We propose a new concept to detect a target molecule on a solid surface directly by spraying biosensing molecules. First, a filter paper as a model of a solid surface was spotted with a target nucleic acid. Then, the biosensing molecule, a DNA-nano-tweezers-structure that exhibits a peroxidase activity recognizing the target, was sprayed with hemin as a cofactor, followed by a second spray containing luminol as a substrate for a light-glowing signal. Finally, an image was recorded using a smartphone. The images revealed that only the places where the corresponding targets were spotted glowed a blue light.
{"title":"Direct detection of a target nucleic acid on a surface by spraying DNA-nano-tweezers-based biosensing molecules","authors":"Hisakage Funabashi, Hiroya Hatano, Hirotaka Nakamura, Reiji Shigematsu, Akio Kuroda","doi":"10.1016/j.ab.2025.116003","DOIUrl":"10.1016/j.ab.2025.116003","url":null,"abstract":"<div><div>We propose a new concept to detect a target molecule on a solid surface directly by spraying biosensing molecules. First, a filter paper as a model of a solid surface was spotted with a target nucleic acid. Then, the biosensing molecule, a DNA-nano-tweezers-structure that exhibits a peroxidase activity recognizing the target, was sprayed with hemin as a cofactor, followed by a second spray containing luminol as a substrate for a light-glowing signal. Finally, an image was recorded using a smartphone. The images revealed that only the places where the corresponding targets were spotted glowed a blue light.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"709 ","pages":"Article 116003"},"PeriodicalIF":2.5,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145407907","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-01Epub Date: 2025-10-30DOI: 10.1016/j.ab.2025.116004
Rolf Wibom , David Alsina , Karin Naess , Martin Engvall , Christoph Freyer , Anna Wedell , Anna Wredenberg
We present an optimised luminometric method for measuring muscle mitochondrial ATP production rate (MAPR), adapted to a 96-well microplate format. The enhanced assay enables quantification of ATP production from 12 or more substrate combinations within 15 min, using only 10 μL of isolated mitochondria. The method demonstrates high accuracy and precision, with a validated measurement range of 0.3–70 nmol/min/L. To support clinical interpretation, a reference dataset was established from 92 individuals aged seven months to 79 years. All these individuals were referred for muscle biopsy but were subsequently deemed unlikely to have a mitochondrial disorder following comprehensive clinical evaluation. An overview of the current version of our assays for oxidative phosphorylation (OXPHOS) enzymes is also provided.
As proof of concept, we present three patients carrying pathogenic variants in mitochondrial DNA (ATP6 and MT-TL1) and the nuclear PDHA1 gene. All exhibited decreased MAPR with one or more substrates, along with additional clinical, biochemical, and morphological features consistent with mitochondrial disease.
Furthermore, we illustrate the age-dependent development of MAPR in muscle across the human lifespan, demonstrating a 60–80 % higher maximal capacity for oxidative ATP production in adults compared with young children. In contrast, MAPR supported by fatty acid-derived substrates remains unchanged over the same period.
In conclusion, the improved MAPR assay offers a robust and efficient tool for assessing mitochondrial function in both clinical diagnostics and research. Its high-throughput format and reliable performance make it particularly well-suited for the investigation of suspected mitochondrial disorders.
{"title":"Advancing a sensitive method for measuring mitochondrial ATP production in small muscle biopsy samples","authors":"Rolf Wibom , David Alsina , Karin Naess , Martin Engvall , Christoph Freyer , Anna Wedell , Anna Wredenberg","doi":"10.1016/j.ab.2025.116004","DOIUrl":"10.1016/j.ab.2025.116004","url":null,"abstract":"<div><div>We present an optimised luminometric method for measuring muscle mitochondrial ATP production rate (MAPR), adapted to a 96-well microplate format. The enhanced assay enables quantification of ATP production from 12 or more substrate combinations within 15 min, using only 10 μL of isolated mitochondria. The method demonstrates high accuracy and precision, with a validated measurement range of 0.3–70 nmol/min/L. To support clinical interpretation, a reference dataset was established from 92 individuals aged seven months to 79 years. All these individuals were referred for muscle biopsy but were subsequently deemed unlikely to have a mitochondrial disorder following comprehensive clinical evaluation. An overview of the current version of our assays for oxidative phosphorylation (OXPHOS) enzymes is also provided.</div><div>As proof of concept, we present three patients carrying pathogenic variants in mitochondrial DNA (<em>ATP6</em> and <em>MT-TL1</em>) and the nuclear <em>PDHA1</em> gene. All exhibited decreased MAPR with one or more substrates, along with additional clinical, biochemical, and morphological features consistent with mitochondrial disease.</div><div>Furthermore, we illustrate the age-dependent development of MAPR in muscle across the human lifespan, demonstrating a 60–80 % higher maximal capacity for oxidative ATP production in adults compared with young children. In contrast, MAPR supported by fatty acid-derived substrates remains unchanged over the same period.</div><div>In conclusion, the improved MAPR assay offers a robust and efficient tool for assessing mitochondrial function in both clinical diagnostics and research. Its high-throughput format and reliable performance make it particularly well-suited for the investigation of suspected mitochondrial disorders.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"709 ","pages":"Article 116004"},"PeriodicalIF":2.5,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145420885","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-01Epub Date: 2025-11-08DOI: 10.1016/j.ab.2025.116009
Anna Makovska , Erik Walinda , Ryo Aoyama , Minsoo Kim , Daichi Morimoto
NEDD8, a ubiquitin-like modifier essential for cullin-RING ligase regulation, is difficult to refold from Escherichia coli inclusion bodies due to aggregation during denaturant removal. Conventional refolding methods such as rapid dilution, one-step dialysis, and stepwise dialysis often produce poorly soluble proteins, limiting structural and functional analyses. To overcome this, we developed a simplified slow dialysis system using a single peristaltic pump to generate a gradual urea gradient over 96 h, enabling efficient refolding of NEDD8 under gentle conditions. Both wild-type and Q40E-mutant NEDD8, which mimics the bacteria-mediated deamidation observed during infection, refolded successfully into monomeric protein, as confirmed by size-exclusion chromatography. NMR spectra showed folded protein that matched reference assignments, demonstrating suitability for atomic level structural analysis. The method also enabled recovery of previously insoluble ISG15 mutants, highlighting its broad applicability for structural studies of diverse challenging proteins.
{"title":"A minimal slow dialysis method for refolding inclusion body proteins: Structural application to NEDD8 and its Q40E mutant","authors":"Anna Makovska , Erik Walinda , Ryo Aoyama , Minsoo Kim , Daichi Morimoto","doi":"10.1016/j.ab.2025.116009","DOIUrl":"10.1016/j.ab.2025.116009","url":null,"abstract":"<div><div>NEDD8, a ubiquitin-like modifier essential for cullin-RING ligase regulation, is difficult to refold from <em>Escherichia coli</em> inclusion bodies due to aggregation during denaturant removal. Conventional refolding methods such as rapid dilution, one-step dialysis, and stepwise dialysis often produce poorly soluble proteins, limiting structural and functional analyses. To overcome this, we developed a simplified slow dialysis system using a single peristaltic pump to generate a gradual urea gradient over 96 h, enabling efficient refolding of NEDD8 under gentle conditions. Both wild-type and Q40E-mutant NEDD8, which mimics the bacteria-mediated deamidation observed during infection, refolded successfully into monomeric protein, as confirmed by size-exclusion chromatography. NMR spectra showed folded protein that matched reference assignments, demonstrating suitability for atomic level structural analysis. The method also enabled recovery of previously insoluble ISG15 mutants, highlighting its broad applicability for structural studies of diverse challenging proteins.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"709 ","pages":"Article 116009"},"PeriodicalIF":2.5,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145487379","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-01Epub Date: 2025-11-03DOI: 10.1016/j.ab.2025.116005
Mingxian Lu , Taigang Liu
Acidophilic, alkaliphilic, and halophilic proteins function under extreme conditions and hold great industrial value. However, their experimental identification is time-consuming and costly. Here, we introduce AHAPC, a unified computational framework for multiclass classification of extremophilic proteins. First, we construct a new benchmark dataset, TriExtrem, by combining three rigorously curated datasets. Then, we extracted two types of protein features, i.e., embeddings from pretrained protein language models (PLMs) and hand-crafted embeddings which include evolutionary descriptors derived from position-specific scoring matrix (PSSM), and sequence features, followed by the feature fusion and selection. Finally, convolutional neural network (CNN), gated recurrent unit (GRU), and bidirectional long short-term memory (BiLSTM) were used for three binary classification tasks respectively, while a multi-branch BiLSTM was adopted for the multiclass setting. Comprehensive evaluation and visualized analysis demonstrate that AHAPC achieves strong performance and provides interpretable predictions, facilitating reliable discovery of extremophilic proteins.
{"title":"AHAPC: Multi-source feature fusion and ensemble learning for multiclass extremophilic protein prediction","authors":"Mingxian Lu , Taigang Liu","doi":"10.1016/j.ab.2025.116005","DOIUrl":"10.1016/j.ab.2025.116005","url":null,"abstract":"<div><div>Acidophilic, alkaliphilic, and halophilic proteins function under extreme conditions and hold great industrial value. However, their experimental identification is time-consuming and costly. Here, we introduce AHAPC, a unified computational framework for multiclass classification of extremophilic proteins. First, we construct a new benchmark dataset, TriExtrem, by combining three rigorously curated datasets. Then, we extracted two types of protein features, i.e., embeddings from pretrained protein language models (PLMs) and hand-crafted embeddings which include evolutionary descriptors derived from position-specific scoring matrix (PSSM), and sequence features, followed by the feature fusion and selection. Finally, convolutional neural network (CNN), gated recurrent unit (GRU), and bidirectional long short-term memory (BiLSTM) were used for three binary classification tasks respectively, while a multi-branch BiLSTM was adopted for the multiclass setting. Comprehensive evaluation and visualized analysis demonstrate that AHAPC achieves strong performance and provides interpretable predictions, facilitating reliable discovery of extremophilic proteins.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"709 ","pages":"Article 116005"},"PeriodicalIF":2.5,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145450530","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-01Epub Date: 2025-10-31DOI: 10.1016/j.ab.2025.116006
Guangxin Yuan , Meng Xia , Ya Li , Honglei Tian , Jianyu Zhu , Yanshuang Wang
Deer placenta and Placenta Hominis are valued traditional Chinese medicines often adulterated with cheaper pig or cow placenta. Current identification methods frequently lack the throughput, speed, or simplicity for effective field monitoring. This study aimed to develop a high-throughput, rapid method to simultaneously identify deer, human, pig, and cow placenta. The approach involved screening a rapid genomic DNA extraction protocol and designing specific primers for the mitochondrial cytochrome b gene of each species, with primers labeled with FAM and Biotin. Following the optimization of PCR amplification conditions, results were visually detected using immunocolloidal gold test strips. The method was rigorously evaluated for its specificity, reproducibility, sensitivity, and stability. The results showed that the extracted DNA was of satisfactory concentration, purity, and integrity. Under optimized conditions (59 °C annealing temperature, 20 cycles), authentic samples for all four placenta types produced two distinct red bands on the test strips, while adulterated and blank controls showed only one control band. Agarose gel electrophoresis confirmed specific amplification for each target with no cross-reactivity. The method demonstrated strong specificity, excellent reproducibility and stability, and a high sensitivity of 0.01 ng μL−1, surpassing conventional electrophoresis. In conclusion, this PCR-based test strip method enables the visual, simultaneous authentication of the four placenta types in a single test, making it highly suitable for on-site rapid monitoring and quality control.
{"title":"A high-throughout PCR test strip method for the rapid identification of four placentas","authors":"Guangxin Yuan , Meng Xia , Ya Li , Honglei Tian , Jianyu Zhu , Yanshuang Wang","doi":"10.1016/j.ab.2025.116006","DOIUrl":"10.1016/j.ab.2025.116006","url":null,"abstract":"<div><div>Deer placenta and Placenta Hominis are valued traditional Chinese medicines often adulterated with cheaper pig or cow placenta. Current identification methods frequently lack the throughput, speed, or simplicity for effective field monitoring. This study aimed to develop a high-throughput, rapid method to simultaneously identify deer, human, pig, and cow placenta. The approach involved screening a rapid genomic DNA extraction protocol and designing specific primers for the mitochondrial <em>cytochrome b</em> gene of each species, with primers labeled with FAM and Biotin. Following the optimization of PCR amplification conditions, results were visually detected using immunocolloidal gold test strips. The method was rigorously evaluated for its specificity, reproducibility, sensitivity, and stability. The results showed that the extracted DNA was of satisfactory concentration, purity, and integrity. Under optimized conditions (59 °C annealing temperature, 20 cycles), authentic samples for all four placenta types produced two distinct red bands on the test strips, while adulterated and blank controls showed only one control band. Agarose gel electrophoresis confirmed specific amplification for each target with no cross-reactivity. The method demonstrated strong specificity, excellent reproducibility and stability, and a high sensitivity of 0.01 ng μL<sup>−1</sup>, surpassing conventional electrophoresis. In conclusion, this PCR-based test strip method enables the visual, simultaneous authentication of the four placenta types in a single test, making it highly suitable for on-site rapid monitoring and quality control.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"709 ","pages":"Article 116006"},"PeriodicalIF":2.5,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145429918","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-01Epub Date: 2025-11-13DOI: 10.1016/j.ab.2025.116011
Rithwik Pradeep , Alexander Zhdanov , Evin Allen
Tumour necrosis factor-alpha (TNF-α) plays a central role in inflammation and autoimmune pathology, making it a key therapeutic target. This study systematically evaluated the performance of monoclonal anti-human TNF-α antibodies derived from hybridoma cells—originally developed during early TNF research—using two widely adopted in vitro potency assays: a cytotoxicity-based assay in L929 fibroblasts and an NF-κB reporter assay in HEK293 Blue cells. Antibody performance was assessed in terms of neutralisation efficiency, signal reproducibility, and assay sensitivity. While both assays yielded comparable IC50 values (1.98 for L929 and 1.34 for HEK293 Blue), they differed in dynamic range, sensitivity, and biological relevance. The HEK293 Blue assay provided rapid, robust, and high-throughput-compatible detection of early TNF-α signalling events, with minimal background and superior reproducibility. In contrast, the L929 assay offered physiologically relevant insights into the later consequences of TNF-α signalling, such as apoptosis and metabolic disruption. ATP quantification using the CellTiter-Glo® assay proved to be a practical and sensitive method for viability assessment, though alternative metabolic and membrane integrity assays may complement interpretation. Overall, our findings highlight the complementary strengths of these assay platforms and support a dual-assay approach for comprehensive evaluation of anti-TNF-α antibodies.
{"title":"Comparative analysis of common potency assays for assessing human TNF-alpha neutralising antibodies","authors":"Rithwik Pradeep , Alexander Zhdanov , Evin Allen","doi":"10.1016/j.ab.2025.116011","DOIUrl":"10.1016/j.ab.2025.116011","url":null,"abstract":"<div><div>Tumour necrosis factor-alpha (TNF-α) plays a central role in inflammation and autoimmune pathology, making it a key therapeutic target. This study systematically evaluated the performance of monoclonal anti-human TNF-α antibodies derived from hybridoma cells—originally developed during early TNF research—using two widely adopted in vitro potency assays: a cytotoxicity-based assay in L929 fibroblasts and an NF-κB reporter assay in HEK293 Blue cells. Antibody performance was assessed in terms of neutralisation efficiency, signal reproducibility, and assay sensitivity. While both assays yielded comparable IC<sub>50</sub> values (1.98 for L929 and 1.34 for HEK293 Blue), they differed in dynamic range, sensitivity, and biological relevance. The HEK293 Blue assay provided rapid, robust, and high-throughput-compatible detection of early TNF-α signalling events, with minimal background and superior reproducibility. In contrast, the L929 assay offered physiologically relevant insights into the later consequences of TNF-α signalling, such as apoptosis and metabolic disruption. ATP quantification using the CellTiter-Glo® assay proved to be a practical and sensitive method for viability assessment, though alternative metabolic and membrane integrity assays may complement interpretation. Overall, our findings highlight the complementary strengths of these assay platforms and support a dual-assay approach for comprehensive evaluation of anti-TNF-α antibodies.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"709 ","pages":"Article 116011"},"PeriodicalIF":2.5,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145530424","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-01Epub Date: 2025-11-04DOI: 10.1016/j.ab.2025.116007
Wenxia Zhang , Guimei Wang , Zhenyu Wang , Guoqi Zhou , Hanliang Jiang
Background
Tyrosine metabolism (TM) plays an important role in the progression of cancer, but its role in lung adenocarcinoma (LUAD) is still unclear. This study aims to construct TM-related prognostic features for LUAD.
Methods
Transcriptomes and clinical data of LUAD were collected from public databases. A TM-related risk score (TMRS) model was constructed using 42 TM-related genes (TMRGs). The prognostic value of the model was comprehensively analyzed through survival analysis, enrichment analysis, immune assessment, and drug sensitivity prediction. The expression of key genes was also verified in LUAD cell lines and patient PBMCs.
Results
A 14-gene prognostic model (TMRS) was constructed. TMRS was an independent prognostic factor for LUAD. The low TMRS group has a more active immune microenvironment and may be more sensitive to immunotherapy. Patients with high TMRS may be more sensitive to various chemotherapy drugs. The model genes were specifically expressed in different cell types, suggesting that they may be involved in metabolic reprogramming and tumor progression.
Conclusion
This study establishes a foundation for personalized risk assessment and treatment decisions in LUAD, highlighting the prognostic significance of TM.
{"title":"The role of tyrosine metabolism-associated genes in predicting lung adenocarcinoma prognosis and their clinical relevance","authors":"Wenxia Zhang , Guimei Wang , Zhenyu Wang , Guoqi Zhou , Hanliang Jiang","doi":"10.1016/j.ab.2025.116007","DOIUrl":"10.1016/j.ab.2025.116007","url":null,"abstract":"<div><h3>Background</h3><div>Tyrosine metabolism (TM) plays an important role in the progression of cancer, but its role in lung adenocarcinoma (LUAD) is still unclear. This study aims to construct TM-related prognostic features for LUAD.</div></div><div><h3>Methods</h3><div>Transcriptomes and clinical data of LUAD were collected from public databases. A TM-related risk score (TMRS) model was constructed using 42 TM-related genes (TMRGs). The prognostic value of the model was comprehensively analyzed through survival analysis, enrichment analysis, immune assessment, and drug sensitivity prediction. The expression of key genes was also verified in LUAD cell lines and patient PBMCs.</div></div><div><h3>Results</h3><div>A 14-gene prognostic model (TMRS) was constructed. TMRS was an independent prognostic factor for LUAD. The low TMRS group has a more active immune microenvironment and may be more sensitive to immunotherapy. Patients with high TMRS may be more sensitive to various chemotherapy drugs. The model genes were specifically expressed in different cell types, suggesting that they may be involved in metabolic reprogramming and tumor progression.</div></div><div><h3>Conclusion</h3><div>This study establishes a foundation for personalized risk assessment and treatment decisions in LUAD, highlighting the prognostic significance of TM.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"709 ","pages":"Article 116007"},"PeriodicalIF":2.5,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145457608","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-01Epub Date: 2025-11-19DOI: 10.1016/j.ab.2025.116013
Hadyn Duncan , Elisabeth Domingo-Contreras , Sotirios Athanasiou , Rosario Fernandez-Godino , Francisco Castillo , Ana Camacho
Traditional diagnostic methods like RT-PCR and proviral DNA PCR have limitations, underscoring the need for swift, sensitive alternatives such as lateral flow immunoassays (LFIA). Yet, efficiently identifying monoclonal antibody (mAb) pairs for robust ternary antigen-antibody interactions remains a universal bottleneck, as conventional ELISA screening is costly and may not accurately reflect native binding. Here, we combined ELISA with Spectral Shift Technology (SST) to minimise reagent use and comprehensively profile anti-FeLV p27 mAbs. This orthogonal approach enabled rapid selection of two low nanomolar mAbs with different epitope targets that form a ternary complex, which translates into a sensitive detection of the FeLV p27 in a LFIA prototype. Our screening methodology supports resource-efficient LFIA development for FeLV and extends to other significant animal and human pathogens like Monkeypox or Helicobacter pylori, improving diagnostics and quality control.
{"title":"Affinity-based selection of anti-Feline Leukaemia Virus p27 monoclonal antibodies for efficient lateral flow assay development","authors":"Hadyn Duncan , Elisabeth Domingo-Contreras , Sotirios Athanasiou , Rosario Fernandez-Godino , Francisco Castillo , Ana Camacho","doi":"10.1016/j.ab.2025.116013","DOIUrl":"10.1016/j.ab.2025.116013","url":null,"abstract":"<div><div>Traditional diagnostic methods like RT-PCR and proviral DNA PCR have limitations, underscoring the need for swift, sensitive alternatives such as lateral flow immunoassays (LFIA). Yet, efficiently identifying monoclonal antibody (mAb) pairs for robust ternary antigen-antibody interactions remains a universal bottleneck, as conventional ELISA screening is costly and may not accurately reflect native binding. Here, we combined ELISA with Spectral Shift Technology (SST) to minimise reagent use and comprehensively profile anti-FeLV p27 mAbs. This orthogonal approach enabled rapid selection of two low nanomolar mAbs with different epitope targets that form a ternary complex, which translates into a sensitive detection of the FeLV p27 in a LFIA prototype. Our screening methodology supports resource-efficient LFIA development for FeLV and extends to other significant animal and human pathogens like Monkeypox or <em>Helicobacter pylori</em>, improving diagnostics and quality control.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"709 ","pages":"Article 116013"},"PeriodicalIF":2.5,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145562491","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-01Epub Date: 2025-10-14DOI: 10.1016/j.ab.2025.115993
Shweta Mishra , Ashlesha J. Chauhan , Jayaveersinh Mahida
Biosensors have become essential analytical tools that integrate biological recognition elements with physical transducers to detect specific analytes with remarkable sensitivity. This review delves into the applications of biosensors in healthcare, environmental monitoring, and other fields. Notable advancements include nanotechnology-enhanced biosensors, wearable devices, and point-of-care systems, all of which are characterized by miniaturization and multiplexing capabilities. In healthcare, biosensors facilitate disease diagnosis and continuous monitoring, whereas in environmental applications, they aid in pollutant detection. In analytical technology, biosensors merge biological recognition with physical transduction, offering unparalleled sensitivity in biological and environmental systems. This review examines the characteristics and applications of biosensors in various fields. They support disease diagnosis and health surveillance in healthcare and enable real-time pollutant monitoring for environmental protection purposes. Recent advances have incorporated nanotechnology to enhance performance through miniaturization and label-free detection. Wearable biosensors have revolutionized point-of-care diagnostics by providing real-time health data outside clinical settings. The integration of artificial intelligence and microfluidics has led to the development of more adaptive biosensing platforms. Despite challenges such as biological component stability and nonspecific binding, innovations in materials science offer potential solutions. The convergence of expertise positions biosensors as vital instruments for improving health outcomes and ensuring environmental sustainability, thereby transforming diagnostics and ecological preservation.
{"title":"Exploring biosensors: Distinctive features and emerging applications","authors":"Shweta Mishra , Ashlesha J. Chauhan , Jayaveersinh Mahida","doi":"10.1016/j.ab.2025.115993","DOIUrl":"10.1016/j.ab.2025.115993","url":null,"abstract":"<div><div>Biosensors have become essential analytical tools that integrate biological recognition elements with physical transducers to detect specific analytes with remarkable sensitivity. This review delves into the applications of biosensors in healthcare, environmental monitoring, and other fields. Notable advancements include nanotechnology-enhanced biosensors, wearable devices, and point-of-care systems, all of which are characterized by miniaturization and multiplexing capabilities. In healthcare, biosensors facilitate disease diagnosis and continuous monitoring, whereas in environmental applications, they aid in pollutant detection. In analytical technology, biosensors merge biological recognition with physical transduction, offering unparalleled sensitivity in biological and environmental systems. This review examines the characteristics and applications of biosensors in various fields. They support disease diagnosis and health surveillance in healthcare and enable real-time pollutant monitoring for environmental protection purposes. Recent advances have incorporated nanotechnology to enhance performance through miniaturization and label-free detection. Wearable biosensors have revolutionized point-of-care diagnostics by providing real-time health data outside clinical settings. The integration of artificial intelligence and microfluidics has led to the development of more adaptive biosensing platforms. Despite challenges such as biological component stability and nonspecific binding, innovations in materials science offer potential solutions. The convergence of expertise positions biosensors as vital instruments for improving health outcomes and ensuring environmental sustainability, thereby transforming diagnostics and ecological preservation.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"709 ","pages":"Article 115993"},"PeriodicalIF":2.5,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145306919","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-01Epub Date: 2025-11-19DOI: 10.1016/j.ab.2025.116014
Djamali Muhoza , Emily P. Esquivel , Stacy R. Hunter , Patience S. Okoto , Thallapuranam K.S. Kumar , Paul D. Adams
The serine/threonine kinase PAK1 serves as a mediator of cytoskeletal reorganization and cancer-related signaling downstream of the small GTPases. Due to the challenges in purifying PAK1 complexes, a 46-residue peptide from PAK1, is widely used to study PAK1-Cdc42 signaling. Traditionally, this purification involved multi-step chromatography of recombinant GST-PBD46 complexes, yielding approximately 1 mg per 1.5 L culture. In this study, a 30 min heat treatment step after thrombin cleavage was used to precipitate GST while leaving pure PBD46 in solution. This step eliminated the need for further affinity and size-exclusion chromatography steps. This improved protocol produces proteins with a 6.5-fold higher yield, halves the purification processing time, and produces peptides with ≥95 % purity. Mass spectrometry, CD, fluorescence, and 1H–15N HSQC NMR confirmed the heat-purified PBD46's identity, structure, folding, and binding to Cdc42. The method also successfully separated other small peptides (e.g., ACK1) but not larger folded proteins. This rapid and scalable approach facilitates peptide production and biochemical studies without compromising the structural or functional integrity. We believe that the method described herein is applicable to other stable GST-fused recombinant proteins.
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