Methods in cell biology最新文献_第2页

Expression, purification and characterization of phosphatidylserine-targeting antibodies for biochemical and therapeutic applications. 用于生化和治疗的磷脂酰丝氨酸靶向抗体的表达、纯化和鉴定。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-11-21 DOI: 10.1016/bs.mcb.2024.10.003

Varsha Gadiyar, David C Calianese, Rachael Pulica, Christopher Varsanyi, Ziren Wang, Ahmed Aquib, Alok Choudhary, Raymond B Birge

引用次数: 0

Development of an in vivo ovarian cancer peritoneal carcinomatosis model for radioimmunotherapy testing. 用于放射免疫治疗试验的卵巢癌腹膜癌模型的建立。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-06-17 DOI: 10.1016/bs.mcb.2024.05.013

Clara Diaz Garcia-Prada, Salima Atis, Jean-Pierre Pouget, Julie Constanzo

{"title":"Development of an in vivo ovarian cancer peritoneal carcinomatosis model for radioimmunotherapy testing.","authors":"Clara Diaz Garcia-Prada, Salima Atis, Jean-Pierre Pouget, Julie Constanzo","doi":"10.1016/bs.mcb.2024.05.013","DOIUrl":"10.1016/bs.mcb.2024.05.013","url":null,"abstract":"Currently, Ovarian Cancer (OC) is the most lethal gynecological malignancy. In most patients, it progresses without clinical signs or symptoms, leading to a late diagnosis when it has already spread in the peritoneal cavity as peritoneal carcinomatosis (PC). To date, OC PC management is based on cytoreductive surgery to remove the macroscopic disease, followed by chemotherapy. Many patients respond to this treatment, but disease recurs in 70-90% of them. Therefore, new therapeutic approaches are needed. The field of targeted radionuclide therapy (TRT) has witnessed considerable progress and several radiopharmaceuticals have been approved in the last decade. In TRT, radiolabeled molecules are injected to specifically recognize, irradiate, and kill tumor cells. TRT is a multisite radiotherapy that delivers dose to all malignant lesions. Therefore, TRT could be an alternative approach for OC PC because conventional external beam radiotherapy cannot be used at curative dose due to toxicity to healthy tissues. Here, we describe an OC PC model based on grafting human SK-OV-3 OC cells in the peritoneal cavity of immunodeficient mice. We also explain how to label trastuzumab with lutetium-177 to specifically target and irradiate SK-OV-3 cell nodules in these mice, and how to monitor the response to this TRT in vivo. With minor variations, the same technique can be conveniently applied to a variety of human (or mouse) tumors.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"192 ","pages":"131-157"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143039848","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}

引用次数: 0

Assessing chromosomal abnormalities in leukemias by imaging flow cytometry. 利用成像流式细胞术评估白血病染色体异常。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2023-05-18 DOI: 10.1016/bs.mcb.2023.04.001

Stephanie J Lam, Henry Y L Hui, Kathy A Fuller, Wendy N Erber

{"title":"Assessing chromosomal abnormalities in leukemias by imaging flow cytometry.","authors":"Stephanie J Lam, Henry Y L Hui, Kathy A Fuller, Wendy N Erber","doi":"10.1016/bs.mcb.2023.04.001","DOIUrl":"10.1016/bs.mcb.2023.04.001","url":null,"abstract":"Chromosome analysis assists in the diagnostic classification and prognostication of leukemias. It is typically performed by karyotyping or fluorescent in situ hybridization (FISH) on glass slides. Flow cytometry offers an alternative high throughput automated methodology to analyze chromosomal content. With the advent of imaging flow cytometers, specific chromosomes and regions of interest can be identified and enumerated within specific cell types. The inclusion of immunophenotyping increases the specificity of this technique to ensure only the leukemic cell is analyzed. With many thousands of cells acquired, and neoplastic cells of interest identified by antigen expression, this technology has expanded the role of flow cytometry for cytogenomics in oncology. Applications to date have focused on hematological malignancies to detect aneuploidy (chromosome gains and losses) and structural defects (e.g., deletions; translocations) of diagnostic or prognostic significance at the time of diagnosis. With limits of detection of 1 cytogenetically abnormal cell in 100,000, also makes this new flow cytometry protocol eminently suitable for monitoring low level disease, detecting clonal evolution after therapy and identifying circulating tumor cells. The technique is equally applicable to solid tumors, many of which have chromosomal aberrations, with selection of appropriate immunophenotypic markers and FISH probes.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"195 ","pages":"71-100"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143780451","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}

引用次数: 0

Retrovirus-based manufacturing of chimeric antigen receptor-modified T cells for cancer therapy research. 基于逆转录病毒制造嵌合抗原受体修饰的T细胞用于癌症治疗研究。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-11-19 DOI: 10.1016/bs.mcb.2024.10.017

Sophia Stock, Luisa Fertig, Vivien Doreen Menkhoff, Thaddäus Strzalkowski, Manuel Caruso, Sebastian Kobold

{"title":"Retrovirus-based manufacturing of chimeric antigen receptor-modified T cells for cancer therapy research.","authors":"Sophia Stock, Luisa Fertig, Vivien Doreen Menkhoff, Thaddäus Strzalkowski, Manuel Caruso, Sebastian Kobold","doi":"10.1016/bs.mcb.2024.10.017","DOIUrl":"10.1016/bs.mcb.2024.10.017","url":null,"abstract":"Treatment with autologous chimeric antigen receptor (CAR)-modified T cells can achieve outstanding clinical response rates in heavily pretreated patients with B and plasma cell malignancies. However, relapses occur, and they limit the efficacy of this promising treatment approach. The complex GMP-compliant production and high treatment costs cause that CAR T cells cannot yet be used in a broad population. Among others, CAR T cell therapy has evolved regarding vector design and manufacturing process. Optimal production of CAR T cells is not yet defined, far from being standardized. Quality, cellular composition and immunophenotype of the administered CAR T cells are influenced by the manufacturing protocol and therefore play a crucial role for therapeutic success. For the gene transfer, viral and non-viral strategies are available. Retrovirus-based protocols for CAR T cell production offer advantages in terms of stable gene integration, sufficient transduction efficiency, proven clinical success, and scalability. Here, we detail a retrovirus-based generation protocol of human CAR-modified T cells for experimental immunotherapeutic treatment of cancer cells. For the CAR generation, HEK-293-based packaging cell lines, CD3+ selection, CD3/CD28-coated bead-based activation and IL-2/IL-15-mediated expansion were used. This protocol can be applied for every possible CAR construct after being successfully transfected in HEK-293-based packaging cell lines.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"191 ","pages":"329-352"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143008161","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}

引用次数: 0

Evaluation of lymphocyte infiltration into cancer spheroids by immunofluorescent staining and 3D imaging. 免疫荧光染色和三维成像评价淋巴细胞浸润癌球体。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-11-14 DOI: 10.1016/bs.mcb.2024.10.015

Mireia Cruz De Los Santos, Andreas Lundqvist

引用次数: 0

Stereotactic injection of murine brain tumor cells for neuro-oncology studies. 立体定向注射小鼠脑肿瘤细胞用于神经肿瘤学研究。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-08-12 DOI: 10.1016/bs.mcb.2024.07.005

Camille Daviaud, María Cecilia Lira, Claire Vanpouille-Box, Mara De Martino

引用次数: 0

Measuring interaction force between T lymphocytes and their target cells using live microscopy and laminar shear flow chambers. 利用活体显微镜和层流剪切室测量T淋巴细胞与靶细胞之间的相互作用力。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-11-08 DOI: 10.1016/bs.mcb.2024.09.001

Sophie Goyard, Amandine Schneider, Jerko Ljubetic, Nicolas Inacio, Marie Juzans, Céline Cuche, Pascal Bochet, Vincenzo Di Bartolo, Andrés Alcover, Thierry Rose

{"title":"Measuring interaction force between T lymphocytes and their target cells using live microscopy and laminar shear flow chambers.","authors":"Sophie Goyard, Amandine Schneider, Jerko Ljubetic, Nicolas Inacio, Marie Juzans, Céline Cuche, Pascal Bochet, Vincenzo Di Bartolo, Andrés Alcover, Thierry Rose","doi":"10.1016/bs.mcb.2024.09.001","DOIUrl":"10.1016/bs.mcb.2024.09.001","url":null,"abstract":"Understanding the immunological synapse formation and dynamics can be enriched by measuring cell-cell interaction forces and their kinetics. Microscopy imaging reveals structural organization of the synapse, while physical methods detail its mechanical construction. Various techniques have been reported for measuring forces needed to rupture the interface between a T lymphocyte and its target cell but most of them measure one pair at a time. We describe here a laminar shear flow-based method that exerts dragging forces on T cell-target cells pairs immobilized on the surface of a flow chamber. Increasing flow rate allows us to observe the detachment of hundreds of cell conjugates on the wide field of a light transmission microscope. Monitoring precisely the flow rate gradient exerted on T cells readily yields synapse rupture measurements. Dragging forces are measured at the point of rupture as a linear function of the flow speed in minutes from 10pN to 20nN for each cell pair among a statistically representative cell population in the whole field of view of a single experiment. The output cells can be collected in multi-well plate sorted in the increasing order of rupture forces. We used this approach to unveil the involvement of the cytoskeleton regulator adenomatous polyposis coli (APC) in the stability of immunological synapses formed between human cytotoxic T cell and tumor target cells. APC is a polarity regulator and tumor suppressor associated with familial adenomatous polyposis and colorectal cancer. Reduced APC expression impairs T cell adhesion with tumor target cells suggesting an impact of APC mutation in anti-tumor immune defense.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"193 ","pages":"175-200"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143370861","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}

引用次数: 0

Ex vivo assessment of human neutrophil motility and migration. 人中性粒细胞运动和迁移的离体评估。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-11-19 DOI: 10.1016/bs.mcb.2024.10.008

Noor A M Bakker, Claudia Burrello, Karin E de Visser

引用次数: 0

In vitro ILC differentiation from human HSCs. 人造血干细胞的体外ILC分化。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-11-19 DOI: 10.1016/bs.mcb.2024.10.004

Silvia Santopolo, Cecilia Ciancaglini, Francesca Romana Mariotti, Lorenzo Moretta, Linda Quatrini

{"title":"In vitro ILC differentiation from human HSCs.","authors":"Silvia Santopolo, Cecilia Ciancaglini, Francesca Romana Mariotti, Lorenzo Moretta, Linda Quatrini","doi":"10.1016/bs.mcb.2024.10.004","DOIUrl":"10.1016/bs.mcb.2024.10.004","url":null,"abstract":"The Innate Lymphoid Cells (ILCs) are a family of innate immune cells composed by the Natural Killer (NK) cells and the helper ILCs (hILCs) (ILC1, ILC2, ILC3), both developing from a common ILC precursor (ILCP) derived from hematopoietic stem cells (HSCs). A correct ILC reconstitution is crucial, particularly in patients receiving HSC transplantation (HSCT), the only therapeutic option for many adult and pediatric high-risk hematological malignancies. Indeed, mainly thanks to their cytotoxic activity, NK cells have a strong Graft-versus-Leukemia (GvL) effect. On the other hand, hILCs, that are mainly tissue resident, are involved in tissue repair and homeostasis, Graft-versus-Host Disease (GvHD) prevention and immune response to infections. Unlike NK cell development, hILC-poiesis is still poorly characterized in humans. Here, we provide a protocol for the in vitro ILC differentiation from healthy donor peripheral blood-derived CD34+ HSCs. This could represent a useful model to dissect the molecular mechanisms by which the distinct ILC subsets are generated from ILCP leading to the development of novel strategies to improve the HSCT clinical outcome.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"191 ","pages":"41-57"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143008130","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}

引用次数: 0

Bacterial predators and BALOs: Growth protocol and relation with mitochondria. 细菌捕食者和BALOs：生长方案和与线粒体的关系。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-08-24 DOI: 10.1016/bs.mcb.2024.07.003

Valerio Iebba

{"title":"Bacterial predators and BALOs: Growth protocol and relation with mitochondria.","authors":"Valerio Iebba","doi":"10.1016/bs.mcb.2024.07.003","DOIUrl":"10.1016/bs.mcb.2024.07.003","url":null,"abstract":"The microbial world is characterized by mechanisms of competition and predation, akin to the animal world. However, while predation's ecological role is well-established in animals, it's less understood in bacteria due to fewer known predators and unclear phylogenetic affiliations. Nevertheless, microorganisms can prey on bacterial cells, including Bacteriophages, Protists, and Predatory Prokaryotes. These predators inhabit various habitats and may play vital roles in bacterial ecology and ecosystem regulation. Predatory interactions between host and parasite are common in nature. Predatory bacteria, such as Bdellovibrio and like organisms (BALOs), employ various strategies, including epibiotic predation and direct invasion. BALOs, which thrive in the periplasmic space of Gram-negative bacterial cells, modulate bacterial populations and could serve as preventive or therapeutic agents against Gram-negative infections. While primarily active against extracellular prey, BALOs may also target mitochondria, which are crucial for cellular processes. The relationship between intracellular bacteria and host mitochondria, including morphology, function, and apoptosis, warrants further exploration. Protocols for growing, propagating, and detecting predatory activities of BALOs, particularly Bdellovibrio bacteriovorus, are provided to assess their presence and activities against potential prey.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"194 ","pages":"151-167"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143586227","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}

引用次数: 0