{"title":"Analysis of Spatial Distributions of the Number of Photon Counts in Fluorescence Fluctuation Spectroscopy","authors":"V. V. Skakun, M. A. Hink, V. V. Apanasovich","doi":"10.1007/s10812-024-01788-8","DOIUrl":null,"url":null,"abstract":"<p>A method is proposed for analysis of the photon-count number distributions (PCD) calculated over the space (pixels) of a stack of fluorescent images obtained in the process of a scanning measurement in fluorescence fluctuation spectroscopy which makes it possible to determine the characteristic brightness and number of molecules of the material being studied. The method is applicable to ergodic systems and is based on a theory of the analysis of Photon Counting Histograms (PCH) developed for single-point measurements. The method is tested on experimentally obtained images of a green fluorescent protein. The obtained results are compared with the results of a single-point experiment and the N&B (Number and Brightness) method, which is most often used for numerical analysis of a stack of fluorescent images obtained in scanning experiments. The estimates of the characteristic brightness and number of molecules of the studied substance are in good agreement with estimates obtained with the aid of the methods of analysis used in this region, which makes it possible to conclude that the theory of the PCH method can be used for analyzing spatial PCDs calculated on the basis of a stack of images. The developed method makes it possible to obtain estimates of the studied parameters based on a selected subregion of a single frame of the image.</p>","PeriodicalId":609,"journal":{"name":"Journal of Applied Spectroscopy","volume":"91 4","pages":"803 - 811"},"PeriodicalIF":0.8000,"publicationDate":"2024-09-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Applied Spectroscopy","FirstCategoryId":"92","ListUrlMain":"https://link.springer.com/article/10.1007/s10812-024-01788-8","RegionNum":4,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"SPECTROSCOPY","Score":null,"Total":0}
引用次数: 0
Abstract
A method is proposed for analysis of the photon-count number distributions (PCD) calculated over the space (pixels) of a stack of fluorescent images obtained in the process of a scanning measurement in fluorescence fluctuation spectroscopy which makes it possible to determine the characteristic brightness and number of molecules of the material being studied. The method is applicable to ergodic systems and is based on a theory of the analysis of Photon Counting Histograms (PCH) developed for single-point measurements. The method is tested on experimentally obtained images of a green fluorescent protein. The obtained results are compared with the results of a single-point experiment and the N&B (Number and Brightness) method, which is most often used for numerical analysis of a stack of fluorescent images obtained in scanning experiments. The estimates of the characteristic brightness and number of molecules of the studied substance are in good agreement with estimates obtained with the aid of the methods of analysis used in this region, which makes it possible to conclude that the theory of the PCH method can be used for analyzing spatial PCDs calculated on the basis of a stack of images. The developed method makes it possible to obtain estimates of the studied parameters based on a selected subregion of a single frame of the image.
期刊介绍：
Journal of Applied Spectroscopy reports on many key applications of spectroscopy in chemistry, physics, metallurgy, and biology. An increasing number of papers focus on the theory of lasers, as well as the tremendous potential for the practical applications of lasers in numerous fields and industries.
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