Search Results (1,894)

Prostate cancer is among the most prevalent malignancies in men worldwide, underscoring the need for early and accurate diagnostic tools. This study presents a proof-of-concept and pilot clinical validation of a novel bioelectric impedance-based biosensor for the detection of prostate-specific antigen (PSA) in human serum. The system integrates Molecular Identification through Membrane Engineering (MIME) with the xCELLigence real-time cell analysis platform, employing Vero cells electroinserted with anti-PSA antibodies. Optimization experiments identified 15,000 cells/well as the optimal configuration for impedance response. The biosensor exhibited specific, concentration-dependent changes in impedance upon exposure to PSA standard solutions and demonstrated significant differentiation between PSA-positive and PSA-negative human serum samples relative to the clinical threshold of 4 ng/mL. The biosensor offered rapid results within one minute, unlike standard immunoradiometric assay (IRMA), while showing strong diagnostic agreement. The system’s specificity, sensitivity, and reproducibility support its potential for integration into point-of-care screening workflows. This bioelectric assay represents one of the fastest PSA detection approaches reported to date and offers a promising solution for reducing overdiagnosis while improving clinical decision-making and patient outcomes. Full article

Background/Objectives: Anti-p200 pemphigoid is a rare and likely underdiagnosed autoimmune blistering disorder. Laminin γ1 and laminin β4 have been implicated as potential target antigens in its pathogenesis. Recently, a novel indirect immunofluorescence assay targeting anti-laminin β4 antibodies has been developed, demonstrating high sensitivity and specificity, and offering a valuable tool for improved diagnosis. Methods: Of the 451 patients, 21 were selected for further laboratory analysis based on medical records. Sera from 10 patients, which showed a positive direct immunofluorescence (DIF) result and negative results in multiplex enzyme-linked immunosorbent assays (ELISAs) and/or mosaic six-parameter indirect immunofluorescence (IIF) for various autoimmune bullous diseases, were tested for the presence of anti-laminin β4 antibodies. Additionally, sera from 11 patients with positive DIF and positive ELISA for antibodies against BP180 and/or BP230 were analyzed. Results: Among the 10 patients with positive DIF and negative ELISA and/or mosaic six-parameter IIF, 6 sera were positive for anti-laminin β4 antibodies. These patients presented with atypical clinical features. In contrast, all 11 sera from patients with both positive DIF and positive ELISA for BP180 and/or BP230 were negative for anti-laminin β4 antibodies. Conclusions: In patients with a positive DIF result but negative ELISA and/or mosaic six-parameter IIF findings, testing for anti-laminin β4 antibodies should be considered. Furthermore, in cases presenting with atypical clinical features—such as acral distribution of lesions, intense pruritus, or erythematous–edematous plaques—the possibility of anti-p200 pemphigoid should be included in the differential diagnosis. Full article

Since carbohydrate antigen 19-9 (CA 19-9) is a significant biomarker for the clinical diagnosis and treatment of pancreatic cancer, a sensitive sandwich-type immunosensor was proposed with an epoxy functionalized covalent organic framework (EP-COF_TTA-DHTA) as the antibody carrier and an electroactive COF_{TTA-2,6-NA(OH)2} as the signal amplification probe for the sensitive detection of CA 19-9. The flexible covalent linkage between the epoxy-functionalized EP-COF_TTA-DHTA and the antibodies was employed to improve the dynamics of the antigen–antibody interaction significantly. Meanwhile, AuNPs@COF_{TTA-2,6-NA(OH)2} with abundant electroactive sites enhanced the current response of the immunoreaction significantly. After optimizing the incubation time and concentration of the antibody, CA 19-9 was quantitatively detected by differential pulse voltammetry (DPV) based on the sensitive sandwich-type immunosensor with a low detection limit of 0.0003 U/mL and a wide linear range of 0.0009–100 U/mL. The electrochemical immunosensor exhibits high specificity, stability and repeatability, and it provides a feasible and efficient method for the pathologic analysis and treatment of tumor markers. Full article

Glioblastoma (GBM) is a highly aggressive and heterogeneous brain tumor. Glioma stem-like cells (GSCs) play a central role in tumor progression, therapeutic resistance, and recurrence. Although immune cells are known to shape the GBM microenvironment, the impact of antigen-presenting-cell (APC) signature genes on tumor-intrinsic phenotypes remains underexplored. We analyzed both bulk- and single-cell RNA sequencing datasets of GBM to investigate the association between APC gene expression and tumor-cell states, including stemness and metabolic reprogramming. Signature scores were computed using curated gene sets related to APC activity, KEGG metabolic pathways, and cancer hallmark pathways. Protein–protein interaction (PPI) networks were constructed to examine the links between immune regulators and metabolic programs. The high expression of APC-related genes, such as HLA-DRA, CD74, CD80, CD86, and CIITA, was associated with lower stemness signatures and enhanced inflammatory signaling. These APC-high states (mean difference = –0.43, adjusted p < 0.001) also showed a shift in metabolic activity, with decreased oxidative phosphorylation and increased lipid and steroid metabolism. This pattern suggests coordinated changes in immune activity and metabolic status. Furthermore, TNF-α and other inflammatory markers were more highly expressed in the less stem-like tumor cells, indicating a possible role of inflammation in promoting differentiation. Our findings revealed that elevated APC gene signatures are associated with more differentiated and metabolically specialized GBM cell states. These transcriptional features may also reflect greater immunogenicity and inflammation sensitivity. The APC metabolic signature may serve as a useful biomarker to identify GBM subpopulations with reduced stemness and increased immune engagement, offering potential therapeutic implications. Full article

Background/Objectives: Chemically or genetically detoxified pertussis toxin (PTx) is a crucial antigen component of the acellular pertussis vaccine. Chemical detoxification using glutaraldehyde generally causes significant structural changes to the toxin. However, how these structural changes in PTx affect its antigenic properties remains unclear. Additionally, there is limited knowledge regarding how many alterations in antigenic properties impact immunogenicity. Methods: To investigate the impact of structural changes on antigenic properties, we developed a sandwich ELISA to quantify the neutralizing epitopes on PTx. Subsequently, we analyzed different PTx toxoid (PTd) preparations with the assay. Additionally, we assessed the immunogenicity of various acellular pertussis vaccine candidates containing these PTd preparations. Finally, the assay was applied to evaluate the consistency of commercial batches of PTx and PTd intermediates. Results: The assay demonstrated reasonable specificity, accuracy, and precision, and it was sensitive enough to quantify variations in neutralizing epitopes among different PTd samples that shared the same protein concentration. Importantly, we found a positive correlation between the number of neutralizing epitopes in detoxified PTx and its immunogenicity, indicating that the amount of neutralizing epitopes present determines the immunogenicity of glutaraldehyde-inactivated PTx. Moreover, commercial batches of PTx and PTd intermediates exhibited minor variations in neutralizing epitopes. Conclusions: These findings have significant implications for developing acellular pertussis vaccines as they highlight the importance of preserving the neutralizing epitopes of PTx during detoxification to ensure the vaccine’s effectiveness. This assay is also valuable for the quality control of PTd as it more accurately represents the actual antigenic changes of PTx. Full article

Following lung cancer, prostate cancer is the leading cause of cancer death in men. High-risk localized tumor burden or metastatic disease often progresses, refractory to initial treatment regimens. There is ongoing development of technology to appropriately identify high-risk patients, stage them correctly, and offer appropriate treatments to obtain the best clinical outcomes. Prostate cancer-specific membrane antigen (PSMA) is a transmembrane glutamate carboxypeptidase, which helps regulate folate absorption, and its overexpression is pathologically directly proportional and associated with prostate cancer. Increased PSMA expression is a known independent risk factor for poorer survival, and most metastatic lesions in CRPC are PSMA positive. Over the last decade, several PSMA-based PET radiopharmaceuticals have demonstrated superior sensitivities and specificities compared to traditional imaging methods. These outcomes have been demonstrated by several large clinical trials. As the data emerges, these diagnostics are being integrated into standard of care protocol to facilitate nuanced identification of malignant lesions. PSMA is also being targeted through several therapeutics, including radioligands and immunotherapies such as CAR-T, BiTEs, and ADCs. This review will discuss the landscape of PSMA-based theranostics in the context of prostate cancer. 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

Biotin (vitamin B7) is a common, naturally occurring water-soluble vitamin. It belongs to the broad group of B vitamins. It is a common ingredient in dietary supplements, cosmetics, medicines, and parapharmaceutical preparations administered orally or applied topically (to the skin, hair, nails). The problem of the relationship between vitamin B supplementation and sensitivity seems to be multi-threaded. There is little literature data that would confirm that oral vitamin B supplementation or local exposure to biotin is a significant sensitizing factor. Moreover, it seems that allergy to vitamin B7 is very rare. It is possible, however, that the relationship between biotin and hypersensitivity is not limited to its direct action, but results from its essential metabolic function. Vitamin B7, as a cofactor of five carboxylases, affects the main pathways of cellular metabolism. Both deficiency and excess of biotin can result in metabolic disorders, which can have a significant impact on the homeostasis of the entire organism, including the efficient functioning of the immune system. Dysregulation of immune systems leads to its dysfunctional functioning, which can also lead to sensitization to various environmental antigens (allergens). Biotin is also used as an element of some methodological models in immunochemical tests (in vitro diagnostics), including methods used to measure the concentration of immunoglobulin E (IgE), both total (tIgE) and allergen-specific (sIgE). For this reason, vitamin B7 supplementation can be a significant interfering factor in some immunochemical tests, which can lead to false laboratory test results, both false positive and false negative, depending on the test format. This situation can have a direct impact on the quality and effectiveness of diagnostics in various clinical situations, including allergy diagnostics. This review focuses on the role of biotin in allergic reactions, both as a causative factor (allergen/hapten), a factor predisposing to the development of sensitization to various allergens, and an interfering factor in immunochemical methods used in laboratory diagnosis of hypersensitivity reactions and how it can be prevented. Full article

Swine infectious diseases, often caused by multiple co-infecting agents, pose severe global threats to pig health and industry economics. Conventional single-plex testing assays, whether relying on pathogen antigens or nucleic acids, exhibit limited efficacy in the face of co-infection events. The modern nucleic acid-based multiplex testing (NAMT) methods demonstrate substantial strengths in the simultaneous detection of multiple pathogens involving co-infections owing to their remarkable sensitivity, exceptional specificity, high-throughput, and short turnaround time. The development, commercialization, and application of NAMT assays in swine infectious disease surveillance would be advantageous for early detection and control of pathogens at the onset of an epidemic, prior to community transmission. Such approaches not only contribute to saving the lives of pigs but also aid pig farmers in mitigating or preventing substantial economic losses resulting from infectious disease outbreaks, thereby alleviating unwanted pressure on animal and human health systems. The current literature review provides an overview of some modern NAMT methods, such as multiplex quantitative real-time PCR, multiplex digital PCR, microarrays, microfluidics, next-generation sequencing, and their applications in the diagnosis of swine infectious diseases. Furthermore, the strengths and weaknesses of these methods were discussed, as well as their future development and application trends in swine disease diagnosis. Full article

The rapid evolution of SARS-CoV-2 has underscored the need for a detailed understanding of antibody binding mechanisms to combat immune evasion by emerging variants. In this study, we investigated the interactions between Class I neutralizing antibodies—BD55-1205, BD-604, OMI-42, P5S-1H1, and P5S-2B10—and the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein using multiscale modeling, which combined molecular simulations with the ensemble-based mutational scanning of the binding interfaces and binding free energy computations. A central theme emerging from this work is that the unique binding strength and resilience to immune escape of the BD55-1205 antibody are determined by leveraging a broad epitope footprint and distributed hotspot architecture, additionally supported by backbone-mediated specific interactions, which are less sensitive to amino acid substitutions and together enable exceptional tolerance to mutational escape. In contrast, BD-604 and OMI-42 exhibit localized binding modes with strong dependence on side-chain interactions, rendering them particularly vulnerable to escape mutations at K417N, L455M, F456L and A475V. Similarly, P5S-1H1 and P5S-2B10 display intermediate behavior—effective in some contexts but increasingly susceptible to antigenic drift due to narrower epitope coverage and concentrated hotspots. Our computational predictions show strong agreement with experimental deep mutational scanning data, validating the accuracy of the models and reinforcing the value of binding hotspot mapping in predicting antibody vulnerability. This work highlights that neutralization breadth and durability are not solely dictated by epitope location, but also by how binding energy is distributed across the interface. The results provide atomistic insight into mechanisms driving resilience to immune escape for broadly neutralizing antibodies targeting the ACE2 binding interface—which stems from cumulative effects of structural diversity in binding contacts, redundancy in interaction patterns and reduced vulnerability to mutation-prone positions. Full article

Immunoassays relying on highly specific antigen–antibody recognition are important tools for effectively measuring the levels of various targets. Efforts have been made in the development of various methods to improve the detection sensitivity and stability of immunoassays. Covalent organic frameworks (COFs), as an emerging class of novel crystalline porous materials, have unique advantages such as flexible designability, high surface area, excellent stability, tunable pore sizes, and multiple functionalities. They have great potential as novel sensory materials. Herein, we summarize the advances of COFs in electrochemical and optical immunoassays serving as electrode modifiers, signal indicators, enzyme or probe carriers, etc. Meanwhile, the design and application of typical COFs-based immunoassays in the determination of different targets are discussed in detail. Finally, challenges and future perspectives are presented. Full article

Synthetic biology has fundamentally advanced cell engineering and helped to develop effective therapeutics such as chimeric antigen receptor (CAR)-T cells. For these applications, the detection, localization, and quantification of heterologous fusion proteins assembled from interchangeable building blocks is of high importance. The V5 tag, a 14-residue epitope tag, offers promising characteristics for these applications but has only rarely been used in this context. Thus, we have systematically evaluated the murine anti-V5 tag antibody mu_SV5-Pk1 as well as its humanized version, hu_SV5-Pk1, to analyze cells expressing V5-tagged receptors in samples from various in vitro and in vivo experiments. We found that the V5 tag signal on cells is affected by certain fixation and detachment reagents. Immunohistochemistry (IHC) on formalin-fixed paraffin-embedded (FFPE) mouse tissue samples was performed to sensitively detect cells in tissue. We improved IHC by applying the hu_SV5-Pk1 monoclonal antibody (mAb) to avoid cross-reactivity within and unspecific background signals arising on fixed mouse tissue. Conversely, the absence of unspecific binding by the mu_SV5-Pk1 mAb was evaluated on 46 human normal or cancer tissues. Our findings present a robust toolbox for utilizing the V5 tag and cognate antibodies in synthetic biology applications. Full article

Background: Measuring frequencies of antigen-specific memory B cells (B_mem), their immunoglobulin (Ig) class and subclass usage, cross-reactivity, and affinity can provide insights into the efficacy of future antibody responses in case of antigen re-encounter. B cell ImmunoSpot^® assays can provide such information; however, like most cell-based tests, they require considerable amounts of blood to be drawn from the donor and this has hindered their inclusion in clinical trials and routine immune diagnostics. Methods: We introduce strategies for reducing the cell numbers required to 2–3 million peripheral blood mononuclear cells (PBMCs) per antigen, obtainable from 2–3 mL of blood from healthy adult donors. Results: Except when B_mem frequencies were very low, we found that testing PBMCs in singlet wells, but in serial dilution, enables as reliable B_mem frequency assessments as when testing replicate wells at a single fixed cell number. Additionally, B cell ImmunoSpot^® assays can be multiplexed for detecting four Ig classes, or IgG subclasses, simultaneously and without loss of sensitivity. The requirement for low cell numbers and the retention of B cell functionality by cryopreserved PBMCs equivalent to freshly isolated material implies that fewer than the standard 10 million PBMCs per vial can be frozen. This would reduce the number of individuals who could not be tested for B_mem due to insufficient availability of PBMCs, a common problem with such assays. Conclusions: The predictable need for and recovery of cryopreserved PBMCs facilitates planning of and optimal cell utilization in B cell ImmunoSpot^® assays and increases the practical feasibility of extensive B_mem characterization in larger cohorts. Full article

Circular RNA (CircRNA) is a single-stranded RNA arising from back splicing. CircRNAs interact with mRNA, miRNA, and proteins. These interactions regulate various life processes, including transcription, translation, cancer progression, anticancer drug resistance, and metabolism. Due to a lack of cap and poly(A) tails, circRNAs show exceptional stability and resistance to RNase degradation. CircRNAs exhibit dysregulated expression patterns in various cancers and influence cancer progression. Stability and regulatory roles in cancer progression make circRNAs reliable biomarkers and targets for the development of anticancer therapeutics. The dysregulated expression of circRNAs is associated with resistance to anticancer drugs. Enhanced glycolysis by circRNAs leads to resistance to anticancer drugs. CircRNAs have been known to regulate the response to chemotherapy drugs and immune checkpoint inhibitors. Exogenous circRNAs can encode antigens that can induce both innate and adaptive immunity. CircRNA vaccines on lipid nanoparticles have been shown to enhance the sensitivity of cancer patients to immune checkpoint inhibitors. In this review, we summarize the roles and mechanisms of circRNAs in anticancer drug resistance and glycolysis. This review discusses clinical applications of circRNA vaccines to overcome anticancer drug resistance and enhance the efficacy of immune checkpoint inhibitors. The advantages and disadvantages of circRNA vaccines are also discussed. Overall, this review stresses the potential value of circRNAs as new therapeutic targets and diagnostic/prognostic biomarkers for cancer Full article

Background: The pro-tumorigenic role of a disintegrin and metalloprotease 15 (ADAM15) is supported by its modified expression in primary tumors and ability to promote tumor growth in colorectal cancer (CRC). Cancer cell-derived ADAM15 promotes the progression of this malignancy by modulating the tumor microenvironment. However, according to our knowledge, this study is the first to assess serum ADAM15 concentrations in CRC patients in comparison to classical tumor markers—carcinoembryonic antigen (CEA) and cancer antigen 19-9 (CA19-9)—and a marker of the inflammatory process, C-reactive protein (CRP). The aim was to evaluate whether circulating serum ADAM15 might be a candidate biomarker for CRC diagnosis and progression. Methods: The study included 110 CRC patients and 54 healthy volunteers. Serum concentrations of ADAM15, CEA, and CA19-9 were measured using immunoenzyme assays, while CRP levels were assessed by the turbidimetric method. Diagnostic characteristics of all tested proteins were calculated. Results: Serum ADAM15 and classical tumor marker (CEA and CA19) levels were higher in CRC patients than in healthy subjects. However, a significant difference was observed only for CEA (p < 0.001). ADAM15 concentrations were significantly higher in CRC patients with distant metastases compared to those without metastases (p = 0.043). The highest diagnostic sensitivity (89%) was achieved by combined analysis of ADAM15 and CRP levels. Conclusions: These findings suggest a significant role of ADAM15 in CRC pathogenesis, indicating the usefulness of this protein in the prediction of distant metastases. Measurement of serum ADAM15, especially in combination with classical tumor markers (CEA) and inflammation markers (CRP), may improve the diagnosis of patients with CRC. Full article