Cold Spring Harbor protocols最新文献_第5页

Examining Protein Localization in Aedes aegypti Cells, Tissues, and Organs: Whole-Mount Immunohistochemistry. 检查埃及伊蚊细胞、组织和器官中的蛋白质定位：整装免疫组化。

Cold Spring Harbor protocols Pub Date : 2024-12-02 DOI: 10.1101/pdb.prot108281

Salwa Afifi, Farwa Sajadi, Jean-Paul V Paluzzi

{"title":"Examining Protein Localization in Aedes aegypti Cells, Tissues, and Organs: Whole-Mount Immunohistochemistry.","authors":"Salwa Afifi, Farwa Sajadi, Jean-Paul V Paluzzi","doi":"10.1101/pdb.prot108281","DOIUrl":"10.1101/pdb.prot108281","url":null,"abstract":"Immunohistochemistry (IHC) is a powerful technique used for visualizing cellular components and determining the presence and/or location of proteins or other macromolecules in tissue samples. The classical IHC process involves the detection of epitopes using a highly specific primary antibody. This is followed by a secondary antibody that is coupled to a reporter molecule or fluorophore and capable of binding to the primary antibody and allowing for protein immunodetection. Although IHC does not routinely provide quantitative results compared to an enzyme-linked immunoassay or western blotting, it permits the localization of the proteins in intact tissues. This protocol describes an IHC assay for whole-body Aedes aegypti mosquito tissues that is used to detect small proteins, specifically neuropeptide hormones. This method is useful for protein detection in whole-mount preparations; however, cross-section IHC is recommended to determine if a protein is localized in the apical versus basolateral membrane of tissues/organs or to visualize immunological distribution in larger, more complex preparations.","PeriodicalId":10496,"journal":{"name":"Cold Spring Harbor protocols","volume":" ","pages":"pdb.prot108281"},"PeriodicalIF":0.0,"publicationDate":"2024-12-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138795544","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Study of Dendrite Differentiation Using Drosophila Dendritic Arborization Neurons. 利用果蝇树突分化神经元研究树突分化

Cold Spring Harbor protocols Pub Date : 2024-12-02 DOI: 10.1101/pdb.top108146

Jason Y Tann, Fangke Xu, Minami Kimura, Oliver R Wilkes, Li-Foong Yoong, Henrik Skibbe, Adrian W Moore

引用次数: 0

Mounting of Embryos, Larvae, and Pupae for Live Drosophila Dendritic Arborization Neuron Imaging. 为果蝇树突分化神经元活体成像安装胚胎、幼虫和蛹。

Cold Spring Harbor protocols Pub Date : 2024-12-02 DOI: 10.1101/pdb.prot108149

Fangke Xu, Jason Y Tann, Oliver R Wilkes, Minami Kimura, Li-Foong Yoong, Adrian W Moore

引用次数: 0

Mosaic Analysis with a Repressible Cell Marker (MARCM) Clone Generation in Drosophila Dendritic Arborization Neurons. 果蝇树突分化神经元可抑制细胞标记（MARCM）克隆生成的镶嵌分析。

Cold Spring Harbor protocols Pub Date : 2024-12-02 DOI: 10.1101/pdb.prot108147

Jason Y Tann, Fangke Xu, Minami Kimura, Adrian W Moore

{"title":"Mosaic Analysis with a Repressible Cell Marker (MARCM) Clone Generation in Drosophila Dendritic Arborization Neurons.","authors":"Jason Y Tann, Fangke Xu, Minami Kimura, Adrian W Moore","doi":"10.1101/pdb.prot108147","DOIUrl":"10.1101/pdb.prot108147","url":null,"abstract":"Mosaic analysis with a repressible cell marker (MARCM) is used in Drosophila research to create labeled homozygous mutant clones of cells in an otherwise heterozygous fly. It allows the study of the effect of embryonically lethal genes and the determination of cell autonomy for a mutant phenotype. When used in dendritic arborization (da) neurons with a fluorescent protein targeted to the plasma membrane, MARCM allows the identification of homozygous mutant neurons and clear imaging of the dendrite arbor in both live and fixed preparations. Previous protocols that outlined experimental procedures to create MARCM clones in da neurons used a heat shock promoter to drive Flippase (FLP) expression; such an approach requires laborious embryo collection and heat shock steps, and it creates clones in other tissues besides the da neurons. The updated protocol described here outlines the use of FLP expression driven by a sensory organ precursor promoter (SOP-FLP); it requires no embryo collection or manipulation steps and creates clones exclusively in the peripheral sensory neuron lineage.","PeriodicalId":10496,"journal":{"name":"Cold Spring Harbor protocols","volume":" ","pages":"pdb.prot108147"},"PeriodicalIF":0.0,"publicationDate":"2024-12-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139039617","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Culture of Larval and Pupal Drosophila Dendritic Arborization Neurons. 幼虫和蛹果蝇树突分化神经元的培养

Cold Spring Harbor protocols Pub Date : 2024-12-02 DOI: 10.1101/pdb.prot108150

Jason Y Tann, Fangke Xu, Li-Foong Yoong, Adrian W Moore

引用次数: 0

Optimized Methods for Applying and Assessing Heat, Drought, and Nutrient Stress of Maize Seedlings in Controlled Environment Experiments. 在受控环境实验中应用和评估玉米幼苗热、旱和养分胁迫的优化方法。

Cold Spring Harbor protocols Pub Date : 2024-11-18 DOI: 10.1101/pdb.top108467

Alejandra Quiñones, Leonardo W Lima, Katherine M Murphy, Anna L Casto, Malia A Gehan, Cory D Hirsch

{"title":"Optimized Methods for Applying and Assessing Heat, Drought, and Nutrient Stress of Maize Seedlings in Controlled Environment Experiments.","authors":"Alejandra Quiñones, Leonardo W Lima, Katherine M Murphy, Anna L Casto, Malia A Gehan, Cory D Hirsch","doi":"10.1101/pdb.top108467","DOIUrl":"10.1101/pdb.top108467","url":null,"abstract":"Maize (Zea mays), also known as corn, is an important crop that plays a crucial role in global agriculture. The economic uses of maize are numerous, including for food, feed, fiber, and fuel. It has had a significant historical importance in research as well, with important discoveries made in maize regarding plant domestication, transposons, heterosis, genomics, and epigenetics. Unfortunately, environmental stresses cause substantial yield loss to maize crops each year. Yield losses are predicted to increase in future climate scenarios, posing a threat to food security and other sectors of the global economy. Developing efficient methods to study maize abiotic stress responses is a crucial step toward a more resilient and productive agricultural system. This review describes the importance of and methods for studying the effects of heat, drought, and nutrient deficiency on early developmental stages of maize grown in controlled environments. Studying the early effects of environmental stressors in controlled environments allows researchers to work with a variety of environmental conditions with low environmental variance, which can inform future field-based research. We highlight the current knowledge of physiological responses of maize to heat, drought, and nutrient stress; remaining knowledge gaps and challenges; and information on how standardized protocols can address these issues.","PeriodicalId":10496,"journal":{"name":"Cold Spring Harbor protocols","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-11-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142459804","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Selection of Affibody Molecules Using Phage Display. 利用噬菌体展示选择亲和体分子。

Cold Spring Harbor protocols Pub Date : 2024-11-01 DOI: 10.1101/pdb.prot108399

Linnea Charlotta Hjelm, Charles Dahlsson Leitao, Stefan Ståhl, John Löfblom, Hanna Lindberg

引用次数: 0

Engineering of Affibody Molecules. Affibody 分子工程。

Cold Spring Harbor protocols Pub Date : 2024-11-01 DOI: 10.1101/pdb.top107760

Stefan Ståhl, Hanna Lindberg, Linnea Charlotta Hjelm, John Löfblom, Charles Dahlsson Leitao

引用次数: 0

Activity Monitoring for Analysis of Sleep in Drosophila melanogaster. 通过活动监测分析黑腹果蝇的睡眠情况

Cold Spring Harbor protocols Pub Date : 2024-11-01 DOI: 10.1101/pdb.top108095

Divya Sitaraman, Christopher G Vecsey, Casey Koochagian

引用次数: 0

Selection of Affibody Molecules Using Escherichia coli Display. 利用大肠杆菌显示筛选亲和体分子。

Cold Spring Harbor protocols Pub Date : 2024-11-01 DOI: 10.1101/pdb.prot108400

Charles Dahlsson Leitao, Linnea Charlotta Hjelm, Stefan Ståhl, John Löfblom, Hanna Lindberg

{"title":"Selection of Affibody Molecules Using Escherichia coli Display.","authors":"Charles Dahlsson Leitao, Linnea Charlotta Hjelm, Stefan Ståhl, John Löfblom, Hanna Lindberg","doi":"10.1101/pdb.prot108400","DOIUrl":"10.1101/pdb.prot108400","url":null,"abstract":"Affibody molecules are small (6-kDa) affinity proteins generated by directed evolution for specific binding to various target molecules. The first step in this workflow involves the generation of an affibody library, which can then be used for selection via multiple display methods. This protocol describes selection from affibody libraries by Escherichia coli cell surface display. With this method, high-diversity libraries of 1011 can be displayed on the cell surface. The method involves two steps for selection of binders from high-diversity libraries: magnetic-activated cell sorting (MACS) and fluorescence-activated cell sorting (FACS). MACS is used first to enrich the library in target-binding clones and to decrease diversity to a size that can be effectively screened and sorted in the flow cytometer in a reasonable time (typically <107 cells). The protocol is based on methodology using an AIDA-I autotransporter for display on the outer membrane, but the general procedures can also be adjusted and used for other types of autotransporters or alternative E. coli display methods.","PeriodicalId":10496,"journal":{"name":"Cold Spring Harbor protocols","volume":" ","pages":"pdb.prot108400"},"PeriodicalIF":0.0,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9924643","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0