Journal of Molecular Recognition最新文献

{"title":"AFM Force Volume for Extracellular Vesicle Detection and Membrane Blebbing Analysis Through Mechanical Signature Mapping.","authors":"F Collacchi, G Corti, M Girasole, G Longo, S Tacconi, D Vardanyan, A Magrini, P Lanuti, L Dini, M Bottini, S Dinarelli","doi":"10.1002/jmr.70038","DOIUrl":"10.1002/jmr.70038","url":null,"abstract":"<p><p>Membrane blebbing is a dynamic cellular phenomenon that plays a key role in numerous biological processes such as migration, apoptosis, cytokinesis, and extracellular vesicle (EV) formation. This phenomenon is traditionally observed via high-resolution imaging techniques on fixed samples, since it is difficult to quantify and monitor on live cells. This aspect limits our understanding of their temporal evolution and mechanical behavior. In this work, we present and validate a novel approach based on Atomic Force Microscopy (AFM) force spectroscopy combined with a custom data analysis pipeline that is capable to detect and map bleb- and EV-like structures directly on live cells. The method leverages the mechanical signatures embedded in force-distance (FD) curves-specifically, the presence of breakthrough points, which indicate the rupture or collapse of soft, vesicular structures. A custom MATLAB algorithm was developed to automatically identify and spatially map FD curves exhibiting EV-like mechanical behavior, based on signal smoothing, inflection point analysis, and user-defined force thresholds. This algorithm was at first validated against isolated EVs in liquid and then applied to live cultures of MDA-MB-231 breast cancer cells, monitored over 3, 7, 10, and 14 days to follow the temporal changes in membrane activity. This method enables the quantitative mapping of vesicle- and bleb-associated curves on the cell surface, highlighting their spatial distribution. A progressive increase in the number of EV-like events was observed up to Day 10, followed by a decline at Day 14-suggesting a dynamic remodeling of the membrane during cell culture maturation or stress. Moreover, they exhibit a preferential localization at the cell periphery. To validate the correspondence between EV-like force signatures and blebs, standard AFM high-resolution images were acquired on the same cells fixed and then analyzed using a Circular Hough Transform-based algorithm to detect and count blebs based on their circular geometry. The time trends obtained from both techniques showed good agreement, confirming the robustness of the force spectroscopy-based detection on live samples. This approach represents a significant advancement in the use of AFM for live-cell analysis. By extracting meaningful biophysical information from FD curves beyond standard elasticity metrics, we demonstrate that AFM spectroscopy can serve as a high-content, non-invasive tool for the detection of nanoscale membrane phenomena.</p>","PeriodicalId":16531,"journal":{"name":"Journal of Molecular Recognition","volume":"39 4","pages":"e70038"},"PeriodicalIF":2.7,"publicationDate":"2026-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147930566","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}

{"title":"Temperature Dependent Motions of N-Terminal Loop in PD-1 Determine the Affinity Towards Nivolumab.","authors":"Mohammad Karbalaeimahdi, Dircia C C Marcelina, Razvan C Stan","doi":"10.1002/jmr.70039","DOIUrl":"10.1002/jmr.70039","url":null,"abstract":"<p><p>Programmed Cell Death-1 receptors on the surface of monocytes and T and B cells modulate various immune responses and are key immunotherapy targets. Immunotherapeutic monoclonal antibodies, including Nivolumab, block the activity of this receptor and are a mainstay of cancer therapy. Use of these drugs has been associated with side effects, including local inflammation and systemic fever in some cancer patients. To understand the role of physiologic (37°C), pathophysiologic (39°C) and standard (25°C) temperatures on the formation of the immune complexes between PD-1 and Nivolumab, we used Surface Plasmon Resonance and Molecular Dynamics simulations, at appropriate temperatures. The mean lives obtained from the dissociation curves of Nivolumab:PD-1 complexes at 37°C or 25°C are ~16 times and ~125 times longer, respectively, than those at 39°C (~7 s). These discrepancies are due to N-terminal loop from PD-1 moving away from Nivolumab only at the higher temperatures, as well as due to changes in the epitope:paratope interface.</p>","PeriodicalId":16531,"journal":{"name":"Journal of Molecular Recognition","volume":"39 4","pages":"e70039"},"PeriodicalIF":2.7,"publicationDate":"2026-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148031385","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}

{"title":"","authors":"","doi":"","DOIUrl":"","url":null,"abstract":"","PeriodicalId":16531,"journal":{"name":"Journal of Molecular Recognition","volume":"39 4","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-05-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148096770","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}

{"title":"","authors":"","doi":"","DOIUrl":"","url":null,"abstract":"","PeriodicalId":16531,"journal":{"name":"Journal of Molecular Recognition","volume":"39 4","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-05-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148074754","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}

{"title":"","authors":"","doi":"","DOIUrl":"","url":null,"abstract":"","PeriodicalId":16531,"journal":{"name":"Journal of Molecular Recognition","volume":"39 4","pages":""},"PeriodicalIF":3.0,"publicationDate":"2026-05-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148062605","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}

{"title":"Artificial Intelligence Clustering Approach for Force Mapping Analysis of Polyacrylic Acid (PAA)/Polyethylene Oxide (PEO) Polymer Brushes for Biosensor Applications.","authors":"D Saad, F Dubreuil, A Steinberger, L Calmettes, E Dellandrea, M Phaner-Goutorbe","doi":"10.1002/jmr.70033","DOIUrl":"10.1002/jmr.70033","url":null,"abstract":"<p><p>Biocompatibility of a biosensor can be achieved by grafting polymer brushes onto a solid surface. These brushes must be able to attract specific analytes or repel unwanted entities. This is obtained with weak polyelectrolyte polymer brushes that shrink or swell depending on external stimuli. In this study, the conformation of polyacrylic acid (PAA) and polyethylene oxide (PEO) polymer brushes was characterized as a function of pH and ionic strength using Atomic Force Microscopy (AFM) in spectroscopic mode. Instead of colloidal tips classically used to measure the mechanical behavior of the brush, force curves were performed with conventional tips for better sensitivity to the interaction between ions and polymer, which is responsible for their conformation. Since force mapping experiments generate thousands of curves, a statistical representation was employed to define the general trend of the curves and facilitate their interpretation. As expected, the neutral PEO is not affected by changes in solution pH and salinity. In contrast, PAA exhibits behaviors depending on the ions present in the solution and increasing salinity; the brush shrinks at low pH with H₃O⁺ ions and swells with the addition of Na⁺ and K⁺ ions. The originality of the study also lies in the implementation of an Artificial Intelligence (AI) clustering model applied to force curves to specifically study a 50% PAA/50% PEO mixed polymer brush. This AI model makes it possible to distinguish areas of the surface where only one type of polymer has been grafted and to identify its nature according to its force curve.</p>","PeriodicalId":16531,"journal":{"name":"Journal of Molecular Recognition","volume":"39 3","pages":"e70033"},"PeriodicalIF":2.7,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13093003/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147723153","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Design of Multi-Epitope DIVA-Compatible Vaccine Candidate Against Canine Parvovirus 2.","authors":"Shubham Sharma, Pratishtha Sharma, Ravindra Pal Mandar, Ram Kumar, Ashok Gaur, Mamta Gurjar, Rajesh Singathia, Braj Nandan Shringi, Sunita Choudhary, Amit Kumar Pandey","doi":"10.1002/jmr.70036","DOIUrl":"10.1002/jmr.70036","url":null,"abstract":"<p><p>Canine parvovirus type 2 (CPV-2) is a highly contagious and often fatal canine pathogen, with antigenic variants (CPV-2a, 2b, 2c), maternally derived antibody (MDA) interference, and lack of Differentiating Infected from Vaccinated Animals (DIVA) capability limiting current vaccines. Using an immunoinformatics-based reverse vaccinology approach, we identified conserved B-cell, cytotoxic T lymphocyte (CTL), and helper T lymphocyte (HTL) epitopes from CPV-2 structural (VP1, VP2) and nonstructural (NS1, NS2) proteins. These were screened for antigenicity, immunogenicity, safety, and stability. The screening identified three B-cell, two CTL, and five HTL epitopes. Out of these, two representative epitopes were utilized for in vitro evaluation. The peripheral blood mononuclear cells (PBMCs) were collected from 10 dogs 28 days after vaccination with a live-attenuated whole-virus CPV vaccine and pooled. This pool was expected to contain memory cytotoxic T cells recognizing a broad viral epitope repertoire. On antigenic stimulation, PCPV2 induced significant proliferation of canine PBMCs, with the highest response at 33.33 μg/mL, alongside ~15.67-fold upregulation of IFN-γ mRNA, indicating a memory T-cell response. A multi-epitope vaccine was designed by integrating validated epitopes with the TLR-4 agonist RS09 and a Capripoxvirus-derived DIVA marker, linked by AAY/EAAAK spacers. The vaccine construct can be effectively delivered via a recombinant canine adenovirus-2 vector for efficient in vivo expression and cellular response. Structural modeling, molecular docking, immune simulations, molecular dynamic simulations, and codon optimization predicted strong receptor binding, stability, IgG/T-cell responses, and expression feasibility in Escherichia coli and canine cells. These findings support PCPV2 as a potent immunogen and demonstrate the promise of the designed construct for a potential adenovirus-vectored vaccine; however, in vivo validation will be essential to confirm efficacy, cross-variant protection, and DIVA performance.</p>","PeriodicalId":16531,"journal":{"name":"Journal of Molecular Recognition","volume":"39 3","pages":"e70036"},"PeriodicalIF":2.7,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147774199","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}

{"title":"RPA Combined With CRISPR/Cas12a for Rapid and Ultrasensitive Detection Dual-Gene of Methicillin-Resistant Staphylococcus aureus (MRSA).","authors":"Lei Chen, Jun Luo, Helin Zhang, Pingsen Zhao","doi":"10.1002/jmr.70035","DOIUrl":"10.1002/jmr.70035","url":null,"abstract":"<p><p>The increasing issue of infections caused by methicillin-resistant Staphylococcus aureus (MRSA) necessitates rapid and reliable diagnostic methods. While existing RPA-CRISPR/Cas12a platforms have demonstrated potential for MRSA detection, most rely on single-gene targets or require multiple Cas enzymes. Here, we have developed a novel dual gene detection strategy that simultaneously detects the S. aureus specific femA gene and the methicillin-resistant mecA gene in a single RPA-CRISPR/Cas12a reaction. This integrated approach enables clear discrimination between MRSA and methicillin-sensitive Staphylococcus aureus (MSSA) in just 30 min, with results visualized via both fluorescence and lateral flow strips. The assay exhibited high specificity (no cross-reactivity with common pathogens) and a sensitivity of 10 copies/μL, comparable to qPCR. Validation with 39 clinical samples showed 100% concordance with antimicrobial susceptibility testing. Our dual-gene RPA-CRISPR/Cas12a platform represents a significant advancement in point-of-care MRSA diagnostics, offering enhanced accuracy and operational simplicity.</p>","PeriodicalId":16531,"journal":{"name":"Journal of Molecular Recognition","volume":"39 3","pages":"e70035"},"PeriodicalIF":2.7,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13092367/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147723121","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Induced Circular Dichroism From the Binding of Achiral Bivalent Ligands to Transthyretin.","authors":"Stephen Wood, Yiwei Guan, James Pitts, Raj Gill, Simon Kolstoe, Támas Jávorfi, Rohanah Hussain, Giuliano Siligardi","doi":"10.1002/jmr.70037","DOIUrl":"10.1002/jmr.70037","url":null,"abstract":"<p><p>Achiral bivalent ligands capable of simultaneous occupation of both hormone binding sites of transthyretin generate a substantial and complex induced near ultraviolet circular dichroism spectrum during protein binding, revealing the dynamics of this process. Reduced temperature and pH slow the interaction and reveal two phases consistent with formation of an encounter complex and progression of the interaction through the core of the transthyretin tetramer. The x-ray structure of the protein-ligand complex confirms the endpoint of the binding trajectory and shows evidence of plasticity in the structure, with substantial disturbances of some mainchain and sidechains within and adjacent to the binding channel. This study highlights the effective complementarity of CD and x-ray investigations.</p>","PeriodicalId":16531,"journal":{"name":"Journal of Molecular Recognition","volume":"39 3","pages":"e70037"},"PeriodicalIF":2.7,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13109678/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147773977","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Meniscus-Mediated Imaging of Soft Biological Membranes Using 200 kN/m Ultra-Stiff 15 MHz MEMS AFM Probes.","authors":"Suyambulingam Subramanian, Nicolas Mauran, Ignacio Casuso, Bernard Legrand","doi":"10.1002/jmr.70034","DOIUrl":"10.1002/jmr.70034","url":null,"abstract":"<p><p>Coronaviruses, among other membrane-enveloped viruses, can remain infectious on surfaces in ambient air for days. Many bacteria and fungi also expose their plasma membranes to air environments. Key survival strategies in air include hydrophobic coatings to prevent water loss and waxy or silica-based resistant cell walls. Yet, many aspects remain largely unexplored due to the lack of suitable nanoscale imaging. While Atomic Force Microscopy (AFM) imaging in the liquid native environment of the biological samples gives access to many details of the membranes, operation in air damages these soft samples, being unable to resolve these fragile structures. Here, we demonstrate that ultra-stiff microelectromechanical (MEMS)-based AFM probes, with an ultra-high stiffness of 200 kN m^-1 and a resonance frequency of 15 MHz, can image soft cell membranes of Halobacterium Sallinarium in ambient air without structural disruption. Although meniscus-mediated tip stabilization has been previously observed for MEMS and optomechanical probes, this study provides the first demonstration of its application to biological imaging. By operating in a capillary-coupled regime with sub-ångström oscillation amplitude, the probe stabilizes at approximately 40% of the meniscus rupture distance, dissipating only ~35% of the maximum interaction energy. This enables noncontact imaging at tip-sample separations of ~5 nm and faithful recovery of the membrane topography (~5 nm thickness). The results establish a pathway for ultra-stiff AFM probe to perform noninvasive imaging of soft biological structures in air.</p>","PeriodicalId":16531,"journal":{"name":"Journal of Molecular Recognition","volume":"39 3","pages":"e70034"},"PeriodicalIF":2.7,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13078902/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147690707","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}