Effectiveness of Xylene and Mineral Oil in DNA Extraction from Formalin-fixed Paraffin-embedded Diffuse Astrocytic Tumor

Edward Jaya Hadi, Hasrayati Agustina, Hermin Aminah Usman


Formalin-fixed paraffin-embedded (FFPE) tissue is an important source of material for molecular analysis. In the anatomical pathology field, molecular testing is needed in certain tumors, such as astrocytic tumors, to confirm the diagnosis. Extracting DNA from FFPE material is still challenging. The first important step in the extraction process is deparaffinization. This study aims to compare two types of deparaffinization methods. The first method used xylene, and the second one used mineral oil. The results of this study can be used to develop a reliable protocol for DNA extraction from FFPE tissue. DNA from 28 FFPE diffuse astrocytic tumor tissue blocks was extracted. The quantity and A260/A280 ratio of the DNA was measured by spectrophotometer. PCR assays were performed to assess the suitability of extracted DNA for molecular analysis. The results showed that the xylene group has significantly higher DNA concentrations than the mineral oil group (p<0.01). In both groups, average and median A260/A280 scores were between 1.8-2.0. In PCR assay, both groups show a similar result (27 of 28 samples were successfully amplified). In conclusion, xylene and mineral oil can be used effectively as deparaffinization agents. Both methods generated good quality extracted DNA. The advantage of mineral oil is that it is non-toxic and has shorter hands-on time. Xylene is preferable for a small-sized sample because it produces a higher DNA yield.


Deoxyribose nucleic acid extraction; formalin-fixed paraffin-embedded; mineral oil; xylene

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Bass, B. P., Engel, K. B., Greytak, S. R., & Moore, H. M. (2014). A review of preanalytical factors affecting molecular, protein, and morphological analysis of formalin-fixed, paraffin-embedded (FFPE) tissue: how well do you know your FFPE specimen? Arch Pathol Lab Med, 138(11), 1520-1530. doi:10.5858/arpa.2013-0691-RA

Christians, A., Adel-Horowski, A., Banan, R., Lehmann, U., Bartels, S., Behling, F., . . . Hartmann, C. (2019). The prognostic role of IDH mutations in homogeneously treated patients with anaplastic astrocytomas and glioblastomas. Acta Neuropathologica Communications, 7(1), 156. doi:10.1186/s40478-019-0817-0

Coura, R., Prolla, J. C., Meurer, L., & Ashton-Prolla, P. (2005). An alternative protocol for DNA extraction from formalin fixed and paraffin wax embedded tissue. J Clin Pathol, 58(8), 894-895. doi:10.1136/jcp.2004.021352

Do, H., & Dobrovic, A. (2015). Sequence Artifacts in DNA from Formalin-Fixed Tissues: Causes and Strategies for Minimization. Clinical Chemistry, 61(1), 64-71. doi:10.1373/clinchem.2014.223040

Donczo, B., & Guttman, A. (2018). Biomedical analysis of formalin-fixed, paraffin-embedded tissue samples: The Holy Grail for molecular diagnostics. J Pharm Biomed Anal, 155, 125-134. doi:10.1016/j.jpba.2018.03.065

Farrugia, A., Keyser, C., & Ludes, B. (2010). Efficiency evaluation of a DNA extraction and purification protocol on archival formalin-fixed and paraffin-embedded tissue. Forensic Sci Int, 194(1-3), e25-28. doi:10.1016/j.forsciint.2009.09.004

Gilbert, M. T. P., Haselkorn, T., Bunce, M., Sanchez, J. J., Lucas, S. B., Jewell, L. D., . . . Worobey, M. (2007). The Isolation of Nucleic Acids from Fixed, Paraffin-Embedded Tissues–Which Methods Are Useful When? PLoS One, 2(6), e537. doi:10.1371/journal.pone.0000537

Glasel, J. A. (1995). Validity of nucleic acid purities monitored by 260nm/280nm absorbance ratios. Biotechniques, 18(1), 62-63.

Greytak, S. R., Engel, K. B., Bass, B. P., & Moore, H. M. (2015). Accuracy of Molecular Data Generated with FFPE Biospecimens: Lessons from the Literature. Cancer Res, 75(8), 1541-1547. doi:10.1158/0008-5472.Can-14-2378

Kandyala, R., Raghavendra, S. P. C., & Rajasekharan, S. T. (2010). Xylene: An overview of its health hazards and preventive measures. Journal of oral and maxillofacial pathology : JOMFP, 14(1), 1-5. doi:10.4103/0973-029X.64299

Louis, D. N., Perry, A., Wesseling, P., Brat, D. J., Cree, I. A., Figarella-Branger, D., . . . Ellison, D. W. (2021). The 2021 WHO Classification of Tumors of the Central Nervous System: a summary. Neuro-Oncology, 23(8), 1231-1251. doi:10.1093/neuonc/noab106

Lucena-Aguilar, G., Sánchez-López, A. M., Barberán-Aceituno, C., Carrillo-Ávila, J. A., López-Guerrero, J. A., & Aguilar-Quesada, R. (2016). DNA Source Selection for Downstream Applications Based on DNA Quality Indicators Analysis. Biopreserv Biobank, 14(4), 264-270. doi:10.1089/bio.2015.0064

Mariño-Enríquez, A., & Bovée, J. V. M. G. (2016). Molecular Pathogenesis and Diagnostic, Prognostic and Predictive Molecular Markers in Sarcoma. Surgical Pathology Clinics, 9(3), 457-473. doi:https://doi.org/10.1016/j.path.2016.04.009

Premalatha, B. R., Patil, S., Rao, R. S., & Indu, M. (2013). Mineral oil--a biofriendly substitute for xylene in deparaffinization: a novel method. J Contemp Dent Pract, 14(2), 281-286. doi:10.5005/jp-journals-10024-1314

Rajan, S., & Malathi, N. (2014). Health hazards of xylene: a literature review. J Clin Diagn Res, 8(2), 271-274. doi:10.7860/jcdr/2014/7544.4079

Rawlings, A. V., & Lombard, K. J. (2012). A review on the extensive skin benefits of mineral oil. International Journal of Cosmetic Science, 34(6), 511-518. doi:https://doi.org/10.1111/j.1468-2494.2012.00752.x

Sarnecka, A. K., Nawrat, D., Piwowar, M., Ligęza, J., Swadźba, J., & Wójcik, P. (2019). DNA extraction from FFPE tissue samples - a comparison of three procedures. Contemp Oncol (Pozn), 23(1), 52-58. doi:10.5114/wo.2019.83875

Sedlackova, T., Repiska, G., Celec, P., Szemes, T., & Minarik, G. (2013). Fragmentation of DNA affects the accuracy of the DNA quantitation by the commonly used methods. Biol Proced Online, 15(1), 5. doi:10.1186/1480-9222-15-5

Sengüven, B., Baris, E., Oygur, T., & Berktas, M. (2014). Comparison of methods for the extraction of DNA from formalin-fixed, paraffin-embedded archival tissues. Int J Med Sci, 11(5), 494-499. doi:10.7150/ijms.8842

Srinivasan, M., Sedmak, D., & Jewell, S. (2002). Effect of fixatives and tissue processing on the content and integrity of nucleic acids. Am J Pathol, 161(6), 1961-1971. doi:10.1016/s0002-9440(10)64472-0

Weller, M., van den Bent, M., Preusser, M., Le Rhun, E., Tonn, J. C., Minniti, G., . . . Wick, W. (2021). EANO guidelines on the diagnosis and treatment of diffuse gliomas of adulthood. Nature Reviews Clinical Oncology, 18(3), 170-186. doi:10.1038/s41571-020-00447-z

Yi, Q. Q., Yang, R., Shi, J. F., Zeng, N. Y., Liang, D. Y., Sha, S., & Chang, Q. (2020). Effect of preservation time of formalin-fixed paraffin-embedded tissues on extractable DNA and RNA quantity. J Int Med Res, 48(6), 300060520931259. doi:10.1177/0300060520931259


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