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Background <p>High-quality RNA isolation from tissues is crucial for transcriptomic analysis. The tissue disruption influences the RNA quality. The aim of this study was to compare different methods of tissue homogenization.</p> Methods <p>Three homogenization methods were used (mortar and pestle, ball mill, and tissue homogenizer) to disrupt head and neck cancerous tissues, healthy tissues from free margin of head and neck cancer patients, and breast skin from breast cancer patients. The comparison of isolated RNA quantity and quality by measuring the concentration and absorbance, RIN, and Ct values of reference genes were performed.</p> Results <p>RNA isolated after tissue homogenizer usage has the highest 260/230 ratio (<i>p</i> = 0.02) and concentration (<i>p</i> = 0.02) also RIN values tend to be highest across all studied tissues. There are no significant differences between Ct values across all tissues processed with different homogenization methods; however, the Ct values of <i>GAPDH</i> and <i>S18</i> are negatively correlated with RIN number (<i>p</i> = 0.002, <i>p</i> = 0.003).</p> Conclusion <p>The tissue homogenizer is the most suitable for obtaining high-quality RNA across all examined tissues, which is essential in various RNA-based analyses. <i>GAPDH</i> and <i>S18</i> Ct can indicate the RNA quality measured by RIN values.</p>

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Impact of soft tissue homogenization methods on RNA quality

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年4月25日

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  • Julia Ostapowicz,
  • Bartosz Maćkowiak,
  • Kamila Ostrowska,
  • Barbara Kaczmarek,
  • Natalia Pietras,
  • Dawid Frąckowiak,
  • Magdalena Fundowicz,
  • Katarzyna Kulcenty,
  • Wojciech Golusiński,
  • Wiktoria Suchorska

摘要

Background

High-quality RNA isolation from tissues is crucial for transcriptomic analysis. The tissue disruption influences the RNA quality. The aim of this study was to compare different methods of tissue homogenization.

Methods

Three homogenization methods were used (mortar and pestle, ball mill, and tissue homogenizer) to disrupt head and neck cancerous tissues, healthy tissues from free margin of head and neck cancer patients, and breast skin from breast cancer patients. The comparison of isolated RNA quantity and quality by measuring the concentration and absorbance, RIN, and Ct values of reference genes were performed.

Results

RNA isolated after tissue homogenizer usage has the highest 260/230 ratio (p = 0.02) and concentration (p = 0.02) also RIN values tend to be highest across all studied tissues. There are no significant differences between Ct values across all tissues processed with different homogenization methods; however, the Ct values of GAPDH and S18 are negatively correlated with RIN number (p = 0.002, p = 0.003).

Conclusion

The tissue homogenizer is the most suitable for obtaining high-quality RNA across all examined tissues, which is essential in various RNA-based analyses. GAPDH and S18 Ct can indicate the RNA quality measured by RIN values.