomous assessment of benzoylecgonine levels in saliva samples measured 24 h after administration of nasal cocaine with a cut-off limit of 200 ng/mL. Secondary outcomes were dichotomous assessments of cocaine in whole blood samples measured 1 and 24 h after administration of nasal cocaine with a cut-off limit of 0.01 mg/kg. Overall, 70 patients had valid saliva samples and 75 had valid blood samples 24 h after cocaine administration. Benzoylecgonine in saliva was traceable above the cut-off in 9/70 patients (13%; CI95%: 6% to 23%), and cocaine in blood was detected above the cut-off in 2/75 patients (3%; CI95%: 0.3% to 9%). We found benzoylecgonine traceable in saliva in 13% of patients and cocaine traceable in blood in 3% of patients 24 h after administration of 80 mg nasal cocaine. Patients should be informed when receiving cocaine and advised not to drive for at least 24 h.

Association between extremely high prognostic nutritional index and all-cause mortality in patients with coronary artery disease: secondary analysis of a prospective cohort study in China

ObjectivesDecreased prognostic nutritional index (PNI) was associated with adverse outcomes in many clinical diseases. This study aimed to evaluate the relationship between baseline PNI value and adverse clinical outcomes in...

Refining the 30-day emergency department visit after initial chemotherapy administration A-PROACCT model.

e23253 Background: This study attempted to address limitations in the A-PROACCT model developed by Stein et al. (JOP, 2023) to predict the rate of acute care events within 30 days (ACE30) of initial chemotherapy administration (ICA). The four most important predictors were drug category (immunotherapy), cancer type (gastrointestinal), age group ( &lt; 40), and BMI category (underweight). Only linear relationships were considered. For patients for whom an ACE30 was predicted by their first and second models, only 0.18 and 0.23 actually had an ACE30, a low positive predictive value (PPV). We attempted to improve the PPV with advanced modeling techniques. Methods: Using Orlando Health data between February of 2012 and April 2021, random sampling was used to split the dataset into training (10,838) and validation (4645). For the training data, resampling was employed to obtain a balanced quota, equal observations followed by an ACE30 and not followed by an ACE30. We included only variables used in Stein (drug category, age group, ED visits and hospitalizations in the prior year, cancer type, insurance category, number of anti-cancer agents, race, and BMI category). Logistic regression with L1-penalty, XGBoost (a non-linear nonparametric tree-based algorithm to account for nonlinear relationships), and artificial neural networks were used to develop and evaluate predictive models. Different sampling methods (bootstrap and SMOTE) as well as cuff-off thresholds for high-risk groups were tested. Results: Based on evaluation with validation data set, the best performing approach was an XGBoost model with SMOTE resampling in the training data. The four most important predictors were ED visits in the prior year, payor category (self-insured), cancer type (genitourinary) and hospitalization in past year. This model reported a PPV of 0.27. Many other combinations of methodologies described above were performed, and the PPV varied between 0.19 to 0.27. Of the 282 ICAs identified as high-risk by the best model, 76 (27.0%) had an ACE30. Of the 4346 ICAs identified as low-risk, 533 (12.3%) had an ACE30. The differences between high risk and low risk ACE30 rates were statistically significant (p &lt; 0.0001). In comparison, using Stein et al. models with our data reported a PPV of 0.27 and 0.25 which are basically at the same level of our best trained model. Conclusions: Using the variables included in A-PROACCT and a variety of machine learning models, advanced sampling methodologies and threshold cut-off limits, the best results were similar to those obtained using basic logistic regression. This suggests improvement in predicting acute care events following chemotherapy administration need to incorporate more extensive multimodal data, such as vital signs, performance status, patient reported outcomes, socioeconomic factors, laboratory and radiology results, remote patient monitoring and wearable device data.

Assessing boar taint in Portuguese pork: A small-scale study of prevalence and classification via established detection thresholds

Aim of study: To assess the boar taint prevalence in the Portuguese pork industry through an observational study model by measuring skatole and androstenone levels in fat and to compare it with the estimated thresholds for these compounds among Portuguese consumers. Area of study: Portugal. Material and methods: Adipose tissue samples from 140 animals (102 entire males and 38 females) were collected in three Portuguese abattoirs for boar taint quantification. Cut-off limits were determined using best-estimate thresholds (BET) for skatole and androstenone. Main results: Boar taint quantification for the 140 animals revealed levels of 36.0±4.7 ng/g of skatole and 64.5±21.3 ng/g of androstenone, and values were significantly higher in males. Group BET values were 35.4 ng/g and 566.3 ng/g for skatole and androstenone, respectively. BET values were exceeded in 28.8% of the samples for skatole and 0.7% for androstenone. Research highlights: The investigation revealed a generally low level of boar taint in the samples of this small-scale study on skatole and androstenone prevalence in Portugal's pork supply chain. However, occasionally a significantly elevated boar taint levels suggest that relying solely on slaughtering sexually immature males might not fully resolve this issue.

Nb2O5/SnO2 Heterojunctions Designed as Optical Absorbers and Microwave Band Filters

Herein, a 200 nm thick films of SnO2 are deposited onto Nb2O5 thin substrates (100 nm) to construct an isotype Nb2O5/SnO2 (NSO) heterojunction devices. The device is constructed using a thermal coating method in a vacuum media of low pressure reaching 10−5 mbar. NSO heterojunctions represent two amorphous/amorphous‐stacked layers that exhibit enhanced light absorption exceeding 170% and 120% in the visible and infrared regions of light spectrum, respectively. In addition, the interface under study displays band conduction offset of 0.60–0.90 eV and valence bad offset of 0.53–0.83 eV, respectively. The NSO device, which is treated as a planner microwaves resonators by contacting it with two Ohmic Pt electrodes, shows AC electrical conduction by the correlated barrier hopping within correlated barriers of average heights of 0.127 eV. Electrical conduction is actualized within scattering time constant of 0.27 ns. NSO microwaves resonators display gigahertz cutoff frequency spectra that suit 6G technology applications. The cutoff limit can be engineered in the range of 1.0–12.0 GHz by altering the driving frequency value. The features of the NSO heterojunctions nominate them for use in optoelectronic, which require high light absorption. The heterojunction devices are also promising for potential use in microwave technology a 6G bands filters.

Cardiac evaluation in free-ranging giant anteaters (Myrmecophaga tridactyla) in the Brazilian Cerrado

The giant anteater (Myrmecophaga tridactyla) is classified as a Vulnerable species, but these animals are regularly brought into rescue centers and are also kept in zoos around the world. They occupy a highly specialized dietary niche with a feeding ecology that is difficult to replicate under human care. This can lead to numerous nutrition-related health problems, including frequent cardiomyopathies. Establishing reference parameters for free-ranging animals is an important tool to assess the health of animals under human care. For this reason, a cardiac assessment through plasma troponin I concentrations, vertebral heart score (VHS) based on thoracic radiographs, and echocardiographic parameters were established for 11 free-ranging giant anteaters in the Cerrado savanna of Mato Grosso do Sul, Brazil. The troponin I concentrations were all lower than 0.2 ng/ml, the cutoff limit for detection in the assay used. The mean VHS in the right lateral projections was 7.49, a value almost identical to cats (7.5) and lower than dogs (9.7). A complete echocardiographic assessment is provided and compared with previous case reports. This paper validates the diet-related origin hypothesis for common dilated cardiomyopathy in giant anteaters. This is the first published study with radiographic and echocardiographic parameters for free-ranging giant anteaters, providing important reference parameters for future giant anteater in situ and ex situ conservation initiatives. This work highlights the benefits of veterinary research comparing health parameters for animals living under and outside of human care.

Enhanced Performance of Fe/WO3 Terahertz Dielectric Lenses

AbstractHerein transparent iron nanosheets deposited by the ionic coating technique onto glass and WO3 dielectric lenses are studied and characterized. The thickness of Fe nanosheets is varied in the range of 70–350 nm. It is observed that the transmittance and reflectance of the Fe nanosheets are highly affected by the layer roughness. Coating of iron nanosheets onto WO3 dielectric lenses increases the light absorption of WO3 by more than 240 times and red‐shifts the energy bandgap. Remarkable enhancements in the dielectric constant and in the optical conductivity are achieved via Fe coatings. In addition, iron coated dielectric lenses show higher terahertz cutoff limits varying in the range of 1.0–30 THz. Iron nanosheets remarkably increase the free charge carrier density and plasmon frequency in the infrared range of light. Moreover, the temperature dependent electrical conductivity shows high temperature stability and an increased electrical conductivity by more than 7 orders of magnitude by coating WO3 with 70 nm thick Fe nanosheets. The stability of the electrical conductivity at low temperatures and the wide range of terahertz cutoff limits in addition to the well‐enhanced light absorbability makes the iron coated tungsten oxide dielectric lenses promising for multifunction optoelectronic applications.

Automatic ultrasensitive lateral flow immunoassay based on a color-enhanced signal amplification strategy

Lateral flow immunoassays (LFIAs) are an essential and widely used point-of-care test for medical diagnoses. However, commercial LFIAs still have low sensitivity and specificity. Therefore, we developed an automatic ultrasensitive dual-color enhanced LFIA (DCE-LFIA) by applying an enzyme-induced tyramide signal amplification method to a double-antibody sandwich LFIA for antigen detection. The DCE-LFIA first specifically captured horseradish peroxidase (HRP)-labeled colored microspheres at the Test line, and then deposited a large amount of tyramide-modified signals under the catalytic action of HRP to achieve the color superposition. A limit of detection (LOD) of 3.9 pg/mL and a naked-eye cut-off limit of 7.8 pg/mL were achieved for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleoprotein. Additionally, in the inactivated virus detections, LOD equivalent to chemiluminescence (0.018 TCID50/mL) was obtained, and it had excellent specificity under the interference of other respiratory viruses. High sensitivity has also been achieved for detection of influenza A, influenza B, cardiac troponin I, and human chorionic gonadotrophin using this DCE-LFIA, suggesting the assay is universally applicable. To ensure the convenience and stability in practical applications, we created an automatic device. It provides a new practical option for point-of-care test immunoassays, especially ultra trace detection and at-home testing.

High frequency of transient congenital hypothyroidism among infants referred for suspected congenital hypothyroidism from the Turkish National screening program: thyroxine dose may guide the prediction of transients

PurposeWe aimed to determine the frequency of transient congenital hypothyroidism (TCH) in 17 participating centers in Türkiye, evaluate the etiological distribution in permanent congenital hypothyroidism (PCH) cases, and investigate the role of laboratory and clinical findings in predicting TCH.MethodsThis retrospective observational multicenter study included patients from 17 pediatric endocrinology centers identified by “National Newborn Screening Program” (NNSP) who were born in 2015 and followed for 6 years. Demographic, clinical, and laboratory information of the cases were compiled through the database http://cedd.saglik-network.org (CEDD-NET).ResultsOf the 239 cases initially treated for CH, 128 (53.6%) were determined as transient in whom a trial of levothyroxine (LT4) withdrawal was performed at a median age of 36 (34–38) months. Among the patients with PCH (n = 111), thyroid dysgenesis was diagnosed in 39.6% (n = 44). The predictive factors for TCH were: LT4 dose at the withdrawal of treatment, and initial newborn blood screening (NBS)-TSH level. Based on the receiver operating characteristic (ROC) curve analysis to predict optimal cut-offs for TCH predictors, LT4 dose < 2.0 µg/kg/day at treatment discontinuation was predictive for TCH and was associated with 94.5% specificity and 55.7% sensitivity, with an area under the curve (AUC) of 0.802. The initial NBS-TSH level value < 45 µIU/mL was predictive for TCH with 93.1% specificity and 45.5% sensitivity, with an AUC of 0.641. In patients with eutopic thyroid gland only LT4 dose < 1.1 µg/kg/day at withdrawal time was predictive for TCH with 84.7% sensitivity and 40.4% specificity, with an AUC of 0.750.ConclusionAccording to our national follow-up data, the frequency of TCH was 53.6%. We determined the LT4 dose < 2.0 µg/kg/day at discontinuation of treatment and the initial NBS-TSH level < 45 µIU/mL as the best cut-off limits to predict TCH.

Screening for central precocious puberty by single basal Luteinizing Hormone levels

PurposeTo identify cut-off for basal LH levels and for pelvic ultrasound uterine and ovarian parameters indicating an Hypotalamic–Pituitary–Gonadal (HPG) axis activation as diagnostic of Central Precocious Puberty (CPP).Methods248 girls referred for suspected precocious/early puberty who had undergone a GnRH stimulation test were enrolled and divided into three groups: Premature Idiopathic Thelarche (PIT), CPP, and Early Puberty (EA). For every patient basal serum Luteinising Hormone (LH) and Follicle Stimulating Hormone (FSH), basal LH/FSH ratio and pelvic ultrasonographic parameters were also collected. Through the use of Receiver Operating Curves (ROCs) the sensitivity (Se) and specificity (Sp) of basal LH, FSH, LH/FSH ratio and ultrasonographic parameters were evaluated at each level and Area Under the Curve (AUC) was measured.ResultsBasal LH model ≥0.14 mIU/mL reached the highest predictability (90.6% and 78.2%, Se and Sp, respectively). Basal LH/FSH ratio ≥0.1 showed a sensitivity of 85.90% and a specificity of 78.14%, while basal FSH cut-off (≥2.36 mIU/mL) had the lowest predictability, with a less favourable sensitivity (71%) and specificity (70.5%). Cut-off point for uterine length as 35 mm, (83.5% and 42.9% of Se and Sp, respectively) was calculated. For ovarian volumes, ROC curves showed very low sensitivity and specificity.ConclusionA single basal LH measurement under the cut-off limit may be adequate to exclude an HPG axis activation as CPP.

Association of urinary calcium excretion with chronic kidney disease in patients with type 2 diabetes.

Herein, we investigated the correlation between urinary calcium excretion (UCaE) and chronic kidney disease (CKD) in patients with type 2 diabetes mellitus (T2DM). From August 2018 to January 2023, a total of 2031 T2DM patients providing 24-h urine samples were included in the final analyses. Patients were separated into four cohorts, based on the UCaE quartiles. We then analyzed renal functional indicators like estimated glomerular filtration rate (eGFR) and urinary albumin excretion (UAE) among the four groups. Lastly, we utilized multivariable logistic regression models to investigate the correlation between UCaE and CKD. After adjusting for confounding factors, we observed a decreasing trend in CKD prevalence (36.3%, 13.0%, 7.5%, and 6.6%, respectively, P < 0.001) across the UCaE quartiles. Albuminuria (55.5% vs. 40.0%, 36.5%, 37.4%) and macroalbuminuria prevalence (20.0% vs. 9.3%, 5.2%, 5.7%) in the lowest quartile were markedly elevated, compared to the remaining three quartiles (P < 0.001). Meanwhile, the eGFR level (P < 0.001) showed a clearly increasing trend across the UCaE quartiles, and patients with moderate-to-severe decreases in eGFR levels (with cutoff limits at 30-59, 15-30, and < 15mL/min/1.73m2) were mostly found in the lowest quartile (P < 0.001). Logistic regression analysis revealed that patients in the lowest quartile experienced an enhanced prevalence of CKD, relative to those in the highest quartile (odds ratio: 5.90, 95% confidence interval: 3.60-9.67, P < 0.001). Decreased UCaE was independently associated with the CKD prevalence in T2DM patients.

Analysis of Demodulation Methods of Tilted Fibre Bragg Gratings Based on the Local Shift of the Cladding Mode Group

Tilted fibre Bragg gratings are optical fibre structures used as sensors of various physical quantities. However, their most popular application is to measure the refractive index of liquids. In such applications, it is important to obtain high measurement accuracy and the ability to distinguish two slightly different values of the refractive index. For this purpose, not only an appropriate periodic structure is needed, but also a demodulation method. We propose averaging the shift of a group of cladding modes. We use the TFBG grating, of which not all cladding modes exceed the cut-off limit. Such modes are not subject to leakage but only to shifts under the influence of SRI changes. To determine the average shift of a group of modes, we analyse cross-correlation algorithms of intensity-transformed optical spectra. Next, the cross-correlation main lobe is analysed by the centroid method, the Fourier phase and the Hilbert transform. Furthermore, phase changes of the main Fourier frequency are used to estimate a shift of part of the optical spectrum. Additionally, we propose the correction of the determined shift using a shift of another group of modes of the same TFBG grating.

High-Harmonic Generation Spectroscopy of Gas-Phase Bromoform.

High-Harmonic Generation (HHG) spectra of randomly aligned bromoform (CHBr3) molecules have been experimentally measured and theoretically simulated at various laser pulse intensities. From the experiments, we obtained a significant number of harmonics that goes beyond the cutoff limit predicted by the three-step model (3SM) with ionization from HOMO. To interpret the experiment, we resorted to real-time time-dependent configuration interaction with single excitations. We found that electronic bound states provide an appreciable contribution to the harmonics. More in detail, we analyzed the electron dynamics by decomposing the HHG signal in terms of single molecular-orbital contributions, to explain the appearance of harmonics around 20-30 eV beyond the expected cutoff due to HOMO. HHG spectra can be therefore explained by considering the contribution at high energy of HOMO-6 and HOMO-9, thus indicating a complex multiple-orbital strong-field dynamics. However, even though the presence of the bromoform cation should be not enough to produce such a signal, we could not exclude a priori that the origin of harmonics in the H29-H45 to be due to the cation, which has more energetic ionization channels.

Determination of cortisol cut-off limits and steroid dynamics in the ACTH stimulation test: a comparative analysis using Roche Elecsys Cortisol II immunoassay and LC-MS/MS

PurposeMeasurement of cortisol concentrations is method dependent. The study aimed to establish assay-specific cut-off limits for cortisol after adrenocorticotropic hormone (ACTH) stimulation, comparing Roche Elecsys Cortisol II immunoassay to liquid chromatography-mass spectrometry (LC-MS/MS), and to assess the impact of patient characteristics, estrogen containing oral contraceptives as well as relation to other adrenocortical steroid hormone dynamics.MethodsOne hundred healthy participants underwent a 250 μg ACTH-test, with plasma samples analyzed using ElecsysCortI, ElecsysCortII, and LC-MS/MS. Cortisone, corticosterone, 17-OH-progesterone, dehydroepiandrosterone sulfate (DHEAS), androstenedione, and testosterone were additionally analyzed with LC-MS/MS. Cut-off limit for a normal cortisol response to the ACTH-test was defined as: 2.5th percentile–1.96 × SE.ResultsElecsysCort II measured cortisol concentrations 21% (95% CI: 19–22%) lower than ElecsysCort I. Cut-off limits for cortisol 30 and 60 min after ACTH were 426 and 485 nmol/L (ElecsysCort II) and 411 and 470 nmol/L (LC-MS/MS). Cut-offs were unaffected by gender, or body-composition. The ACTH-test resulted in significantly increased adrenocortical steroid hormones, except for decreased cortisone concentrations (both sexes), and decreased testosterone in men (1.9 nmol/L, 95% CI: 1.3–2.5). Testosterone was increased in women (0.07 nmol/L, 95% CI: 0.02–0.13).ConclusionElecsysCort II has high analytical performance and yields significantly lower cortisol concentrations than prior polyclonal immunoassays. This clinically relevant difference underscores the necessity for revised cut-off limits for improved diagnostic precision. Suggested 30-minute cortisol cutoff limits are 411 nmol/L (LC-MS/MS) and 426 nmol/L (ElecsysCort II). Adrenocortical steroids increased upon ACTH stimulation, except for cortisone in both sexes and testosterone in men, both of which decreased.

A multicomponent LC-MS/MS method for drugs of abuse testing using volumetric DBS and a clinical evaluation by comparison with urine

BackgroundDrug testing commonly use urine as a specimen and immunoassays for screening. The need for supervised urine collection has led to an interest in alternative specimens and a need for using mass spectrometry methods already for screening. In addition, mass spectrometry methods allow for broad multipanel screening which of great value because of the increased number of substances that needs to be covered has increased over time. One alternative specimen of interest for drugs of abuse testing is dried blood spots (DBS) and this work aimed at developing multipanel screening methods based on selected reaction monitoring liquid chromatography - mass spectrometry for both urine and dried finger blood as specimens. Materials and MethodsThe urine method comprised 37 analytes and utilised salted out liquid/liquid extraction in 96-well format, respectively, and the blood method comprised 35 analytes, a 10 µL volumetric DBS device and a two-step solvent extraction procedure. In both cases stable isotope labelled internal standards were used for almost all analytes. ResultsThe methods were validated according to forensic standard. The lowest reporting limits were generally set at 100 ng/mL for urine and 1 ng/mL for blood and the accuracy and imprecision were within limits of 15 and 20%. The methods were applied in a clinical study on patients receiving methadone maintenance treatment for opioid dependence. Methadone was detected in all urine and DBS samples, for urine sometimes below the commonly applied screening cutoff limit of 300 ng/mL. In 20 out of 99 cases no other drug was detected in any specimen. The most commonly other detected substances were pregabalin, amphetamine, alprazolam, zopiclone and THCCOOH. Findings in urine and DBS generally agreed well but more positives were detected in DBS. ConclusionMultipanel methods using liquid chromatography – mass spectrometry suitable for clinical drug screening were successfully developed for urine and blood collected by finger-pricking and stored as DBS.

Development of a colloidal gold immunochromatographic strip for rapid detection of ribavirin in pig urine

ABSTRACT A specific and sensitive monoclonal antibody (mAb) for ribavirin (RBV) was generated by immunizing BALB/c mice with RBV – bovine serum albumin. Using the mAb, a highly sensitive indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and a colloidal gold immunochromatographic (CGI) test strip were developed for detecting RBV in pig urine. Under optimum assay conditions, IC50 was 0.08 ng/mL in 0.05 M phosphate-buffered saline and the recovery rate in real-world samples ranged between 77.3% and 99%. The cut-off limit of the CGI test strip for RBV was 5 ng/mL in pig urine. The cross-reactivity rate of the test strip to the tested antiviral drugs including acyclovir and moroxydine hydrochloride and the metabolites 1,2,4-triazole-3-carboxamide and 1-β-D-ribofuranosyl-1,2,4-triazole-3-carboxylic acid was below 0.1%. The results indicate that ic-ELISA and the CGI test strip have high sensitivity and specificity and that these methods are suitable for the rapid detection of RBV in pig urine.