Tissue Fixation with a Formic Acid-Deprived Formalin Better Preserves DNA Integrity over Time.

IF 3.5 4区 医学 Q3 CELL BIOLOGY Pathobiology Pub Date : 2023-01-01 DOI:10.1159/000525523
Enrico Berrino, Laura Annaratone, Paolo Detillo, Dora Grassini, Alberto Bragoni, Anna Sapino, Benedetta Bussolati, Giovanni Bussolati, Caterina Marchiò
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引用次数: 3

Abstract

Introduction: Optimization of pre-analytic procedures and tissue processing is a basic requirement for reliable and reproducible data to be obtained. Tissue fixation in formalin represents the extensively favored method for surgical tissue specimen processing in diagnostic pathology; however, formalin fixation exerts a blasting effect on DNA and RNA.

Methods: A formic acid-deprived formaldehyde solution was prepared by removing acids with an ion-exchange basic resin and the concentrated, acid-deprived formaldehyde (ADF) solution was employed to prepare a 4% ADF solution in 0.1 M phosphate buffer, pH 7.2-7.4. Human (n = 27) and mouse (n = 20) tissues were fixed in parallel and similar conditions in either ADF or neutral buffered formalin (NBF). DNAs and RNAs were extracted, and fragmentation analyses were performed.

Results: Besides no significant differences in terms of extraction yield and absorbance ratio, ADF fixation reduced DNA fragmentation, i.e., the largest fragments (>5,000 bp) were significantly more prevalent in the DNAs purified from ADF-fixed tissues (p < 0.001 in both cohorts). Moreover, we observed that DNA preservation is more stable in ADF-fixed tissue compared to NBF-fixed tissues.

Conclusion: Although DNA fragmentation in FFPE tissues is a multifactor process, we showed that the removal of formic acid is responsible for a significant improvement in DNA preservation.

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组织固定与甲酸剥夺福尔马林更好地保存DNA完整性随着时间的推移。
分析前程序和组织处理的优化是获得可靠和可重复数据的基本要求。在福尔马林组织固定代表广泛青睐的方法,外科组织标本处理诊断病理学;然而,福尔马林固定对DNA和RNA产生爆炸效应。方法:采用离子交换碱性树脂脱酸法制备甲酸去甲醛溶液,将浓缩的去酸甲醛(ADF)溶液在0.1 M磷酸盐缓冲液中配制4% ADF溶液,pH值为7.2 ~ 7.4。人(n = 27)和小鼠(n = 20)的组织在平行和类似的条件下固定在ADF或中性缓冲福尔马林(NBF)中。提取dna和rna,进行片段化分析。结果:除了提取率和吸光比方面没有显著差异外,ADF固定减少了DNA片段化,即从ADF固定组织中纯化的DNA中,最大片段(> 5000 bp)明显更普遍(两组的p < 0.001)。此外,我们观察到与nbf固定组织相比,adf固定组织中的DNA保存更稳定。结论:尽管FFPE组织中的DNA断裂是一个多因素过程,但我们发现甲酸的去除对DNA保存有显著改善。
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来源期刊
Pathobiology
Pathobiology 医学-病理学
CiteScore
8.50
自引率
0.00%
发文量
47
审稿时长
>12 weeks
期刊介绍: ''Pathobiology'' offers a valuable platform for the publication of high-quality original research into the mechanisms underlying human disease. Aiming to serve as a bridge between basic biomedical research and clinical medicine, the journal welcomes articles from scientific areas such as pathology, oncology, anatomy, virology, internal medicine, surgery, cell and molecular biology, and immunology. Published bimonthly, ''Pathobiology'' features original research papers and reviews on translational research. The journal offers the possibility to publish proceedings of meetings dedicated to one particular topic.
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