Search Results (2,299)

Background: The mucosal barrier presents a significant challenge for non-invasive delivery of macromolecular therapeutics, often requiring administration with poor bioavailability and increased toxicity risks. The polymeric immunoglobulin receptor (pIgR) contains an extracellular secretory component (SC) for immunoglobulin binding and a membrane-anchored stem domain capable of apical-to-basolateral transcytosis. We hypothesized that targeting the stem domain could enable active drug transport across mucosal barriers. Methods: Using phage display, we identified four high-affinity nanobodies against human and murine pIgR. Two lead candidates (3LTHMP-4 and 3LTHMP-5) demonstrated efficient apical-to-basolateral transport in vitro (Transwell assays) and in vivo (fluorescence imaging). Engineered bispecific antibodies fusing these nanobodies with anti-IL-5 mAb reslizumab were administered via inhalation in a murine asthma model at one-tenth the intraperitoneal reslizumab dose. Resluts: The bispecific antibodies showed significant therapeutic efficacy, while reslizumab alone at equivalent concentrations failed to demonstrate efficacy. Hydrogen–Deuterium Exchange Mass Spectrometry (HDX-MS) revealed that both 3LTHMP-4 and 3LTHMP-5 specifically bind to the pIgR stem domain (residues 578–612), a region distinct from the dimeric IgA binding site. Conclusions: These findings suggest that stem domain-specific binding may facilitate transport across the mucosal barrier while preserving native receptor physiology, offering a potential strategy for effective transmucosal delivery of biologics. Full article

Food contamination by Aflatoxin M1 and Atrazine remains a critical food-safety concern, requiring sensitive detection methods that can operate reliably in complex matrices. Here, we report an AI-assisted antibody-functionalized electrochemical sensing platform for the detection and classification of Aflatoxin M1 and Atrazine across corn, corn flour, and protein matrices. The sensor used analyte-specific antibodies immobilized on an electrochemical electrode surface, where target binding produced measurable changes in the interfacial electrochemical response. Sensor performance was evaluated using cyclic voltammetry, coulometry, and electrochemical impedance spectroscopy (EIS), with EIS providing strong frequency-dependent signatures for concentration-dependent analysis. Spike-and-recovery studies further demonstrated the applicability of the platform in food-matrix conditions. To improve interpretation of complex electrochemical signals, full-spectrum EIS features were integrated with machine learning models for concentration-level classification into low, mid, and high groups. The AI workflow achieved an overall classification accuracy of 93.33%, with 96.67% specificity, 93.44% PPV, 96.66% NPV, and 0.982 AUC for Atrazine, and 96.70% specificity, 93.38% PPV, 96.67% NPV, and 0.987 AUC for Aflatoxin M1. In addition, analyte classification between Aflatoxin M1 and Atrazine reached 97.4% accuracy and 0.994 ROC-AUC. Overall, this work demonstrates a matrix-aware electrochemical immunosensing strategy enhanced by AI-based signal interpretation for food contaminant detection. Full article

The endothelin system is critical in chronic kidney disease (CKD) pathogenesis. However, the relative contribution of circulating endothelin-related biomarkers versus genetic variability remains unclear, particularly in diabetic nephropathy (DN) and after kidney transplantation (KTx). This study evaluated plasma concentrations of endothelin-1 (ET-1), endothelin A receptor (ETAR), and anti-ETAR antibodies (ETAR-Ab) in healthy controls, diabetic nephropathy (DN) patients, and DN patients after kidney transplantation (post-KTx). The influence of polymorphisms rs5370 (EDN1) and rs5333 (EDNRA) on endothelin-related parameters was analyzed. Polymorphisms were genotyped via PCR-RFLP, and endothelial-related parameters were determined by ELISA. Significant endothelin system activation was observed in both DN and post-KTx patients. ET-1 and ETAR concentrations were markedly elevated compared to controls, with the highest ET-1 levels detected in the post-KTx group, whereas ETAR-Ab levels were reduced. A sex-specific association for rs5370 was observed in male patients with the TG genotype (a nearly 4.5-fold higher risk of renal replacement therapy than in female patients). In proteinuric DN patients, the TC genotype (rs5333) was associated with elevated ETAR and ETAR-Ab. Endothelin system dysregulation is a prominent and persistent feature of CKD, noted after kidney transplantation. The endothelin activation, observed in transplant patients particularly, highlights the potential clinical relevance of endothelin-related biomarkers and supports the rationale for therapeutic strategies targeting the endothelin pathway, including endothelin receptor antagonists. Full article

The clinical relevance of food-specific IgG antibodies in pediatric gastrointestinal disorders remains controversial. Although current international guidelines discourage their use as standalone diagnostic tools, their significance within a broader immune–gut inflammatory framework has not been sufficiently explored. This study aimed to investigate associations between food-specific IgG reactivity, inflammatory and permeability biomarkers, microbiological findings, and abdominal ultrasound abnormalities in children with chronic gastrointestinal symptoms. Methods: (1) Children presenting chronic gastrointestinal symptoms associated with food-specific IgG polysensitization, elevated inflammatory and permeability biomarkers, and abdominal ultrasound abnormalities (number (n) = 196); (2) a symptomatic gastrointestinal group without the complete multimodal profile (n = 146); and (3) a control group with normal abdominal ultrasound findings and biomarkers within reference ranges (n = 210). All participants underwent food-specific IgG testing using a 216-antigen ELISA panel, abdominal ultrasound examination, and assessment of intestinal inflammatory and permeability biomarkers. Food-specific IgG antibodies were not interpreted as diagnostic markers of food allergy or food intolerance. Comparative analyses, correlation analyses, multivariable logistic regression, and receiver operating characteristic (ROC) analyses were performed. Results: Food-specific IgG polysensitization was significantly more frequent among children presenting the multimodal inflammatory profile compared with symptomatic and control groups (all p < 0.001). Reactivity predominantly involved gluten-containing cereals, dairy proteins, and mixed gluten–dairy patterns. Elevated fecal calprotectin, zonulin, and fecal histamine concentrations were more frequently observed in this subgroup, together with a higher prevalence of ultrasound abnormalities, including bowel wall thickening and mesenteric lymphadenopathy. Correlation analyses demonstrated significant associations between cumulative IgG burden and bowel wall thickness (r = 0.48, p < 0.001), while fecal calprotectin showed the strongest association with ultrasound abnormalities (r = 0.62, p < 0.0001). Multivariable logistic regression identified elevated calprotectin, increased zonulin, IgG polysensitization, and mixed gluten–dairy reactivity as independent predictors of pathological ultrasound findings. The integrated multimodal model demonstrated higher classification performance than isolated biomarkers. Conclusions: Children presenting chronic gastrointestinal symptoms, food-specific IgG polysensitization, inflammatory biomarker abnormalities, and ultrasound changes represented a multimodal clinical subgroup within the study population. These findings support evaluating food-specific IgG reactivity within a broader immune–gut assessment framework rather than as a standalone diagnostic biomarker. The observed associations should be considered exploratory and hypothesis-generating, requiring prospective validation and mechanistic investigation. Full article

Background/Objectives: Minipuberty represents the second phase of physiological activation of the reproductive axis and may play a role in postnatal genital development. Its course has been shown to be affected by untreated or inadequately treated maternal hypothyroidism. The aim of the present study was to investigate minipuberty in the sons of women with euthyroid autoimmune thyroiditis during pregnancy. Methods: This prospective matched cohort study included three groups of apparently healthy infant boys. Two groups comprised the male offspring of levothyroxine-naive, euthyroid women with autoimmune thyroiditis: one group was unsupplemented, and the other received vitamin D and selenium supplementation. The control group consisted of boys born to healthy women. Salivary concentrations of testosterone, androstenedione, DHEA-S, estradiol, and progesterone, along with urinary FSH and LH levels, were assessed longitudinally over the first 12 months of life. These hormonal measurements were evaluated in relation to genital development, including testicular volume and penile length, which were recorded at each study visit. Results: Compared with the offspring of healthy mothers, sons of women with autoimmune thyroiditis who did not receive supplementation exhibited lower concentrations of LH and testosterone, without a distinct peak, while the duration of hormone detectability did not differ between the groups. These hormonal alterations were accompanied by reduced penile length, with no differences observed in testicular volume. This group also exhibited lower DHEA-S concentrations, whereas levels of other hormones were comparable. In contrast, in the group receiving vitamin D and selenium supplementation, the dynamics of hormonal changes and genital organ growth did not differ from those observed in the control group. LH concentrations were inversely correlated with thyroid peroxidase antibody titers, which were lower in the supplemented group. Conclusions: The findings indicate an altered course of minipuberty in the sons of women with euthyroid autoimmune thyroiditis during pregnancy and suggest a potential benefit of exogenous vitamin D and selenium supplementation in this population. Full article

4-Chlorophenoxyacetic acid (4-CPA), a synthetic auxin analog, is employed in agriculture both as a plant growth regulator and as a constituent of herbicide formulations. Consequently, the establishment of simple and rapid detection methods is essential for effective environmental monitoring. This study reports the first development of a homogeneous fluorescence polarization immunoassay (FPIA) for the determination of 4-CPA. The monoclonal antibody (M1), raised against 4-CPA, was evaluated as a recognition element. Furthermore, two fluorescently labeled 4-CPA tracers—with ethylenediamine fluorescein thiocarbamate and aminohexylaminocarbonylfluorescein—were synthesized and purified, and their structures were unequivocally confirmed by high-performance liquid chromatography coupled with high-resolution mass spectrometric detection (HPLC-HRMS). Optimal concentrations of monoclonal antibodies and tracers were established, yielding a limit of detection of 1.2 ng/mL. The assay demonstrated a broad dynamic range of 2.3–300 ng/mL and a rapid analysis time of 15 min. Validation via the standard addition method in authentic open water samples resulted in recovery rates of 98–112%. To address the cross-reactivity with the prevalent herbicide 2,4-dichlorophenoxyacetic acid (2,4-D), two novel strategies were devised and successfully implemented. The first approach involves the concurrent execution of two separate FPIAs—one for 2,4-D and one for 4-CPA—followed by the mathematical resolution of two analyte concentrations from the two measured binding values. The second strategy entails the preliminary selective removal of 2,4-D from sample matrices using affinity chromatography columns with immobilized anti-2,4-D antibodies prior to FPIA for 4-CPA. These proposed methodologies appear highly promising for overcoming the inherent limitations of traditional immunoassays when faced with significant cross-reactivity among structurally analogous compounds. Full article

During construction of the Baku–Tbilisi–Kars railroad between Kars City in Türkiye and Tbilisi in Georgia, blasting operations in an anthrax-endemic region disrupted a burial pit containing carcasses of cattle that had died of anthrax. Railroad workers expressed concerns that release of this material could result in them developing anthrax infection. We therefore undertook a seroprevalence study six months later to seek evidence of exposure to Bacillus anthracis spores. We used an optimised Enzyme-Linked Immunosorbent Assay (ELISA) to screen serum for antigen-specific IgG antibodies to the anthrax toxin subunits Protective Antigen (PA) and Lethal Factor (LF). Stepwise linear regressions and t-tests were performed to compare results from railroad workers (n = 64) with a group of long-term Kars City residents (urban dwellers, n = 16), who had no history of possible contact with anthrax antigens. Anti-PA IgG concentrations were higher (p = 0.038) in railroad workers than in urban dwellers, but anti-LF IgG concentrations did not differ (p = 0.932) between the two groups. The anti-PA response is known to be dominant, and the difference was small. The lack of LF response did not preclude an antibody response to B. anthracis. Following the blasting operations, no cases of anthrax infection occurred in either railroad workers or villagers living nearby, suggesting that the spore exposure (evidenced by higher antibody titres) was at levels insufficient to initiate clinical infection. The elevated PA-specific antibody responses in railroad workers compared with urban dwellers might be consistent with the former having had previous subclinical exposure to B. anthracis. In anthrax-endemic regions, therefore, construction activities that involve blasting or large-scale excavation may pose risks of occupational exposure to Bacillus anthracis spores. Full article

Introduction: The 3 Rs principle advocates developing alternative, biologically relevant models. Thus, 3D fish liver in vitro models have been increasingly used for ecotoxicological studies. We previously optimized spheroids from the rainbow trout non-tumoral liver cell line RTL-W1 and employed them to assess the effects of aquatic pollutants. Although they demonstrated potential for assessing ecotoxicological effects, further optimization is warranted to enhance their physiological relevance. Incorporating an extracellular matrix (ECM), such as collagen, has been shown to be a promising strategy to improve spheroids’ structural organization and functionality. Objective: This study aimed to optimize 3D culturing conditions of RTL-W1 spheroids by evaluating the effects of collagen supplementation on viability, morphology, and functional response. Methodology: Spheroids from the RTL-W1 cell line (60,000 cells per well) were cultured in 96-well ultra-low attachment (ULA) plates at 18 °C. After spheroids’ formation, rat tail collagen was supplemented at concentrations of 15 (C15), 30 (C30), and 60 (C60) µg/mL at culture days 7, 8, and 9. Spheroids were collected at two sampling days (10 and 14). Viability was assessed using alamarBlue and lactate dehydrogenase (LDH) assays, while morphology was assessed by optical microscopy. Collagen penetration was evaluated using Masson’s trichrome staining technique. Protein expression of cytochrome P450(CYP)1A was assessed by quantifying immunocytochemistry staining using an anti-CYP1A antibody. Results: On day 10, LDH leakage decreased in C15 and C60, compared with the control, whilst C15 spheroids showed lower absorbance levels in the alamarBlue assay. On day 14, LDH showed no significant differences; however, C30 and C60 had higher alamarBlue absorbance, indicating greater metabolic capacity. Spheroid morphology appeared intact in all conditions. Masson trichrome revealed collagen fibrils at the periphery of the spheroids, especially in C30 and C60, indicating that spheroids incorporated collagen. CYP1A immunostain was present in all conditions, localized in the spheroids’ border, and tended to be higher when supplementation occurred in earlier days. Conclusions: Our results suggest that RTL-W1 spheroids interacted with the collagen matrix and appeared to functionally improve. Data suggest that incorporating ECM may increase the complexity and physiological relevance of RTL-W1 spheroids, thereby better supporting mechanistic and ecotoxicological applications. Full article

Background/Objectives: Nipah virus (NiV) is a highly pathogenic zoonotic virus with fatality rates exceeding 70% and causes recurring outbreaks in South and Southeast Asia. Reliable serological assays are critical for outbreak surveillance, diagnosis, and evaluation of vaccine-induced immune responses. This study aimed to develop and qualify an indirect enzyme-linked immunosorbent assay (ELISA) based on recombinant NiV glycoprotein G for the detection of virus-specific IgG antibodies in human serum. Methods: An indirect ELISA was developed and optimized for antigen concentration, blocking conditions, and serum dilution. The assay performance was evaluated using convalescent human sera from Bangladesh, along with the World Health Organization (WHO) International Standard for anti-Nipah virus antibodies, maintained and distributed by the National Institute for Biological Standards and Control (NIBSC). Analytical validation was conducted in accordance with ICH Q2 (R2) guidelines, including assessments of sensitivity, specificity, Precision, Linearity, and detection limits. Results: The assay demonstrated 100% sensitivity and specificity relative to reference sera. Intra-assay coefficients of variation ranged from 0.36% to 5.73%, and inter-assay variation was 4.16%, indicating high precision. The ELISA showed excellent Linearity (R² > 0.995). The lower limit of detection was 0.51 IU/mL, and the lower limit of quantification was 0.98 IU/mL. Conclusions: The developed ELISA is a BSL-2-compatible, robust, and scalable platform suitable for serosurveillance and the assessment of vaccine-induced immunity in endemic regions. Calibration against an international standard supports its applicability for standardized antibody measurement. This assay provides a practical tool for NiV outbreak response and vaccine evaluation. Full article

People living with HIV (PLWH) represent a high-risk population for hepatitis A virus (HAV) infection, with exposure risk equal to or higher than that of the general population, particularly within adult risk networks. Anti-HAV immunoglobulin G (IgG) serves as a neutralizing antibody and is considered a key marker of protective immunity against HAV infection. However, the serologic profile of anti-HAV IgG and IgM among unvaccinated PLWH remains insufficiently characterized, especially in South China. A total of 1232 unvaccinated adults were enrolled in the study, including 800 PLWH and 432 HIV- negative controls, to evaluate the serological markers of HAV immunity. Serum anti-HAV IgG and immunoglobulin M (IgM) were measured using enzyme-linked immunosorbent assays, and demographic, immunological, and biochemical data were collected. We observed that PLWH had significantly lower anti-HAV IgG concentrations (0.27 ± 0.16 vs. 0.31 ± 0.14 ng/mL, p < 0.001) and a lower IgG seropositivity rate (22.5% vs. 35.6%, p < 0.001) compared with HIV-negative controls, whereas no differences were found in IgM levels or positivity between the two groups. Multivariable logistic regression identified HIV infection (OR = 0.599, 95% CI 0.413–0.869, p = 0.007) and age (OR = 1.019, 95% CI 1.007–1.031, p = 0.002) as independent factors associated with IgG seropositivity. Among PLWH, those who were IgG-positive tended to be older (p = 0.003) and had higher serum globulin levels (p < 0.001), whereas IgM positivity was linked to younger age (p < 0.001) and a higher CD4/CD8 ratio (p = 0.030). Age-stratified analyses further revealed that IgG seroprevalence increased with age, while IgM positivity showed a declining trend. These findings indicate a considerable immunity gap in a population at elevated risk of HAV infection and support the need for targeted serologic screening and vaccination strategies among PLWH. Full article

Background. Newborn ruminants are born agammaglobulinemic, and colostrum quality (IgG concentration) is often insufficient, especially in heifers, leading to high morbidity and mortality from diarrheal infections. Objective. To develop and evaluate colostrum enrichment with specific egg yolk immunoglobulins (IgY) for the prevention of diarrhea in newborn calves. Methods. Hens were hyperimmunized with an inactivated vaccine against rotavirus, coronavirus, and E. coli. Yolk melange was prepared. Total and specific IgY were measured by chromatography and ELISA. Trials were conducted in three farms with high diarrhea incidence: experimental calves (n = 25) received 100 mL of melange (5 yolks) with the first two colostrum feedings; controls (n = 25) received native colostrum. Results. One yolk contained up to 100 mg of polyclonal IgY, with 8% specific antibodies. Diarrhea occurred in 12% of experimental calves (mild, no drugs) vs. 76% in controls (24% required antibiotics/rehydration). Testing in two other farms (n = 42, n = 38) reduced incidence 5.2–6.8-fold compared to the previous period. Conclusions. Enriching colostrum with specific IgY from hyperimmunized hens is highly effective and affordable for preventing diarrheal infections in newborn calves, especially in herds with poor colostrum quality in heifers. Full article

Antibody-drug conjugates (ADCs) have transitioned from clinically marginal agents into a defining therapeutic class for solid tumor oncology. In DESTINY-Breast03, trastuzumab deruxtecan achieved a four-fold progression-free survival advantage over trastuzumab emtansine, attributable not to antibody engineering but to the linker-payload axis: a cleavable peptide linker and a topoisomerase I payload with bystander activity. Sacituzumab govitecan extends the same logic to Trop-2-positive disease via extracellular payload release, and the framework now spans breast, urothelial, gynecologic, lung, gastric, and colorectal cancers, with enfortumab vedotin plus pembrolizumab displacing platinum chemotherapy as first-line therapy for urothelial cancer in EV-302 (median overall survival 31.5 versus 16.1 months). This review synthesizes ADC biology along three analytical axes. The mechanistic axis links each linker-payload-DAR configuration to a specific tumor-biology barrier: vascular limitation, which delivers approximately 0.1% of the administered dose to tumor tissue; the binding-site barrier, which concentrates exposure at the perivascular margin; and antigen mosaicism, which defeats internalization-dependent killing. The translational axis examines resistance as a coordinated failure across antigen modulation, trafficking, efflux, apoptotic execution, and lysosomal processing. The clinical axis traces the platform’s migration toward earlier-line and curative-intent settings. We close by examining whether the ADC delivery architecture translates to precision immunosuppression in autoimmune disease, where the glucocorticoid receptor modulator ADC ABBV-154 met placebo-controlled efficacy endpoints in rheumatoid arthritis but was discontinued because its benefit-risk profile did not differentiate it from existing biologic therapies. Full article

Background/Objectives: Minipuberty is a transient activation of the hypothalamic–pituitary–gonadal axis in infancy that contributes to the postnatal development of sexual organs. Its course has been shown to be influenced by maternal hypothyroidism. This study aimed to evaluate the reproductive axis and genital development in infant girls born to women with euthyroid autoimmune thyroiditis. Methods: The study involved three groups of infants: two groups were daughters of euthyroid women with autoimmune thyroiditis, while the third group (control) consisted of daughters of women without thyroid disease during pregnancy. Half of the mothers with thyroiditis received additional vitamin D and selenium supplementation during pregnancy, whereas the other half did not. During the first 18 months of life, periodic assessments were conducted of gonadotropin concentrations in urine, as well as salivary levels of estradiol, progesterone, testosterone, androstenedione, and DHEA-S. Additionally, ovarian volume, uterine length, and breast diameter were measured in the infants. Results: Daughters of women with autoimmune thyroiditis who did not receive supplementation during pregnancy exhibited lower levels of LH, estradiol, and progesterone, as well as a more rapid decline in LH and estradiol to below detectable levels, compared with daughters of healthy women. These hormonal differences were accompanied by smaller uterine length and breast diameter in this group. No differences were observed between the offspring of non-supplemented women with thyroiditis and daughters of healthy women regarding the levels of other hormones or ovarian volume. The dynamics of all assessed hormone levels and organ measurements did not differ between daughters of euthyroid women with thyroiditis who received vitamin D and selenium supplementation and daughters of healthy women. LH and progesterone levels showed inverse correlations with anti-thyroid peroxidase antibody titers, whereas uterine and breast dimensions positively correlated with estradiol levels. Conclusions: These findings suggest that maternal euthyroid autoimmune thyroiditis can affect the progression of female minipuberty, while supplementation with vitamin D and selenium during pregnancy may mitigate this effect. Full article

Vitamin D₃ is an essential fat-soluble vitamin for the human body. Its metabolite, 25-hydroxyvitamin D₃ (25(OH)D₃), serves as the primary biomarker to assess vitamin D levels. The monitoring of 25(OH)D₃ concentration is crucial for human health assessment. While traditional detection methods offer high sensitivity and accuracy, they are operationally complex and costly. This review systematically summarizes the most recent progress in 25(OH)D₃ detection technologies. Special attention is given to the recognition modes of 25(OH)D₃ by antibodies, nucleic acids, and molecularly imprinted recognition elements. Subsequently, the design strategies of diverse types of biosensors, including fluorescent, colorimetric, and electrochemical biosensors, are analyzed. Moreover, the development of portable devices, smartphone software, and flexible wearable devices for detection applications is also examined. Biosensing detection platforms are compared from the perspectives of target recognition, signal conversion, signal output, and application scenarios. Additionally, the potential of biosensor detection platforms in clinical diagnosis, health management, and community health surveillance is further investigated. Finally, the future trends of intelligent, portable, accurate, and home-use 25(OH)D₃ detection systems are delineated. This review offers a comprehensive reference for researchers developing next-generation 25(OH)D₃ diagnostic sensors and provides insights for the early prevention and treatment of vitamin D deficiency-related diseases. Full article

Acute respiratory diseases caused by viral pathogens such as Influenza A continue to represent a major global health challenge, emphasizing the need for rapid, sensitive, and accessible diagnostic tools. In this work, a carbon screen-printed electrode (SPCE)-based electrochemical immunosensor for the detection of an Influenza A (H1) antigen is reported, incorporating a comparative electrochemical evaluation of four electrode materials. Fe₃O₄ nanoparticles, Fe₃O₄@C nanoparticles, graphene quantum dots (GQDs), and gold nanoparticles (AuNPs) were systematically assessed by cyclic voltammetry to evaluate their electrocatalytic performance. The highest electrochemical response was selected for biosensor construction. The immunosensor was fabricated by immobilizing antibodies on a modified SPCE and characterized using differential pulse voltammetry (DPV). A concentration-dependent response was observed for H1 antigen concentrations ranging from 0 to 300 ng/mL, with a minimum detectable concentration (MDC) of 1 ng/mL and limit of detection (LOD) of 176 ng/mL and 45 ng/mL for serum and nasopharyngeal swabs, respectively. The biosensor performance was specifically evaluated in complex biological fluids, demonstrating reproducible performance and moderate selectivity against non-target influenza subtypes. Overall, this study highlights the critical role of electrode material selection in determining electrochemical immunosensor performance and supports the potential of SPCE-based platforms for the screening of an Influenza A (H1) antigen in point-of-care-oriented applications. Full article