Search Results (361)

Background: B7-H3, an immunoregulatory protein of the B7 family, has been associated with both anti-cancer immunity and tumor promotion, with its expression commonly correlated with poor prognosis. Although it is frequently expressed across cancers, its heterogeneity may limit the effectiveness of B7-H3-targeted therapies. Consequently, a sensitive and non-invasive method is needed to assess B7-H3 expression for patient selection and stratification. Single-domain antibody fragments (sdAbs) offer a promising platform for developing such a diagnostic tool. Methods: To generate B7-H3 sdAbs, two Ilamas were immunized with the recombinant human B7-H3 protein. Positive clones were selected through Phage biopanning and characterized for thermal stability, binding specificity, and affinity to human and murine B7-H3 proteins. Selected sdAbs were radiolabeled with Technetium-99m (^99mTc) and evaluated for B7-H3 detection in two xenograft tumor models using micro-SPECT/CT imaging and dissection studies. Results: Sixteen purified sdAbs bound specifically to recombinant B7-H3 proteins and cells expressing native B7-H3 antigens, with nanomolar affinities. The four best-performing sdAbs bound promiscuously to tested mouse and human B7-H3 isoforms. Lead sdAb C51 labeled with ^99mTc displayed specific accumulation across two human B7-H3⁺ tumor models, achieving high contrast with a tumor-to-blood ratio of up to 10 ± 3.16, and a tumor uptake of up to 4.96 ± 1.4%IA/g at 1.5 h post injection. Conclusions: The lead sdAb enabled rapid, specific, and non-invasive imaging of human B7-H3⁺ tumors. Its isoform promiscuity supports broad applicability across cancers expressing different human B7-H3 isoforms. These results support further development for clinical translation to enable patient selection and improved B7-H3-targeted therapies. Full article

Brucellosis, a zoonotic infection caused by the intracellular pathogen Brucella, leads to chronic multi-organ damage. Currently, rapid, accurate, and sensitive diagnostic technologies are crucial for the prevention and control of brucellosis. This study describes the development of a chemiluminescent immunoassay (Bru-CLIA) for sheep and bovine brucellosis antibody detection, utilizing Brucella abortus strain A19 lipopolysaccharide-coated magnetic particles (LPS-MPs) as the serum antigen and acridinium ester-labeled recombinant streptococcal protein G (AE-SPG) for signal generation. After optimizing the assay’s parameters, the Bru-CLIA demonstrated a sensitivity of approximately 1 IU/mL and 2 IU/mL for detecting sheep and bovine brucellosis, respectively. No cross-reactivity was observed with sera from animals immunized with Escherichia coli O157:H7, Mycobacterium tuberculosis, Vibrio cholerae, Legionella, Salmonella, Foot and Mouth Disease virus types O and A, Bovine viral diarrhea virus, Sheep contagious pleuropneumonia, Goat pox virus, or Peste des Petits Ruminants virus, indicating strong specificity. The testing of 81 sheep serum samples and 96 bovine serum samples revealed that Bru-CLIA showed 87.65% and 93.75% concordance with the ID-VET commercial kits for sheep and bovine brucellosis detection, respectively. These results demonstrate that Bru-CLIA offers high specificity, sensitivity, repeatability, and reliability, making it a viable rapid diagnostic tool for the epidemiological surveillance of brucellosis. Full article

The ongoing COVID-19 pandemic has underscored the urgent need for rapid, sensitive, and versatile diagnostic tools. In this study, we developed a nanogold-based lateral flow immunoassay (LFIA) capable of detecting both SARS-CoV-2 nucleocapsid (N) protein antigens and anti-N IgG antibodies at the point of care. Following optimization of colloidal gold nanoparticle size, pH, and protein conjugation parameters, LFIA strips were assembled in two formats: a competitive assay for antigen detection and a sandwich assay for antibody detection. In the competitive format, gold nanoparticles (AuNPs)-conjugated N protein were used to detect varying concentrations of free N protein. The test line signal inversely correlated with antigen concentration, confirming the assay’s specificity and effectiveness. For antibody detection, the sandwich LFIA format employed immobilized anti-human IgG to capture anti-N antibodies in serum samples from COVID-19 patients. Strong test line signals were observed in samples collected ≥11 days post-symptom onset, indicating a time-dependent increase in IgG detectability. These results demonstrate that the AuNP-based LFIA platform provides a flexible, rapid, and low-cost diagnostic solution, suitable for both early antigen detection and serological monitoring during SARS-CoV-2 infection and recovery. Full article

Background: Veterinary antibiotics are widely used in food-producing animals, raising public health concerns due to drug residues and the risk of antimicrobial resistance. Rapid and reliable detection systems are critical to ensure food safety and regulatory compliance. Colloidal gold immunoassay (CGIA)-based antigen–antibody test cards are widely used in food safety for the rapid screening of veterinary antibiotic residues. However, manual interpretation of test cards remains inefficient and inconsistent. Methods: To address this, we propose a complete AI-based detection system for veterinary antibiotic residues. The system is built on the Rockchip RK3568 platform and integrates a five-megapixel OV5640 autofocus USB camera (60° field of view) with a COB LED strip (6000 K, rated 5 W/m). It enables high-throughput, automated interpretation of colloidal gold test cards and can generate structured detection reports for regulatory documentation and quality control. The core challenge lies in achieving accurate and fast inference on resource-constrained embedded devices, where traditional detection networks often struggle to balance model size and performance. To this end, we propose VetStar, a lightweight detection algorithm specifically optimized for this task. VetStar integrates StarBlock, a shallow feature extractor, and Depthwise Separable-Reparameterization Detection Head (DR-head), a compact, partially decoupled detection head that accelerates inference while preserving accuracy. Results: Despite its compact size, with only 0.04 M parameters and 0.3 GFLOPs, VetStar maintains strong performance after distillation with the Bridging Cross-task Protocol Inconsistency Knowledge Distillation (BCKD) method. For our custom Veterinary Drug Residue Rapid Test Card (VDR-RTC) dataset, it achieves an mAP50 of 97.4 and anmAP50-95of 89.5. When deployed on the RK3568 device, it delivers results in just 5.4 s—substantially faster than comparable models. Conclusions: These results highlight the system’s strong potential for high-throughput, cost-effective, and rapid veterinary antibiotic residue screening, supporting food safety surveillance efforts. Full article

Over a decade ago, WHO introduced the ASSURED (Affordable, Sensitive, Specific, User-friendly, Rapid and Robust, Equipment-free, and Deliverable to end-users) criteria to guide diagnostic assay development. Today, lateral flow assays (LFAs) best meet these standards, evolving from simple rapid tests to advanced diagnostics integrating AI and nanotechnology for precise, quantitative results. Notably, nanoparticle-enhanced LFAs have achieved limits of detection (LOD) as low as 0.01 pg/mL (a 100-fold improvement over conventional methods), while AI algorithms have reduced interpretation errors by 40% in low-contrast conditions. The COVID-19 pandemic underscored the societal impact of LFAs, with over 3 billion antigen tests deployed globally, demonstrating 98% specificity in real-world surveillance. Beyond infectious diseases, LFAs are revolutionizing cancer screening through liquid biopsy, achieving a 92% concordance rate with gold-standard assays, food safety and environmental monitoring. Despite these advancements, challenges remain in scalability, reproducibility, sustainable manufacturing, and how to enhance the sensitivities and lower the LOD. However, innovations in biodegradable materials, roll-to-roll printing, CRISPR-integrated multiplexing, and efficient functionalization methods like photochemical immobilization technique offer promising solutions, with projected further cost reductions and scalability. This review highlights the technological evolution, diverse applications, and future trajectories of LFAs, highlighting their critical role in democratizing diagnostics. Full article

Wild animals often serve as reservoirs for vector-borne zoonoses, which are on the rise worldwide but have not yet been sufficiently researched. Vector-borne zoonoses, such as those caused by Anaplasma phagocytophilum, Borrelia burgdorferi sensu lato, and Dirofilaria immitis, are a growing public health concern due to their increasing incidence and broad host range. The aim of this study was to determine the prevalence and risk factors for vector-borne bacterial (borreliosis, anaplasmosis, ehrlichiosis) and parasitic (dirofilariasis) pathogens and to detect some of these pathogens in the red fox (Vulpes vulpes) population in Croatia. A total of 179 blood samples from foxes from nine districts were analysed. The SNAP ^® 4Dx ^® Plus rapid test was used to detect circulating D. immitis antigen and antibodies against B. burgdorferi, A. phagocytophilum/Anaplasma platys, and Ehrlichia canis/Ehrlichia ewingii. Circulating D. immitis antigen was detected in 6.70% of the samples (95% CI: 3.20–10.19%), while antibodies against A. phagocytophilum/A. platys were found in 10.06% (95% CI: 5.8–14.25%). Only one sample was positive for B. burgdorferi, while no antibodies were detected for E. canis/E. ewingii. Spatial analysis revealed statistically significant differences in prevalence by geographical region (district) and age, while no significant correlations were found. In the standard PCR analysis, DNA of D. immitis was not detected in any of the eight positive and eight negative SNAP ^® 4Dx ^® Plus samples. D. repens, A. reconditum, or co-infections were also not detected by PCR. Of the nine samples that tested positive for A. phagocytophilum/A. platys antibodies, four were confirmed to be positive for A. phagocytophilum by nested and semi-nested PCR targeting the 16S rRNA and GroEL genes. Phylogenetic analysis revealed similarities with various European strains, including zoonotic strains. This study is the first molecular detection of A. phagocytophilum from blood samples of red foxes in Croatia. The results show that red foxes are not free from infections such as anaplasmosis and dirofilariasis, emphasising their possible role in the maintenance and transmission of these pathogens in certain regions of Croatia. These results underline the need for further research to better understand the epidemiological importance of red foxes in the spread of vector-borne diseases. Full article

Co-infection of Orientia tsutsugamushi and influenza A virus complicates diagnosis and treatment in endemic regions because of overlapping clinical features and potential synergistic inflammation. We describe a 68-year-old woman from Hainan, China, who presented with five days of high fever (39.2 °C), nonproductive cough, eschar formation, lymphadenopathy, cytopenias, elevated liver enzymes, and raised inflammatory markers. On the day of admission, influenza A was confirmed by rapid antigen test and Orientia tsutsugamushi IgM/IgG was detected via colloidal-gold immunochromatography, prompting concurrent oseltamivir and doxycycline therapy. Quantitative PCR on day 2 measured an Orientia tsutsugamushi load of 2.85 × 10⁴ copies/mL (Cq 28.86), and targeted next-generation sequencing on day 3 revealed a high H1N1pdm09 viral burden (>1 × 10⁶ copies/mL) with low-level human herpesvirus 1 co-detection. Nested PCR and Sanger sequencing assigned Orientia tsutsugamushi to the Karp_A lineage and influenza A to clade 6B.1A.5a.2a. The patient defervesced by hospital day 2, laboratory indices normalized by day 3, and radiographic abnormalities resolved by day 6. This first documented Orientia tsutsugamushi–influenza A co-infection in China highlights the value of integrating rapid serology, qPCR quantification, nested PCR genotyping, and tNGS for early, precise dual-pathogen identification. Systematic multi-pathogen screening during overlapping transmission seasons is recommended to guide timely combination therapy and enhance epidemiological surveillance. Full article

Background and Objectives: A correct diagnosis of beta-hemolytic group A streptococcus (GAS)-pharyngitis allows the prevention of complications and unnecessary use of antibiotics. The aim of this study was to assess the management of pediatric GAS-pharyngitis in Romanian general practitioners (GPs)’ practice. Material and Methods: a cross-sectional study was conducted using a questionnaire distributed to Romanian GPs. Results: In total, 56 GPs completed the questionnaire, mostly females (83.9%, n = 47) from an urban area (60.7%, n = 34). They treated 5–10 (35.7%) or more than 10 (32.1%) cases of GAS monthly and considered white exudate on tonsils (92.9%, n = 52) to be the most suggestive clinical sign. Of the GPs, 25% (n = 14) used the Centor Criteria, 10.7% (n = 6) performed a rapid antigen detection test, and 42.9% (n = 24) requested throat culture for diagnosis. The younger GPs used the Centor Criteria significantly more often (p = 0.027) than the older ones. Most GPs (69.6%, n = 39) preferred targeted antibiotic therapy. Amoxicillin-clavulanate was the most commonly used antibiotic (55.4%, n = 31). Most GPs preferred oral antibiotics (89%, n = 50) for 10 days (55.4%, n = 31). Conclusions: Antibiotic treatment was initiated mostly based on clinical symptoms and in a short-course therapy. GPs stated that they prefer targeted antibiotic therapy, but they did not use proper diagnostic tools. Full article

Extrapulmonary tuberculosis (EPTB) remains diagnostically challenging due to its paucibacillary nature and variable presentation. Xpert and culture are limited in EPTB diagnosis due to sampling challenges, low sensitivity, and long turnaround times. This study evaluated the performance of the MPT64 antigen detection test for diagnosing EPTB, particularly tuberculous lymphadenitis (TBLN) and tuberculous pleuritis (TBP), in a high-TB, low-HIV setting. Conducted at Gulab-Devi Hospital, Lahore, Pakistan, this study evaluated the MPT64 test’s performance against conventional diagnostic methods, including culture, histopathology, and the Xpert MTB/RIF assay. Lymph node biopsies were collected, and cell blocks were made from aspirated pleural fluid from patients clinically presumed to have EPTB. Of 338 patients, 318 (94%) were diagnosed with EPTB. For TBLN, MPT64 demonstrated higher sensitivity (84%) than Xpert (48%); for TBP, the sensitivity was 51% versus 7%, respectively. Among histopathology-confirmed TBLN cases, MPT64 outperformed both culture and Xpert (85% vs. 58% and 47%). Due to the low number of non-TB cases, specificity could not be reliably assessed. The MPT64 test shows promise as a rapid, sensitive diagnostic tool for EPTB, particularly TBLN, in routine settings. While sensitivity is notably superior to Xpert, further studies are needed to evaluate its specificity and broader diagnostic utility. Full article

Nipah virus (NiV) and Hendra virus (HeV), which both belong to the genus henipavirus, are zoonotic pathogens that cause severe systemic, neurological, and/or respiratory disease in humans and a variety of mammals. Therefore, monitoring viral prevalence in natural reservoirs and rapidly diagnosing cases of henipavirus infection are critical to limiting the spread of these viruses. Current laboratory methods for detecting NiV and HeV include virus isolation, reverse transcription quantitative real-time PCR (RT-qPCR), and antigen detection via an enzyme-linked immunosorbent assay (ELISA), all of which require highly trained personnel and specialized equipment. Here, we describe the development of a point-of-care customized immunochromatographic lateral flow (ILF) assay that uses recombinant human ephrin B2 as a capture ligand on the test line and a NiV-specific monoclonal antibody (mAb) on the conjugate pad to detect NiV and HeV. The ILF assay detects NiV and HeV with a diagnostic specificity of 94.4% and has no cross-reactivity with other viruses. This rapid test may be suitable for field testing and in countries with limited laboratory resources. Full article

Heavy metal ions pose a significant threat to the environment and human health due to their high toxicity and bioaccumulation. Traditional instrumentations, although sensitive, are often complex, costly, and unsuitable for on-site rapid detection of heavy metal ions. Lateral flow assay technology has emerged as a research hotspot due to its rapid, simple, and cost-effective advantages. This review summarizes the applications of lateral flow assay technology based on nucleic acid molecules and antigen–antibody interactions in heavy metal ion detection, focusing on recognition mechanisms such as DNA probes, nucleic acid enzymes, aptamers, and antigen–antibody binding, as well as signal amplification strategies on lateral flow testing strips. By incorporating these advanced technologies, the sensitivity and specificity of lateral flow assays have been significantly improved, enabling highly sensitive detection of various heavy metal ions, including Hg²⁺, Cd²⁺, Pb²⁺, and Cr³⁺. In the future, the development of lateral flow assay technology for detection of heavy metal ions will focus on multiplex detection, optimization of signal amplification strategies, integration with portable devices, and standardization and commercialization. With continuous technological advancements, lateral flow assay technology will play an increasingly important role in environmental monitoring, food safety, and public health. Full article

Breast cancer is a leading cause of cancer-related mortality worldwide, requiring efficient diagnostic tools for early detection and monitoring. Human epidermal growth factor receptor 2 (HER2) is a key biomarker for breast cancer classification, typically assessed using immunohistochemistry (IHC). However, IHC requires invasive biopsies and time-intensive laboratory procedures. In this study, we present a biosensor integrated with a reusable printed circuit board (PCB) and functionalized glucose test strips designed for rapid and non-invasive HER2 detection in saliva. The biosensor achieved a limit of detection of 10⁻¹⁵ g/mL, 4 to 5 orders of magnitude more sensitive than the enzyme-linked immunosorbent assay (ELISA), with a sensitivity of 95/dec and a response time of 1 s. In addition to HER2, the biosensor also detects cancer antigen 15-3 (CA15-3), another clinically relevant breast cancer biomarker. The CA15-3 test demonstrated an equally low limit of detection, 10⁻¹⁵ g/mL, and a higher sensitivity, 190/dec, further validated using human saliva samples. Clinical validation using 29 saliva samples confirmed our biosensor’s ability to distinguish between healthy, in situ breast cancer, and invasive breast cancer patients. The system, which integrates a Bluetooth Low-Energy (BLE) module, enables remote monitoring, reduces hospital visits, and enhances accessibility for point-of-care and mobile screening applications. This ultra-sensitive, rapid, and portable biosensor can serve as a promising alternative for breast cancer detection and monitoring, particularly in rural and underserved communities. Full article

Diagnosis of histoplasmosis is challenging. A rapid, sensitive, and specific method is essential. Serum is a non-invasive and easy sample to obtain in any hospital. The diagnostic accuracy of different techniques that use serum has been evaluated. Forty-one serum samples from patients with proven or probable histoplasmosis were analyzed. Different diagnostic techniques based on the detection of antibodies (ID Fungal Antibody System), antigens (Histoplasma GM EIA and Platelia^TM Aspergillus Ag), and DNA (“in-house” real-time PCR (RT-PCR) were tested and compared. Additionally, the quantification of cytokines and biomarkers related to histoplasmosis was performed. Global results from 27 samples in which all the tests were performed showed that the sensitivity of the Histoplasma GM EIA kit was 87.5% in patients with disseminated infection and HIV as an underlying disease; in immunocompetent (IC) patients, it was 54.5%. The detection of Histoplasma spp. with the ID Fungal Antibody System was positive in 90.9% of IC and in 62.5% of HIV patients. The Platelia-Asp kit had a low performance in both groups of patients (37.5% in HIV and 9% in non-HIV), and, finally, RT-PCR was better in immunosuppressed patients (44% in HIV vs. 27% in non-HIV). The combination of diagnostic techniques increased the detection of Histoplasma infection in inmunosupressed patients. Overall, patient groups infected with H. capsulatum (Hc) showed higher IL-8, IL-6, IL-1β, TNF-α, and IL-18 median values compared to non-Hc-infected controls. The effectiveness of diagnostic techniques on serum samples is highly influenced by the patient’s clinical presentation and underlying condition. Consequently, a thorough assessment of the patient’s clinical presentation and disease phenotype is crucial in selecting the most suitable diagnostic method. Full article

Norovirus, a foodborne pathogen, causes a significant economic and health burden globally. Although detection methods exist, they are expensive and non-field deployable. A flow-based dielectrophoretic biosensor was designed for the detection of foodborne pathogenic viruses and was tested using bacteriophage MS2 as a norovirus surrogate. The flow-based MS2 sensor comprises a concentrator and a detector. The concentrator is an interdigitated electrode array designed to impart dielectrophoretic effects to manipulate viral particles toward the detector in a fluidic channel. The detector is made of a silver electrode conjugated with anti-MS2 IgG to allow for antibody–antigen biorecognition events and is supplied with the electrical current for the purpose of measurement. Serially diluted MS2 suspensions were continuously injected into the fluidic channel at 0.1 mL/min. A cyclic voltammogram indicated that current measurements from single-walled carbon nanotube (SWCNT)-coated electrodes increased compared to uncoated electrodes. Additionally, a drop in the current measurements after antibody immobilization and MS2 capture was observed with the developed electrodes. Antibody immobilization at the biorecognition site provided greater current changes with the antibody-MS2 complexes vs. the assays without antibodies. The electric field applied to the fluidic channel at 10 V_pp and 1 MHz contributed to an increase in current changes in response to MS2 bound on the detector and was dependent on the MS2 concentrations in the sample. The developed biosensor was able to detect MS2 with a sensitivity of 10² PFU/mL within 15 min. Overall, this work demonstrates a proof of concept for a rapid and field-deployable strategy to detect foodborne pathogens. Full article

The COVID-19 pandemic has necessitated urgent measures to curb its spread, with vaccination emerging as a pivotal strategy. This prospective observational study evaluated breakthrough COVID-19 infections among healthcare workers (HCWs) vaccinated with Comirnaty (Pfizer-BioNTech) at a tertiary care hospital in Greece. Over an 8-month period, from February to September 2021, 1958 fully vaccinated HCWs were monitored. Rapid antigen tests and RT-PCR tests were conducted weekly for asymptomatic HCWs. Contact tracing and whole-genome sequencing were performed. Results showed that 2.75% (54 cases) of HCWs experienced breakthrough infections. Among these, 25 (45%) were asymptomatic, mild symptoms occurred in 21 (37%), while 7 (13%) had a fever (≥38 °C) alone and 3 (5%) developed high fever (≥39 °C) with respiratory symptoms. Physicians and nursing staff were the most affected groups. Dominant SARS-CoV-2 variants detected included Delta, British, and Wild type variants. Comparison with existing literature underscored the effectiveness of Comirnaty in reducing breakthrough infections. The findings emphasize the importance of continued booster vaccinations and ongoing surveillance to mitigate breakthrough infections among HCWs. Full article