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Published on: December 25, 2017
The use of FTA cards for preserving unfixed cytological material for high-throughput molecular analysis
Mauro Ajaj Saieg1, William R Geddie, Scott L Boerner
1Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada.
Insights
FTA Classic cards offer superior DNA integrity for molecular analysis compared to Elute cards, enabling high-throughput mutation screening from fine-needle aspiration samples.
Area of Science:
- Molecular Biology
- Genomics
- Biobanking
Background:
- High-throughput molecular technologies necessitate efficient cell and tissue specimen collection and storage.
- FTA cards offer a cryopreservation alternative for biobanking fresh, unfixed cells.
- This study evaluates DNA quality and integrity from two FTA card types and extraction protocols for fine-needle aspiration (FNA) samples.
Purpose of the Study:
- To compare DNA quality and integrity from FTA Classic and Elute cards.
- To assess DNA extraction protocols for FTA cards.
- To determine the feasibility of multiplex mutational screening using FNA samples stored on FTA cards.
Main Methods:
- Residual material from 42 FNA biopsies was collected on 21 FTA Classic and 21 FTA Elute cards.
- DNA was extracted using 'Classic' protocol for Classic cards and both 'Classic' and 'Elute' protocols for Elute cards.
- Polymerase chain reaction (PCR) for p53 and CARD11 genes was performed to evaluate DNA integrity.
Main Results:
- Successful p53 amplification rates were significantly higher with Classic cards (95.2%) compared to Elute cards (80.9% with Classic protocol, 28.5% with Elute protocol; P = .001).
- All samples yielded CARD11 amplification, indicating sufficient DNA for smaller targets.
- No significant differences in DNA concentration or purity (260/280 ratio) were observed between card types; multiplex MassARRAY identified a KRAS mutation in one sample.
Conclusions:
- High molecular weight DNA can be successfully extracted from FTA cards in adequate quantity and quality for high-throughput multiplex mutation assays.
- FTA Classic cards demonstrate superior DNA integrity for molecular applications compared to FTA Elute cards.
- FTA cards are suitable for preserving FNA samples for molecular analysis, including mutation screening.
Background:
Novel high-throughput molecular technologies have made the collection and storage of cells and small tissue specimens a critical issue. The FTA card provides an alternative to cryopreservation for biobanking fresh unfixed cells. The current study compared the quality and integrity of the DNA obtained from 2 types of FTA cards (Classic and Elute) using 2 different extraction protocols ("Classic" and "Elute") and assessed the feasibility of performing multiplex mutational screening using fine-needle aspiration (FNA) biopsy samples.
Methods:
Residual material from 42 FNA biopsies was collected in the cards (21 Classic and 21 Elute cards). DNA was extracted using the Classic protocol for Classic cards and both protocols for Elute cards. Polymerase chain reaction for p53 (1.5 kilobase) and CARD11 (500 base pair) was performed to assess DNA integrity.
Results:
Successful p53 amplification was achieved in 95.2% of the samples from the Classic cards and in 80.9% of the samples from the Elute cards using the Classic protocol and 28.5% using the Elute protocol (P = .001). All samples (both cards) could be amplified for CARD11. There was no significant difference in the DNA concentration or 260/280 purity ratio when the 2 types of cards were compared. Five samples were also successfully analyzed by multiplex MassARRAY spectrometry, with a mutation in KRAS found in 1 case.
Conclusions:
High molecular weight DNA was extracted from the cards in sufficient amounts and quality to perform high-throughput multiplex mutation assays. The results of the current study also suggest that FTA Classic cards preserve better DNA integrity for molecular applications compared with the FTA Elute cards.
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