Fundación instituto valenciano de oncología (fivo)
A sustainable alternative to deep-freezing for DNA and plasma preservation.
DNA is a naturally stable molecule that is well suited to long-term storage. However, inadequate storage makes it susceptible to degradation processes that can compromise its quality—especially when it comes to meeting the demands of new and more sensitive molecular biology techniques.
As a result, despite its supposed stability, DNA is still routinely stored in ultra-low temperature freezers, occupying between 10% and 20% of available storage capacity in biobanks and laboratories.
Plasma, on the other hand, is a complex biological fluid with a high protein content and significant clinical relevance. It is particularly sensitive to freeze-thaw cycles and storage conditions, which can affect the stability of its components and compromise downstream analyses.
Objective
To assess the technical feasibility of 300K Solutions technology for the dry stabilization of plasma and DNA extracted from different primary samples. The study aims to compare, in parallel, the quality of lyophilized samples against those stored by conventional freezing, using standardized quality control metrics commonly employed in biobank settings. This validation will determine the suitability of lyophilization as an alternative storage method without compromising sample integrity or downstream analytical performance.
Materials and Methods
Samples used for the study
20 DNA samples
extracted from peripheral blood
20 DNA samples
extracted from FFPE
20 DNA samples
frozen at -20 ºC for up to 5 years
20 plasma samples
Procedure
The validation process is structured to enable a direct comparison between DNA samples stored through conventional freezing and those preserved via lyophilization using the S3 system developed by 300K Solutions. The procedure is standardized across all sample types to ensure consistency and scientific robustness.
The study includes four types of samples: high molecular weight genomic DNA extracted from peripheral blood, DNA extracted from formalin-fixed paraffin-embedded (FFPE) tissue, plasma for downstream circulating DNA (cfDNA) extraction, and previously frozen DNA obtained from whole blood.
For genomic DNA and FFPE-derived DNA, extraction is performed at the beginning of the study using validated methods. Upon extraction, quality control is conducted using electrophoresis and Nanodrop spectrophotometry to assess DNA purity and integrity. Each sample is then divided into two identical aliquots: one stored at -20ºC under standard biobank conditions and the other lyophilized using the S3 Sample Stabilization System (300K Solutions). In the case of frozen DNA, it is thawed before being either lyophilized or returned to frozen storage, depending on the assigned condition.
In plama samples, half of the aliquots are stored at −80 ºC, and the other half are lyophilized using 300K technology. After the storage period, cfDNA is extracted from both sets of samples, enabling a direct comparison of DNA yield and quality between lyophilized and frozen conditions.
Lyophilized samples are stored at room temperature in clearly labeled boxes. Frozen control samples remain stored at −20ºC (DNA) or −80ºC (plasma), following standard biobanking protocols. All samples are stored under their respective conditions for one and six months. At both time points, paired aliquots—lyophilized and frozen—from the same original sample are processed in parallel. DNA is either rehydrated from lyophilized pellets or thawed from frozen state before undergoing quality control analysis. This design allows intra-sample comparisons to assess the technical feasibility and performance of lyophilized storage relative to conventional freezing.
Quality Assurance
Ratio A260/280
DIN
DNA Integrity Number
PicoGreen
Quantification
PCR
Long PCR/amplification of the beta-2-microglobulin gene
Quality assessments are performed at two timepoints—one month and six months post-storage—for both lyophilized and frozen samples to evaluate purity, integrity, concentration, and long-fragment amplifiability
DNA extracted from fresh blood is analyzed using:
- Nanodrop spectrophotometry
- TapeStation electrophoresis
- Picogreen quantification
- Long-PCR
DNA from FFPE samples undergoes analysis with:
- Nanodrop
- TapeStation
- Picogreen
- PCR amplification of the beta-2-microglobulin gene, a standard quality marker for FFPE DNA.
DNA previously frozen is assessed with:
- Nanodrop
- TapeStation.
Circulating DNA extracted from plasma is evaluated using:
- TapeStation
Results
Coming soon…
Conclusion
Coming soon…
