Fresh frozen tissue is a method for preserving biological samples, capturing their original state at collection. This technique rapidly freezes tissue specimens, halting cellular processes and molecular degradation. This preservation allows scientists and medical professionals to study the tissue’s biological properties with minimal alteration. These samples are used in various scientific and medical applications, from understanding disease to developing new diagnostic tools.
Understanding Fresh Frozen Tissue
Fresh frozen tissue differs from other preservation methods, such as formalin-fixed, paraffin-embedded (FFPE) tissue, by avoiding chemical fixatives. Instead, it undergoes rapid freezing. This method maintains the tissue’s original biological properties, including the integrity of its nucleic acids (DNA and RNA) and proteins.
Rapid freezing minimizes ice crystal formation, which can damage cellular structures and alter molecular composition. Preserving these components in their native state provides a more accurate representation of the living biological environment. This makes fresh frozen tissue a preferred choice for analyses requiring pristine molecular information, unlike FFPE tissue where chemical fixation can introduce modifications and degrade nucleic acids.
The Freezing and Preservation Process
The preparation of fresh frozen tissue begins immediately after collection to minimize degradation. Specimens are typically snap-frozen by submerging them directly into liquid nitrogen. This rapidly cools the tissue to extremely low temperatures, often below -180°C (-292°F), arresting all biological and chemical activity. Some protocols use isopentane chilled by liquid nitrogen for freezing.
Once snap-frozen, samples are transferred to ultra-low temperature storage units, such as specialized freezers at -80°C (-112°F) or liquid nitrogen vapor phase tanks for long-term preservation. Maintaining these ultra-low temperatures without interruption, known as the “cold chain,” is important to prevent thawing that could compromise sample integrity. The tissue must remain frozen during storage, transport, and analysis to prevent damaging ice crystals or enzyme activation, ensuring it remains close to its original living condition for future use.
Why Fresh Frozen Tissue is Indispensable
Fresh frozen tissue is indispensable in biomedical research and diagnostics due to its preserved molecular integrity. It is favored for genomic studies because it maintains DNA and RNA in their natural, intact forms. This allows for accurate sequencing, gene expression analyses, and other molecular biology techniques like RNA-Seq and microarray testing.
Fresh frozen tissue is also invaluable for proteomic analyses, as proteins are maintained in their native, functional conformations. Unlike chemically fixed tissues, fresh frozen samples allow for reliable mass spectrometry-based proteomic analyses, enabling researchers to identify and quantify proteins and study their interactions. The preserved activity of enzymes and other molecules in fresh frozen samples also makes them suitable for functional studies. These capabilities provide comprehensive insights into biological processes and disease mechanisms.
Ensuring Quality for Reliable Results
Maintaining the quality of fresh frozen tissue samples requires strict adherence to established protocols. Rapid processing, ideally within 30 to 60 minutes of surgical excision, is important to minimize degradation. During collection, transport, and storage, samples must be kept consistently at ultra-low temperatures, such as -80°C or in liquid nitrogen vapor phase, to prevent thawing or temperature fluctuations that could compromise their integrity.
Quality control measures verify sample suitability, including assessing RNA integrity using metrics like the RNA Integrity Number (RIN). Histological evaluation, such as hematoxylin and eosin (H&E) staining, is also performed to confirm diagnosis and assess tumor content in cancer samples. These steps minimize issues like ice crystal formation or enzymatic degradation, ensuring samples yield accurate and reliable results for downstream analyses.