{"title":"共聚焦显微镜中半透明层深度分辨率建模","authors":"Maximilian Maier, Thomas Böhm","doi":"10.1002/smsc.202400120","DOIUrl":null,"url":null,"abstract":"Confocal microscopy is an established technique with manifold applications that offers the capability to perform nondestructive through-plane imaging. However, depth resolution typically decreases when focusing below the surface of a sample, which limits the applicability. A computational model is introduced that calculates the axial resolution, its decay, and the attenuation coefficient from confocal through-plane scans of translucent layers. The model is benchmarked with different polymers and objectives (air, water, oil) using a confocal Raman microscope. The algorithm requires a single through-plane scan that allows to identify the sample by signal intensity differences. It fits the point spread function of the objective at the top and bottom interface of the specimen to extract the resolution at both interfaces and the attenuation coefficient of the sample. It provides robust outputs on various and even multilayered samples if the signal-to-noise ratio of the input is sufficient and if the layers are planar and homogeneous. The algorithm of the model is provided open-source for MATLAB and Python. Quantifying microscope resolution in through-plane scans can improve image analysis in multiple fields, and this study is a comprehensive proof-of-concept for the presented model. It establishes an accessible tool to quantify the depth resolution of confocal microscopy.","PeriodicalId":29791,"journal":{"name":"Small Science","volume":"18 1","pages":""},"PeriodicalIF":11.1000,"publicationDate":"2024-06-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Modeling the Depth Resolution of Translucent Layers in Confocal Microscopy\",\"authors\":\"Maximilian Maier, Thomas Böhm\",\"doi\":\"10.1002/smsc.202400120\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"Confocal microscopy is an established technique with manifold applications that offers the capability to perform nondestructive through-plane imaging. However, depth resolution typically decreases when focusing below the surface of a sample, which limits the applicability. A computational model is introduced that calculates the axial resolution, its decay, and the attenuation coefficient from confocal through-plane scans of translucent layers. The model is benchmarked with different polymers and objectives (air, water, oil) using a confocal Raman microscope. The algorithm requires a single through-plane scan that allows to identify the sample by signal intensity differences. It fits the point spread function of the objective at the top and bottom interface of the specimen to extract the resolution at both interfaces and the attenuation coefficient of the sample. It provides robust outputs on various and even multilayered samples if the signal-to-noise ratio of the input is sufficient and if the layers are planar and homogeneous. The algorithm of the model is provided open-source for MATLAB and Python. Quantifying microscope resolution in through-plane scans can improve image analysis in multiple fields, and this study is a comprehensive proof-of-concept for the presented model. It establishes an accessible tool to quantify the depth resolution of confocal microscopy.\",\"PeriodicalId\":29791,\"journal\":{\"name\":\"Small Science\",\"volume\":\"18 1\",\"pages\":\"\"},\"PeriodicalIF\":11.1000,\"publicationDate\":\"2024-06-25\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Small Science\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.1002/smsc.202400120\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"MATERIALS SCIENCE, MULTIDISCIPLINARY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Small Science","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1002/smsc.202400120","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"MATERIALS SCIENCE, MULTIDISCIPLINARY","Score":null,"Total":0}
Modeling the Depth Resolution of Translucent Layers in Confocal Microscopy
Confocal microscopy is an established technique with manifold applications that offers the capability to perform nondestructive through-plane imaging. However, depth resolution typically decreases when focusing below the surface of a sample, which limits the applicability. A computational model is introduced that calculates the axial resolution, its decay, and the attenuation coefficient from confocal through-plane scans of translucent layers. The model is benchmarked with different polymers and objectives (air, water, oil) using a confocal Raman microscope. The algorithm requires a single through-plane scan that allows to identify the sample by signal intensity differences. It fits the point spread function of the objective at the top and bottom interface of the specimen to extract the resolution at both interfaces and the attenuation coefficient of the sample. It provides robust outputs on various and even multilayered samples if the signal-to-noise ratio of the input is sufficient and if the layers are planar and homogeneous. The algorithm of the model is provided open-source for MATLAB and Python. Quantifying microscope resolution in through-plane scans can improve image analysis in multiple fields, and this study is a comprehensive proof-of-concept for the presented model. It establishes an accessible tool to quantify the depth resolution of confocal microscopy.
期刊介绍:
Small Science is a premium multidisciplinary open access journal dedicated to publishing impactful research from all areas of nanoscience and nanotechnology. It features interdisciplinary original research and focused review articles on relevant topics. The journal covers design, characterization, mechanism, technology, and application of micro-/nanoscale structures and systems in various fields including physics, chemistry, materials science, engineering, environmental science, life science, biology, and medicine. It welcomes innovative interdisciplinary research and its readership includes professionals from academia and industry in fields such as chemistry, physics, materials science, biology, engineering, and environmental and analytical science. Small Science is indexed and abstracted in CAS, DOAJ, Clarivate Analytics, ProQuest Central, Publicly Available Content Database, Science Database, SCOPUS, and Web of Science.