[{"data":1,"prerenderedAt":-1},["ShallowReactive",2],{"doc-seo-357335-105":3,"detail-sidebar-cat-0-en-105":81,"doc-detail-357335-en":130},{"code":4,"msg":5,"data":6},0,"ok",{"site_id":7,"language":8,"slug":9,"title":10,"keywords":11,"description":12,"schema_data":13,"social_meta":74,"head_meta":76,"extra_data":78,"updated_unix":80},105,"en","optimizing-an-ethanol-based-fixative-for-enhanced-nucleic-acid-preservation-in-cervical-samples-using-a-central-composite-design-approach","Optimizing an ethanol-based fixative for enhanced nucleic acid preservation in cervical samples using a central composite design approach","","High-quality DNA and RNA from cervical specimens are essential for accurate cervical cancer diagnosis, yet commonly used preservation approaches can compromise RNA stability and raise safety concerns. This study develops an optimized ethanol-based fixative for ambient-condition storage. HeLa cells and clinical samples were evaluated using a central composite design, varying pH, sorbitol, and PEG 8000. Nucleic acid integrity was assessed by gel electrophoresis, PCR, qRT-PCR, and HPV genotyping, alongside Papanicolaou-based morphology.",{"@graph":14,"@context":73},[15,34,56],{"@type":16,"itemListElement":17},"BreadcrumbList",[18,23,27,31],{"item":19,"name":20,"@type":21,"position":22},"https://docshare.wps.com","Home","ListItem",1,{"item":24,"name":25,"@type":21,"position":26},"https://docshare.wps.com/document/","Document",2,{"item":28,"name":29,"@type":21,"position":30},"https://docshare.wps.com/document/healthcare/","Healthcare",3,{"item":32,"name":10,"@type":21,"position":33},"https://docshare.wps.com/document/optimizing-an-ethanol-based-fixative-for-enhanced-nucleic-acid-preservation-in-cervical-samples-using-a-central-composite-design-approach/357335/",4,{"url":32,"name":10,"@type":35,"image":36,"author":41,"headline":10,"publisher":44,"fileFormat":47,"inLanguage":8,"description":12,"dateModified":48,"datePublished":49,"encodingFormat":47,"isAccessibleForFree":50,"interactionStatistic":51},"DigitalDocument",{"url":37,"@type":38,"width":39,"height":40},"https://docshare.wps.com/thumbnails/optimizing-an-ethanol-based-fixative-for-enhanced-nucleic-acid-preservation-in-cervical-samples-using-a-central-composite-design-approach/357335.png","ImageObject",300,407,{"name":42,"@type":43},"Aldword","Person",{"url":19,"name":45,"@type":46},"DocShare","Organization","application/pdf","2026-10-23","2026-09-23",true,{"@type":52,"interactionType":53,"userInteractionCount":55},"InteractionCounter",{"@type":54},"ViewAction",7,{"@type":57,"mainEntity":58},"FAQPage",[59,65,69],{"name":60,"@type":61,"acceptedAnswer":62},"What is the main goal of the study on cervical samples?","Question",{"text":63,"@type":64},"To optimize an ethanol-based fixative that preserves DNA and RNA from cervical specimens stored under ambient conditions for reliable molecular and cytological testing.","Answer",{"name":66,"@type":61,"acceptedAnswer":67},"Which experimental factors were optimized using the central composite design?",{"text":68,"@type":64},"pH, Sorbitol concentration, and PEG concentration, assessed for their effects on nucleic acid preservation quality.",{"name":70,"@type":61,"acceptedAnswer":71},"How did the optimized fixative perform compared with the commercial PreservCyt solution?",{"text":72,"@type":64},"It significantly improved DNA and RNA preservation, showing high DNA integrity with successful PCR amplification (including HPV-18 oncogene targeting) and clear rRNA bands with lower qRT-PCR Ct values.","https://schema.org",{"og:url":32,"og:type":75,"og:title":10,"og:site_name":45,"og:description":12},"article",{"robots":77,"canonical":32},"index,follow",{"doc_id":79,"site_id":7},357335,1790377393,{"code":4,"msg":82,"data":83},"success",[84,88,92,96,101,106,109,114,119,122,126],{"id":22,"doc_module":4,"doc_module_name":25,"category_name":85,"show_sort_weight":86,"slug":87},"Story & Novel",90,"story-novel",{"id":26,"doc_module":4,"doc_module_name":25,"category_name":89,"show_sort_weight":90,"slug":91},"Literature",80,"literature",{"id":33,"doc_module":4,"doc_module_name":25,"category_name":93,"show_sort_weight":94,"slug":95},"Exam",70,"exam",{"id":97,"doc_module":4,"doc_module_name":25,"category_name":98,"show_sort_weight":99,"slug":100},5,"Comic",60,"comic",{"id":102,"doc_module":4,"doc_module_name":25,"category_name":103,"show_sort_weight":104,"slug":105},6,"Technology",50,"technology",{"id":55,"doc_module":4,"doc_module_name":25,"category_name":29,"show_sort_weight":107,"slug":108},40,"healthcare",{"id":110,"doc_module":4,"doc_module_name":25,"category_name":111,"show_sort_weight":112,"slug":113},8,"Research & Report",30,"research-report",{"id":115,"doc_module":4,"doc_module_name":25,"category_name":116,"show_sort_weight":117,"slug":118},9,"Religion & Spirituality",20,"religion-spirituality",{"id":117,"doc_module":4,"doc_module_name":25,"category_name":120,"show_sort_weight":117,"slug":121},"World Cup","world-cup",{"id":123,"doc_module":4,"doc_module_name":25,"category_name":124,"show_sort_weight":123,"slug":125},10,"Lifestyle","lifestyle",{"id":127,"doc_module":4,"doc_module_name":25,"category_name":128,"show_sort_weight":97,"slug":129},19,"General","general",{"code":4,"msg":82,"data":131},{"doc_id":79,"user_id":132,"nickname":42,"user_avatar":133,"doc_module":4,"category_id":55,"category_name":29,"doc_title":10,"doc_description":12,"doc_content":134,"file_id":135,"file_url":136,"file_type":137,"file_size":138,"view_count":55,"is_deleted":4,"is_public":22,"is_downloadable":22,"audit_status":22,"page_count":117,"language":139,"language_code":8,"site_id":7,"html_lang":8,"table_of_contents":140,"faqs":141,"seo_title":142,"seo_description":12,"update_tm":143,"read_time":104},2336478940917,"https://ap-avatar.wpscdn.com/davatar_6f874abed73319feea01a86fa6f0fab8","OPEN ACCESS  \nCitation: Sedaghat Shayegan G, Salehipour P, Najafi A, Modarressi MH (2026) Optimizing an ethanol-based fixative for enhanced nucleic acid preservation in cervical samples using a central composite design approach. PLoS One 21(6): e0349088. [https://doi.org/10.1371/journal](https://doi.org/10.1371/journal). pone.0349088  \nEditor: Devesh U Kapoor, Dr. Dayaram Patel Pharmacy College, INDIA  \nReceived: July 4, 2025  \nAccepted: April 23, 2026  \nPublished: June 26, 2026  \nCopyright: © 2026 Sedaghat Shayegan et al. This is an open access article distributed under the terms of the Creative Commons Attribution  License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.  \nData availability statement: All relevant data are within the manuscript and its Supporting  Information files.  \nFunding: This work was supported by the Tehran University of Medical Sciences, under  \nRESEARCH ARTICLE  \nOptimizing an ethanol-based fixative for enhanced nucleic acid preservation in cervical samples using a central composite design approach  \nGhazal Sedaghat Shayegan1, Pouya Salehipour1, Ali Najafi1,  \nMohammad Hossein Modarressi1,2*  \n1 Department of Medical Genetics, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran 2 Genomics Department, Nilou Medical Laboratory, Tehran, Iran  \n* [Modaresi@tums.ac.ir](Modaresi@tums.ac.ir)  \nAbstract  \nBackground  \nTo accurately diagnose cervical cancer, high-quality genetic material (DNA and RNA) from clinical samples is crucial. Current preservation methods often have limitations, including poor RNA stability and safety concerns. This study aimed to develop an optimized ethanol-based fixative to preserve DNA and RNA in cervical samples under ambient conditions.  \nMethods  \nHeLa cells were fixed in ethanol-based fixatives containing polyhydric compounds (Sorbitol and polyethylene glycol [PEG]) and chelating agents. We used a central composite design (CCD) approach to evaluate the effects of pH, Sorbitol concentration, and PEG concentration on nucleic acid preservation. DNA and RNA quality were assessed using agarose gel electrophoresis, PCR, and real-time PCR. Cellular morphology was evaluated using Papanicolaou-stained slides. HPV genotyping of clinical samples was conducted using real-time PCR.  \nResults  \nThe optimized fixative, developed in this study consisted of 40% ethanol, 4.3% Sorbitol, 1.2% PEG 8000, with a pH of 5.6. This new formula significantly improved DNA and RNA preservation compared to the commercial solution, PreservCyt. DNA showed high integrity and was successfully amplified in PCR test targeting HPV-18 oncogenes. RNA quality was confirmed through clear 28S and 18S rRNA bands and lower threshold cycle (Ct) values in qRT-PCR. HPV genotyping and morphological  \nPLOS One | [https://doi.org/10.1371/journal.pone.0349088](https://doi.org/10.1371/journal.pone.0349088) June 26, 2026 1 / 20  \nGrant number 1400-1-410-52883. The fundershad no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.  \nCompeting interests: The authors have declared that no competing interests exist.  \nanalysis revealed excellent preservation, enabling both molecular and cytological evaluations.  \nConclusion  \nThe new ethanol-based fixative represents a promising cost-effective and environmentally friendly solution for preserving nucleic acids and cellular morphology in cervical samples. Its good performance under ambient conditions suggests it may serve as an option for Human Papillomavirus (HPV) testing and cervical cancer prevention, particularly in places with limited resources.  \n1. Introduction  \nCervical cancer is an important global health issue that particularly affects the female reproductive system. Cervical cancer is the fourth most common cancer in women worldwide, according to the World Health Organization (WHO) scale [ 1] . Human papillomavirus (H","cbCaink3Ab6iH9VU","https://ap.wps.com/l/cbCaink3Ab6iH9VU","pdf",3715736,"English","# Abstract\n## Background\n## Methods\n## Results\n## Conclusion\n# Introduction\n## Cervical cancer and HPV risk\n## Molecular diagnostics and the need for nucleic acid integrity\n## Approved HPV diagnostic methods and sample requirements\n## Limitations of morphology and DNA-only screening","[{\"question\":\"What is the main goal of the study on cervical samples?\",\"answer\":\"To optimize an ethanol-based fixative that preserves DNA and RNA from cervical specimens stored under ambient conditions for reliable molecular and cytological testing.\"},{\"question\":\"Which experimental factors were optimized using the central composite design?\",\"answer\":\"pH, Sorbitol concentration, and PEG concentration, assessed for their effects on nucleic acid preservation quality.\"},{\"question\":\"How did the optimized fixative perform compared with the commercial PreservCyt solution?\",\"answer\":\"It significantly improved DNA and RNA preservation, showing high DNA integrity with successful PCR amplification (including HPV-18 oncogene targeting) and clear rRNA bands with lower qRT-PCR Ct values.\"}]","Optimizing an ethanol-based fixative for enhanced nucleic acid preservation in cervical samples using a central composite design approach | PDF",1790129649]