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Traditional flat agar pads restrict control of worm orientation, often yielding predominantly lateral views that reduce clarity of dorsal-ventral structures, especially in older, more pigmented animals. A new approach uses channeled agarose pads to precisely rotate and fix worms in defined positions, bringing targets closer to the objective, enabling simultaneous imaging of multiple structures, and improving resolution for regenerated neuronal fiber studies. The method uses the same time and materials as standard pads, supporting broad laboratory adoption.",{"@graph":69,"@context":122},[70,84,105],{"@type":71,"itemListElement":72},"BreadcrumbList",[73,77,79,82],{"item":74,"name":75,"@type":76,"position":8},"https://docshare.wps.com","Home","ListItem",{"item":78,"name":9,"@type":76,"position":14},"https://docshare.wps.com/document/",{"item":80,"name":40,"@type":76,"position":81},"https://docshare.wps.com/document/research-report/",3,{"item":83,"name":65,"@type":76,"position":19},"https://docshare.wps.com/document/fine-adjustment-of-caenorhabditis-elegans-orientation-on-channeled-agar-pads-for-imaging-neuroregeneration/432951/",{"url":83,"name":65,"@type":85,"image":86,"author":91,"headline":65,"publisher":94,"fileFormat":97,"inLanguage":63,"description":67,"dateModified":98,"datePublished":99,"encodingFormat":97,"isAccessibleForFree":100,"interactionStatistic":101},"DigitalDocument",{"url":87,"@type":88,"width":89,"height":90},"https://docshare.wps.com/thumbnails/fine-adjustment-of-caenorhabditis-elegans-orientation-on-channeled-agar-pads-for-imaging-neuroregeneration/432951.png","ImageObject",300,407,{"name":92,"@type":93},"pop out","Person",{"url":74,"name":95,"@type":96},"DocShare","Organization","application/pdf","2026-09-30","2026-09-29",true,{"@type":102,"interactionType":103,"userInteractionCount":14},"InteractionCounter",{"@type":104},"ViewAction",{"@type":106,"mainEntity":107},"FAQPage",[108,114,118],{"name":109,"@type":110,"acceptedAnswer":111},"Why do traditional flat agar pads limit imaging quality in C. elegans?","Question",{"text":112,"@type":113},"They provide limited control over animal orientation, so worms tend to revert to lateral views after immobilization. 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Defined orientation helps place relevant structures closer to the microscope objective, improving image resolution and visualization.","https://schema.org",{"og:url":83,"og:type":124,"og:title":65,"og:site_name":95,"og:description":67},"article",{"robots":126,"canonical":83},"index,follow",{"doc_id":128,"site_id":62},432951,1790741800,{"code":4,"msg":5,"data":131},{"doc_id":128,"user_id":132,"nickname":92,"user_avatar":133,"doc_module":4,"category_id":39,"category_name":40,"doc_title":65,"doc_description":67,"doc_content":134,"file_id":135,"file_url":136,"file_type":137,"file_size":138,"view_count":14,"is_deleted":4,"is_public":8,"is_downloadable":8,"audit_status":8,"page_count":139,"language":140,"language_code":63,"site_id":62,"html_lang":63,"table_of_contents":141,"faqs":142,"seo_title":143,"seo_description":67,"update_tm":144,"read_time":145},962090760408,"https://ap-avatar.wpscdn.com/davatar_085a072bc5b1113ac321206ff7593b45","Author Manuscript Author Manuscript Author Manuscript Author Manuscript  \n\n| | HHS Public Access\u003Cbr>Author manuscript\u003Cbr>J Vis Exp. Author manuscript; available in PMC 2026 January 04. |\n| --- | --- |\n\nFine Adjustment of Caenorhabditis elegans Orientation on Channeled Agar Pads for Imaging Neuroregeneration  \nTina Thuy N. Nguyen Hoang 1 , Chirayu P. Sanganeria 1 , Samuel H. Chung 1  \n1 Department of Bioengineering, Northeastern University  \nAbstract  \nCaenorhabditis elegans is a model organism widely used for studying biological processes. Its transparency and small size make it ideal for imaging tissues, cells, and subcellular structures.  \nTraditional flat agar pads for imaging C. elegans limit control over the animal’s orientation, restricting views primarily to lateral perspectives. This limitation complicates the visualization of dorsal-ventral structures and reduces image clarity, especially in older animals with increased pigmentation and larger diameters. To overcome these challenges, we developed channeled agarose pads that allow precise control of animal orientation. These channels enable researchers to rotate and fix C. elegans in specified positions, facilitating the simultaneous imaging of multiple structures and improving image resolution by bringing target cells closer to the microscope objective. This is particularly useful for imaging regenerated neuronal fibers after surgery, which may grow in directions difficult to capture with traditional flat agar pads. This method is accessible, as fabricating channeled agar pads requires the same time and materials as flat pads, making it a practical option for most laboratories.  \nIntroduction  \nThe nematode Caenorhabditis elegans ( C. elegans) is a widely utilized model organism for studying biological processes such as development and behavior due to its strongly conserved genetics with mammalian counterparts 1. Researchers cultivate C. elegans on a nematode growth medium (NGM) plate in a tightly controlled environment that standardizes experimental conditions and allows for more precise attribution of results to experimental variables. The animal is transparent, enabling clear imaging of tissues, cells, and subcellular structures. C. elegans are small, with the adult animals being ~1 mm in length and 80 μmin diameter, which permits the placement of multiple animals on a single slide mount for imaging 1. Their small size also allows a single image to capture the whole organism while resolving individual cells, which is crucial for visualizing the regeneration of neuron fibers. Synergistic to these advantages for microscopy, the rapid development, stereotyped anatomy, and facile genetics of C. elegans allow for large-scale studies 1.  \nCreative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License Corresponding Author: Samuel H. Chung, [schung@northeastern.edu](schung@northeastern.edu).  \nA complete version of this article that includes the video component is available at [http://dx.doi.org/10.3791/67811](http://dx.doi.org/10.3791/67811) . Disclosures  \nThe authors have no conflicts of interest to disclose.  \nAuthor Manuscript Author Manuscript Author Manuscript Author Manuscript  \nNguyen Hoang et al. Page 2  \nThe typical procedure for imaging of cells in C. elegans was established decades ago and has largely remained unchanged. Researchers use flat agar pads on slides to mount and immobilize the animals for visualization of their cells2. A coverslip on top of the pad holds the animals in place and protects the lens ofthe microscope. Importantly, the coverslip has a high refractive index that increases the numerical aperture of captured light and improves imaging resolution. Additionally, the coverslip reduces distortion in light transmission. Thus, a coverslip improves the visualization of target cells and structures.  \nWhen used with a coverslip, however, flat agar pads often restrict animal orientation. C.  \nelegans bend along their dorsal-ventral (DV) di","cbCaimzLV7pUSMdJ","https://ap.wps.com/l/cbCaimzLV7pUSMdJ","pdf",1138766,11,"English","# Abstract\n# Introduction","[{\"question\":\"Why do traditional flat agar pads limit imaging quality in C. elegans?\",\"answer\":\"They provide limited control over animal orientation, so worms tend to revert to lateral views after immobilization. This complicates imaging of dorsal-ventral structures and can reduce clarity, particularly for older, more pigmented animals.\"},{\"question\":\"How do channeled agarose pads improve orientation control?\",\"answer\":\"The channels let researchers rotate and fix C. elegans into specified positions. The pad design positions animals in depressions between raised agar surfaces, reducing forces during coverslip placement and eliminating undesired rotation.\"},{\"question\":\"Why is this method especially useful for imaging regenerated neuronal fibers?\",\"answer\":\"Regenerated neuronal fibers may grow in directions that are difficult to capture with predominantly lateral views. Defined orientation helps place relevant structures closer to the microscope objective, improving image resolution and visualization.\"}]","Fine Adjustment of Caenorhabditis elegans Orientation on Channeled Agar Pads for Imaging Neuroregeneration | PDF",1790662056,28]