Figure 1. NucleoSpin 96 DNA FFPE procedure. The protocol can be performed using centrifugation or vacuum processing and can be automated on common liquid-handling platforms.
Product overview
NucleoSpin 96 DNA FFPE
The NucleoSpin 96 DNA FFPE kit is designed for high-throughput DNA purification from formalin-fixed, paraffin-embedded samples. Paraffin is first removed from the sample using odorless, xylene-free Paraffin Dissolver, which enables effective lysis in a convenient, two-phase system. Samples are then digested by Proteinase K to solubilize fixed tissues and release DNA into solution. Subsequently, crosslinks are removed via heat incubation in a specially designed buffer, and large amounts of chaotropic salt and ethanol are added to the lysate, providing conditions suitable for binding of DNA to the silica membrane in the NucleoSpin DNA FFPE Binding Plate. The binding process is reversible and specific to nucleic acids. While DNA is kept on the silica membrane, contaminants are removed by washing with two different wash buffers. Pure genomic DNA is eluted under low ionic strength conditions in a slightly alkaline elution buffer.
- Isolation of DNA from FFPE samples in a high-throughput, 96-well plate format
- Blue-colored, xylene-free Paraffin Dissolver for easy paraffin removal
- Decrosslinking buffer to eliminate formalin crosslinking
- High-quality DNA for improved performance in downstream applications
- Suitable for manual and automated processing under centrifugation or vacuum
Procedure
Reliable DNA yield
Figure 2. Reliable DNA yield with NucleoSpin 96 DNA FFPE. DNA was purified from seven FFPE samples derived from human ileum and stomach and quantified using the Quantifiler Human DNA Quantification Kit. Average DNA yields of 0.3 µg and 1.7 µg were obtained from the ileum and stomach, respectively.
Consistent PCR performance
Figure 3. Consistent PCR performance with NucleoSpin 96 DNA FFPE. Genomic DNA was isolated from mouse liver, lung, muscle, and kidney FFPE tissues (8 samples each) with the NucleoSpin 96 DNA FFPE kit. Panel A. Analysis by qPCR. Purified DNA was quantified using Maxima SYBR Green qPCR Master Mix (amplicon size: 100 bp). The similar CT values observed for each sample type demonstrate the consistent performance of the kit in yielding DNA suitable for PCR amplification. Panel B. Analysis by gel electrophoresis. Agarose gel electrophoresis of PCR products from the 8 different samples of mouse liver genomic DNA indicated that DNA quality and yield were comparable for each.
At-a-glance
| Technology | Silica membrane |
| Format | 96-well plate |
| Processing | Manual or automated, vacuum or centrifugation |
| Starting material | ≤10 mg tissue ≤15 mg paraffin |
| Fragment size | 50 bp–approx. 5 kb |
| Typical yield | Depends on the amount and quality of the sample |
| A260/280 | Strongly depends on sample quality |
| Elution volume | 100 µl |
| Preparation time | ~60 min/plate (excl. lysis) |
| Binding capacity | 20 µg |
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