Methods and Protocols最新文献_第7页

Protocol for the Implementation of a Targeted Maternal and Newborn Service Delivery Bundle in Sierra Leone. 在塞拉利昂实施有针对性的孕产妇和新生儿一揽子服务提供议定书。

IF 2.5

Methods and Protocols Pub Date : 2026-06-10 DOI: 10.3390/mps9030096

Robert B Clark, Joseph Odu, Annette Ofodum, Rondi Anderson

{"title":"Protocol for the Implementation of a Targeted Maternal and Newborn Service Delivery Bundle in Sierra Leone.","authors":"Robert B Clark, Joseph Odu, Annette Ofodum, Rondi Anderson","doi":"10.3390/mps9030096","DOIUrl":"10.3390/mps9030096","url":null,"abstract":"Sierra Leone faces persistently high neonatal and maternal mortality rates, driven largely by delayed recognition and treatment of newborn respiratory distress and postpartum hemorrhage. In this protocol, we describe the planned implementation of a bundle of maternal and newborn clinical practices over a 36-month period across nine public health facilities in the Greater Freetown area and Bo District to address these critical gaps. The service delivery improvements include the World Health Organization (WHO) Essential Newborn Care Course (ENCC) Parts 1 and 2; Vayu bubble continuous positive airway pressure (bCPAP) and oxygen blenders for respiratory support; the WHO Postpartum Hemorrhage package; and obstetric risk stratification using point-of-care ultrasound (POCUS) and complementary diagnostics for maternal care improvement. We anticipate that this bundle of evidence-based clinical tools and training, reinforced by mentorship, structured checklists, and low-dose high-frequency (LDHF) practice, will significantly reduce perinatal and maternal mortality and morbidity. The bundle will be evaluated using a Hybrid Type 1 effectiveness-implementation design, utilizing routine health information system data, supplemented by project registers, skills assessments, and observations. By aligning with the Ministry of Health's Child Survival Action Plan, the aim of this project protocol is to provide a sustainable and scalable model for reducing preventable maternal and newborn deaths in resource-constrained settings.","PeriodicalId":18715,"journal":{"name":"Methods and Protocols","volume":"9 3","pages":""},"PeriodicalIF":2.5,"publicationDate":"2026-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13305997/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148318234","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Methodological Framework for a Multimodal Rat Model of Bleomycin-Induced Fibrosis and Autologous Tissue Grafting. 博莱霉素诱导大鼠纤维化和自体组织移植的多模态模型的方法学框架。

IF 2.5

Methods and Protocols Pub Date : 2026-06-10 DOI: 10.3390/mps9030094

Razvan George Bogdan, Iulian-Alexandru Ciprian Blidisel, Ionut Ciobota, Anca Maria Campean, Alina Helgiu, Claudiu Helgiu, Ioan Catalin Bodea, Dan Ionel Orbulescu, Rodica Elena Heredea, Zorin Petrisor Crainiceanu

{"title":"Methodological Framework for a Multimodal Rat Model of Bleomycin-Induced Fibrosis and Autologous Tissue Grafting.","authors":"Razvan George Bogdan, Iulian-Alexandru Ciprian Blidisel, Ionut Ciobota, Anca Maria Campean, Alina Helgiu, Claudiu Helgiu, Ioan Catalin Bodea, Dan Ionel Orbulescu, Rodica Elena Heredea, Zorin Petrisor Crainiceanu","doi":"10.3390/mps9030094","DOIUrl":"10.3390/mps9030094","url":null,"abstract":"Reproducible experimental models of localized dermal-hypodermal fibrosis are essential for standardized investigation of regenerative interventions. Variability in bleomycin dosing, anatomical targeting, and assessment strategies limits cross-study comparability. This study describes a methodological framework for standardized induction of early dermal-hypodermal remodeling in a rat model followed by autologous subcutaneous tissue grafting and multimodal longitudinal evaluation. Female Wistar rats underwent subcutaneous bleomycin administration at 1 mg/kg/day for three consecutive days. Clinical documentation, high-frequency ultrasonography with fixed imaging parameters, and sequential biopsies from a predefined thoracic anatomical site were performed at baseline, intermediate reassessment, and final evaluation. Autologous subcutaneous tissue grafting was conducted at Day 17 after study initiation. The protocol enabled controlled induction of early structural remodeling and consistent longitudinal documentation of dermal-hypodermal thickness, echogenicity changes, and histological architecture within a standardized anatomical region. This protocol development study establishes a reproducible and spatially consistent experimental platform integrating imaging and histological assessment, facilitating future hypothesis-driven investigations of fibrotic remodeling and regenerative strategies.","PeriodicalId":18715,"journal":{"name":"Methods and Protocols","volume":"9 3","pages":""},"PeriodicalIF":2.5,"publicationDate":"2026-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13304985/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148318168","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Content Generalizability: Generalizing Qualitative Research to the General Study Population. 内容概括性：将定性研究推广到一般研究人群。

IF 2.5

Methods and Protocols Pub Date : 2026-06-04 DOI: 10.3390/mps9030090

Costas S Constantinou

{"title":"Content Generalizability: Generalizing Qualitative Research to the General Study Population.","authors":"Costas S Constantinou","doi":"10.3390/mps9030090","DOIUrl":"10.3390/mps9030090","url":null,"abstract":"This paper highlights an important topic in qualitative research: the generalizability of qualitative findings to the general study population. While the concept of generalizability in qualitative inquiry has been discussed, with types such as analytical generalizability and transferability being proposed, the direct generalization of qualitative results to the general population has received limited attention. This oversight stems largely from the prevailing belief among researchers that qualitative research cannot be generalized due to its reliance on small, non-randomized samples. This perception has led many scientists, particularly those outside the social sciences, to regard qualitative research primarily as exploratory or supplementary to quantitative findings. This paper is a technical note that presents a new concept and procedure. Based on concept analysis, content generalizability is proposed, which offers a new type of generalizability through which qualitative findings can be applied to the general population, thereby enabling them to inform policy and drive social change. Content generalizability could expand the uses of qualitative research within the scientific community by providing knowledge and insights into the experiences of the general study population. This paper also contributes to developing empirical research by testing and applying content generalizability.","PeriodicalId":18715,"journal":{"name":"Methods and Protocols","volume":"9 3","pages":""},"PeriodicalIF":2.5,"publicationDate":"2026-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13304781/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148318206","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A Lightweight Workflow for Targeted Long-Read Transcriptomic Profiling Using Oxford Nanopore Sequencing. 使用牛津纳米孔测序靶向长读转录组分析的轻量级工作流程。

IF 2.5

Methods and Protocols Pub Date : 2026-06-04 DOI: 10.3390/mps9030091

Mariya Levkova

{"title":"A Lightweight Workflow for Targeted Long-Read Transcriptomic Profiling Using Oxford Nanopore Sequencing.","authors":"Mariya Levkova","doi":"10.3390/mps9030091","DOIUrl":"10.3390/mps9030091","url":null,"abstract":"Long-read sequencing technologies provide portable and flexible service, making them attractive for small-scale sequencing studies. However, many existing RNA-sequencing analysis frameworks are designed for transcriptome-wide analyses and require substantial computational resources. Here we present a lightweight and reproducible computational pipeline for targeted long-read transcriptomic profiling using Oxford Nanopore Technologies (ONT) cDNA sequencing data. The pipeline was evaluated using targeted long-read transcriptomic datasets generated from formalin-fixed paraffin-embedded (FFPE) colorectal carcinoma samples previously classified as microsatellite instability-high (MSI-high) by PCR-based testing. Libraries were sequenced on the Oxford Nanopore MinION platform using R10.4.1 flow cells. Application of the workflow enabled rapid quantification of mismatch repair gene expression and detection of immune-related transcripts including CD8A, PDCD1, and HAVCR2 across multiplexed barcode samples. The pipeline performs targeted alignment of long-read sequencing data to a custom transcript reference panel using minimap2, followed by gene-level read counting and normalization using reads-per-million (RPM). Optional modules enable immune marker profiling, detection of reads aligning to multiple genes, exploratory variant analysis, and visualization of expression patterns. By combining simplicity, reproducibility, and minimal computational overhead, the present pipeline provides an accessible framework for targeted transcriptomic analysis of long-read sequencing data. It may facilitate adoption of ONT-based transcriptomic profiling in settings with restricted computational resources.","PeriodicalId":18715,"journal":{"name":"Methods and Protocols","volume":"9 3","pages":""},"PeriodicalIF":2.5,"publicationDate":"2026-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13304758/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148318151","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Comparative Study of Bacteriophage Pharmacokinetics by Different Enteral Administration Routes. 不同肠内给药途径噬菌体药代动力学的比较研究。

IF 2.5

Methods and Protocols Pub Date : 2026-06-03 DOI: 10.3390/mps9030089

Maria Anurova, Aleksey Kuzmin, Aleksey Vorobev, Nataliya Feldman, Elena Zinurova, Andrey Aleshkin

引用次数: 0

A Multiresolution Breast Cancer CIBERSORTx Resource Validated for Accuracy, Interpretive Limits, and Biological and Clinical Coherence in Tumor Microenvironment Deconvolution. 多分辨率乳腺癌CIBERSORTx资源在肿瘤微环境反褶积中的准确性，解释限制和生物学和临床一致性验证。

IF 2.5

Methods and Protocols Pub Date : 2026-06-02 DOI: 10.3390/mps9030088

Toru Hanamura, Akinori Takase, Masanori Oshi, Naoki Niikura

{"title":"A Multiresolution Breast Cancer CIBERSORTx Resource Validated for Accuracy, Interpretive Limits, and Biological and Clinical Coherence in Tumor Microenvironment Deconvolution.","authors":"Toru Hanamura, Akinori Takase, Masanori Oshi, Naoki Niikura","doi":"10.3390/mps9030088","DOIUrl":"10.3390/mps9030088","url":null,"abstract":"Accurate deconvolution of bulk transcriptomes is essential for characterizing the breast cancer tumor microenvironment (TME), yet existing reference matrices incompletely capture tumor-specific cellular diversity. Here, we developed breast cancer-specific multiresolution CIBERSORTx signature matrices from single-cell RNA sequencing data and systematically evaluated their analytical performance and interpretability. Major-, minor-, and subset-level matrices were constructed and assessed using pseudo-bulk mixtures and pure cell profiles, while biological and clinical coherence were evaluated in TCGA-BRCA and the I-SPY2 cohort. All matrices demonstrated high accuracy in reconstructing pseudo-bulk compositions, with performance declining at finer resolution. Spillover increased with granularity but was largely restricted within related lineages. Lineage-wise deconvolution modestly reduced spillover but consistently decreased accuracy, highlighting the importance of cross-lineage transcriptional contrast. In external datasets, most inferred cell populations showed biologically coherent associations with canonical markers and pathways, whereas some fine-resolution subsets exhibited non-canonical patterns, likely reflecting intra-lineage trade-offs or context-dependent transcriptional states. In the I-SPY2 cohort, plasmablasts and selected myeloid populations were positively associated with pathological complete response, whereas fibroblastic and perivascular-like populations showed negative associations. These findings establish a validated and interpretable resource for breast cancer TME deconvolution and clarify its performance characteristics and limitations.","PeriodicalId":18715,"journal":{"name":"Methods and Protocols","volume":"9 3","pages":""},"PeriodicalIF":2.5,"publicationDate":"2026-06-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13304752/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148318106","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

An Overview of In Vitro Release Methods for Long-Acting Injectable Products Based on PLGA. 基于PLGA的长效注射类药物体外释放方法综述

IF 2.5

Methods and Protocols Pub Date : 2026-06-01 DOI: 10.3390/mps9030087

Maja Lusina Kregar, Iva Krtalić, Ivana Šagud

{"title":"An Overview of In Vitro Release Methods for Long-Acting Injectable Products Based on PLGA.","authors":"Maja Lusina Kregar, Iva Krtalić, Ivana Šagud","doi":"10.3390/mps9030087","DOIUrl":"10.3390/mps9030087","url":null,"abstract":"Long-acting injectables (LAIs) are widely used for chronic conditions such as schizophrenia, opioid use disorder, and HIV. Their prolonged efficacy improves adherence and reduces dosing frequency. Among these systems, poly(lactide-co-glycolide) (PLGA)-based formulations are commonly used to deliver drugs ranging from small molecules to peptides and proteins. In vitro release (IVR) tests play a critical role in evaluating drug product performance for both immediate- and prolonged-release dosage forms. However, there is a lack of standardized compendial IVR methods for the assessment of LAIs. This lack impedes the development of new drug products in this area and also complicates their regulatory approval process. Considering the complexity of drug release mechanisms and the diversity of various formulation design approaches, it is not possible to devise a universal IVR method that is applicable to all LAI products. The in vitro release test applied for quality control should be simple, robust, reproducible, and discriminatory. On the other hand, more complex biorelevant media and methods are often used during development to better reflect physiological conditions. This article provides a comprehensive review of compendial and non-compendial methods used for in vitro release testing of PLGA-based LAIs (microspheres and in situ forming implants), with the goal of aiding the development and standardization of future methodologies.","PeriodicalId":18715,"journal":{"name":"Methods and Protocols","volume":"9 3","pages":""},"PeriodicalIF":2.5,"publicationDate":"2026-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13304579/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148318146","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Evaluation of Deep Eutectic Solvents for Cryopreservation of the Fish Pathogen Saprolegnia parasitica. 深共熔溶剂对鱼寄生腐殖菌冷冻保存的评价。

IF 2.5

Methods and Protocols Pub Date : 2026-06-01 DOI: 10.3390/mps9030085

Sara Delimar, Ela Šarić, Marina Cvjetko Bubalo, Ana Bielen

{"title":"Evaluation of Deep Eutectic Solvents for Cryopreservation of the Fish Pathogen Saprolegnia parasitica.","authors":"Sara Delimar, Ela Šarić, Marina Cvjetko Bubalo, Ana Bielen","doi":"10.3390/mps9030085","DOIUrl":"10.3390/mps9030085","url":null,"abstract":"Saprolegnia parasitica (Oomycota) causes saprolegniosis and poses significant ecological and economic challenges in aquaculture. Experimental research on this pathogen is constrained by the lack of reliable long-term preservation methods, as routine maintenance by serial subculturing is labor-intensive and may result in genetic and phenotypic instability. Deep eutectic solvents (DESs), tunable low-melting mixtures, have recently gained attention as alternative cryoprotectants. However, their application has not been evaluated in oomycetes. Here, twelve glycerol-based two- and multicomponent DESs were assessed for cryopreservation of S. parasitica at -80 °C and compared with glycerol as a conventional cryoprotectant. Cryopreservation efficiency was assessed based on post-thaw survival and mycelial regeneration. Several two-component DESs, particularly glycerol-trehalose, supported 100% survival and high post-thaw mycelial regeneration, performing comparably to glycerol under the tested conditions. Shorter pre-incubation (30 min vs. 1 h and 3 h) and controlled-rate freezing (vs. direct freezing) significantly improved post-thaw growth. Although survival remained 100% under optimized conditions, extending storage from 7 to 32 days significantly reduced mycelial regeneration in the glycerol-trehalose treatment, indicating that survival alone, as done in existing literature, does not reflect physiological recovery. Overall, our results support the use of selected DESs as alternative cryoprotectants in oomycetes and contribute to the development of cryopreservation strategies for S. parasitica.","PeriodicalId":18715,"journal":{"name":"Methods and Protocols","volume":"9 3","pages":""},"PeriodicalIF":2.5,"publicationDate":"2026-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13304509/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148318200","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Boron-Doped Carbon Dots for Organelle Labeling and Mitochondrial Bioimaging. 硼掺杂碳点用于细胞器标记和线粒体生物成像。

IF 2.5

Methods and Protocols Pub Date : 2026-06-01 DOI: 10.3390/mps9030086

Aasia Bibi, Daniela De Benedictis, Giuseppe Capitanio, Alessandra Gabriele, Alessandro Buccolieri, Mariapompea Cutroneo, Lorenzo Torrisi, Daniela E Manno, Antonio Serra, Domenico De Rasmo, Anna Signorile

{"title":"Boron-Doped Carbon Dots for Organelle Labeling and Mitochondrial Bioimaging.","authors":"Aasia Bibi, Daniela De Benedictis, Giuseppe Capitanio, Alessandra Gabriele, Alessandro Buccolieri, Mariapompea Cutroneo, Lorenzo Torrisi, Daniela E Manno, Antonio Serra, Domenico De Rasmo, Anna Signorile","doi":"10.3390/mps9030086","DOIUrl":"10.3390/mps9030086","url":null,"abstract":"Background: Carbon dots (CDs) are promising fluorescent nanomaterials with great application potential in bioimaging and organelle-targeted diagnostics. This study compares nitrogen-doped (N-CDs) and boron-nitrogen co-doped CDs (BN-CDs) in normal NIH3T3 fibroblasts and KRAS-transformed cells.Methods: CDs were synthesized via a microwave-assisted method. Their fluorescence, cytocompatibility, and intracellular localization were evaluated using confocal microscopy, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays, and organelle colocalization. Cellular metabolism was assessed by Seahorse analysis. Oxidative stress and cAMP levels were pharmacologically modulated.Results: BN-CDs exhibited stronger intracellular fluorescence than N-CDs, indicating enhanced uptake and imaging performance, with no cytotoxicity up to 100 µg/mL. They localized to multiple organelles, particularly mitochondria. However, fluorescence was significantly reduced in KRAS-transformed cells despite similar mitochondrial mass. BN-CDs did not affect mitochondrial respiration or glycolytic activity. Induced oxidative stress or elevated cAMP in normal cells reduced BN-CD fluorescence.Conclusions: Boron doping improves N-CD imaging properties without affecting cell viability or metabolism. Reduced fluorescence in KRAS cells is associated with altered intracellular conditions, suggesting that BN-CDs could be used to discriminate between normal and cancer cells.","PeriodicalId":18715,"journal":{"name":"Methods and Protocols","volume":"9 3","pages":""},"PeriodicalIF":2.5,"publicationDate":"2026-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13306065/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148318141","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Triple-Survival Stereotactic Brain Surgeries for the Intracranial Injections of Glioblastoma Stem-like Cells and Oncolytic Herpes Simplex Viruses. 颅内注射胶质母细胞瘤干细胞样细胞和溶瘤性单纯疱疹病毒的三存活立体定向脑手术。

IF 2.5

Methods and Protocols Pub Date : 2026-05-31 DOI: 10.3390/mps9030082

Sourav Chakraborty, Connor Howard, Checo J Rorie, Samuel D Rabkin, Hiroaki Wakimoto, Dipongkor Saha

{"title":"Triple-Survival Stereotactic Brain Surgeries for the Intracranial Injections of Glioblastoma Stem-like Cells and Oncolytic Herpes Simplex Viruses.","authors":"Sourav Chakraborty, Connor Howard, Checo J Rorie, Samuel D Rabkin, Hiroaki Wakimoto, Dipongkor Saha","doi":"10.3390/mps9030082","DOIUrl":"10.3390/mps9030082","url":null,"abstract":"Glioblastoma (GBM) is an aggressive primary brain tumor associated with poor prognosis and resistance to therapy, underscoring the need for reliable preclinical models to evaluate emerging treatments. The orthotopic implantation of GBM stem-like cells (GSCs), combined with the intratumoral delivery of therapeutic agents, represents a widely used approach for modeling GBM tumor growth and studying treatment response. In particular, oncolytic herpes simplex viruses (oHSVs) have emerged as a promising strategy to selectively target malignant cells while inducing antitumor immune responses with minimal systemic toxicity. However, performing repeated survival stereotactic neurosurgeries in the same animal poses significant technical challenges. Here, we describe a comprehensive and reproducible protocol for triple survival stereotactic neurosurgery in mice. This approach involves (i) the intracranial implantation of GSCs to establish orthotopic tumors, (ii) the intratumoral delivery of oHSV using the same stereotactic coordinates, and (iii) contralateral intracranial rechallenge with GSCs to evaluate therapeutic efficacy and resistance to tumor rechallenge as a measure of immune memory. Using this protocol, consistent tumor establishment was achieved, and mice tolerated repeated neurosurgical procedures with stable postoperative recovery. Successful intracranial rechallenge in the same animal demonstrates the technical feasibility of multiple survival surgeries while minimizing procedure-related variability and complications. This method enables longitudinal assessment of tumor progression, therapeutic response, and durable memory protection within a single subject. Furthermore, this protocol provides a versatile platform for evaluating oncolytic virotherapy and other localized treatment strategies for GBM.","PeriodicalId":18715,"journal":{"name":"Methods and Protocols","volume":"9 3","pages":""},"PeriodicalIF":2.5,"publicationDate":"2026-05-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13305860/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148318253","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0