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Related Concept Videos

Fixation and Sectioning01:03

Fixation and Sectioning

Two basic types of preparation are used to visualize specimens with a light microscope: wet mounts and fixed specimens.
The simplest type of preparation is the wet mount, in which the specimen is placed in a drop of liquid on the slide. A liquid specimen can be directly deposited on the slide using a dropper. Solid specimens, such as skin scraping, can be placed on the slide before adding a drop of liquid to prepare the wet mount. Sometimes the liquid is simply water, but stains are often added...

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Protocol for HER2 FISH Using a Non-cross-linking, Formalin-free Tissue Fixative to Combine Advantages of Cryo-preservation and Formalin Fixation
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Formaldehyde substitute fixatives: effects on nucleic acid preservation.

Cathy B Moelans1, Daphne Oostenrijk, Michiel J Moons

  • 1Department of Pathology, University Medical Center Utrecht, Utrecht, The Netherlands. cmoelans@umcutrecht.nl

Journal of Clinical Pathology
|July 1, 2011
PubMed
Summary

New alcohol-based non-crosslinking fixatives (FineFIX, RCL2) offer superior DNA and RNA yield and quality for molecular pathology compared to traditional formalin. This enhances diagnostic applicability and may reduce required biopsy size.

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Area of Science:

  • Molecular Pathology
  • Surgical Pathology
  • Biomolecular Analysis

Background:

  • Formalin-fixed paraffin-embedded (FFPE) tissues are crucial for molecular assays alongside histopathology.
  • Standard formalin fixative is carcinogenic and its crosslinking properties degrade nucleic acid quality.
  • There is a need for safer, high-quality fixatives in molecular pathology.

Purpose of the Study:

  • To evaluate the suitability of novel alcohol-based fixatives (F-Solv, FineFIX, RCL2) for molecular pathology.
  • To compare these fixatives against neutral buffered formalin (NBF) for DNA and RNA extraction and analysis.
  • To assess the impact of crosslinking versus non-crosslinking properties on molecular assay performance.

Main Methods:

  • Three alcohol-based fixatives (F-Solv, FineFIX, RCL2) and NBF were tested.
  • Molecular assays included: size ladder PCR, epidermal growth factor receptor (EGFR) sequencing, microsatellite instability (MSI) analysis, chromogenic in situ hybridisation (CISH), fluorescence in situ hybridisation (FISH), and quantitative PCR (qPCR).
  • DNA and RNA yield, quality, and assay performance were evaluated.

Main Results:

  • Non-crosslinking alcohol-based fixatives (FineFIX, RCL2) yielded higher DNA quantity and quality than NBF and F-Solv.
  • PCR amplicons were larger with non-crosslinking fixatives (400 bp) compared to crosslinking ones (300 bp).
  • FineFIX and RCL2 showed superior RNA yield and quality, and lower qPCR Ct values; all fixatives were suitable for MSI and EGFR analysis, and most for CISH/FISH.

Conclusions:

  • Alcohol-based non-crosslinking fixatives significantly outperform crosslinking fixatives for DNA/RNA yield, quality, and molecular diagnostic applications.
  • These improved fixatives may enable molecular studies with less starting material, increasing biopsy utility.
  • FineFIX and RCL2 represent promising alternatives to formalin for routine molecular pathology.