FormaPure Total Performance and Data

Manual or Automated RNA, DNA or Total Nucleic Acid Extraction from FFPE Tissues

Nucleic acid extraction from formalin-fixed, paraffin-embedded (FFPE) tissue is challenging due to the nature of the tissue preparation. FormaPure reagent kits represent a single chemistry system designed and optimized for use in Next-Generation Sequencing (NGS) as well as other downstream applications including qPCR/ddPCR/PCR. Maximizing integrity, yield and purity from an FFPE sample is required to minimize the risk of losing important genetic information.

  • Consistent extraction of NGS compatible RNA and/or DNA from a single FFPE sample
  • Scalable single chemistry system for manual or automated usage
  • Extract higher integrity nucleic acids supporting improved sensitivity for NGS applications

FormaPure Total for RNA Extraction

NGS performance starts with RNA integrity. Fragment analysis shows FormaPure RNA reagent kits retains the best possible RNA integrity even with 10+ year old tissue blocks.

Genomics NGS Performance with FormaPure RNA

FormaPure RNA isolates RNA with superior integrity. Fragment analyses were performed with RNA isolated with FormaPure RNA (blue traces) and an alternative column based supplier’s method (orange traces) from four different FFPE sample types. All extractions were performed with one 10 µm curl. DV200 values are presented for each of the chromatograms. DV200 values represent the percentage of RNA fragments > 200 nucleotides.

Genomic DNA Isolation from FFPE

FormaPure DNA maintains detectability of low frequency variants without introducing extraction based experimental bias. DNA extracted using FormaPure DNA is suitable for the most stringent downstream applications including NGS.

FormaPure DNA and FormaPure Total provide:

  • Consistent and efficient extraction performance
  • Simple workflow with minimal hands-on time
  • Proven compatibility with NGS and other downstream applications
  • Flexibility to accommodate sample input variation

 

Next-Generation Sequencing Compatibility
Position Gene FormaPure DNA Samples with Variant Call (Max 6) FormaPure DNA Variant Freq (%) Expected Allelic Freq (%)
140453136 BRAF 6 12.2 10.5
55599321 KIT 6 9.2 10
55242464 EGFRΔ 6 1.8 2
55259515 EGFR 6 3.1 3
55241707 EGFR 6 25 24.5
25398281 KRAS 6 13.7 15
25398284 KRAS 6 5.3 6
115256530 NRAS 6 17.2 12.5
178952085 PIK3CA 6 17.5 17.5
178936091 PIK3CA 6 9.1 9

 

Nucleic Acid Yields

Determined by the following variables:

Tissue Type | Tissue Size | Tumor Type | Fixation Method | Paraffin Embedding Method | Reagents | Storage Conditions | Tissue Input Amount | Extraction Chemistry | Age of Tissue Block

Tissue Type Block Age (years) RNA Yield (ng) DNA Yield (ng)
Breast >3 866 368
Large Intestine >10 576 410
Liver >10 580 1134
Lung >10 376 951

 

Visual Workflow

Genomics Products Sequencing RNA and DNA Tubes graphic

  1. Deparaffinize FFPE tissue
  2. Lyse FFPE tissue and decrosslink RNA
  3. Transfer half of the lysate for RNA workflow
  4. Bind RNA to magnetic beads
  5. Separate magnetic beads from contaminants
  6. Wash magnetic beads with 80% ethanol to remove contaminants
  7. Treat samples with DNase I
  8. Rebind RNA to magnetic beads with RBA
  9. Separate magnetic beads from contaminants
  10. Wash magnetic beads with 70% ethanol to remove contaminants
  11. Elute RNA from magnetic beads
  12. Process remaining lysate for DNA workflow
  13. Decrosslink DNA
  14. Transfer remaining lysate for DNA workflow
  15. Treat samples with RNase A
  16. Bind DNA to magnetic beads
  17. Separate magnetic beads from contaminants
  18. Wash magnetic beads with 80% ethanol to remove contaminants
  19. Elute DNA from magnetic beads
  20. Transfer eluted RNA and DNA to a storage plate

 

Automation vs. Manual Timing

FormaPure Total, RNA and DNA are flexible set of extraction kits suitable for manual or automated workflows.

  • Scalable based on throughput
  • Quick transition with ready-to-implement methods
  • Knowledgeable support for reagents, automation and methods from a single vendor

Estimated hands-on-time and total time in hours, required to perform 8, 24, 48 and 96 FormaPure Total, RNA or DNA nucleic acid extractions. The methods can be performed either manually or automated on a liquid handling system. Data represented in this table is based on a Biomek i7 Hybrid. Difference in time between manual and automation is indicated. NR=Not recommended.

 

FormaPure Total FormaPure DNA FormaPure RNA
Manual Automated Manual Automated Manual Automated
Batch Size 8 Hands-on Time 3.5 0.5 1 0.5 2 0.5
Total Time 6.5 6 3.5 5.25 4.5 5
24 Hands-on Time 4 0.5 1.5 0.5 2.5 0.5
Total Time 7 6.25 4 5.5 5 5
48 Hands-on Time NR 0.5 NR 0.5 NR 0.5
Total Time NR 6.5 NR 5.5 NR 5.5
96 Hands-on Time NR 0.5 NR 0.5 NR 0.5
Total Time NR 6.75 NR 5.75 NR 5.5

 

Products: FormaPure Reagent Kits

FormaPure reagent kits are available in multiple kit sizes and extraction types based on your application and throughput needs. These products are enabled for purchase or you can request a quote. Please see the specific products and part numbers below.

PART # NAME PREPS
C16675 FormaPure Total 50
C16676 FormaPure Total 96
PART # NAME PREPS
C19157 FormaPure RNA 50
C19158 FormaPure RNA 96
PART # NAME PREPS
B89230 FormaPure DNA 50
B89231 FormaPure DNA 96

  

 

Beckman Coulter makes no warranties of any kind whatsoever express or implied, with respect to this protocol, including but not limited to warranties of fitness for a particular purpose or merchantability or that the protocol is non-infringing. All warranties are expressly disclaimed. Your use of the method is solely at your own risk, without recourse to Beckman Coulter. Not intended or validated for use in the diagnosis of disease or other conditions. This protocol is for demonstration only, and is not validated by Beckman Coulter.

© 2019 Beckman Coulter, Inc. All rights reserved. Beckman Coulter, the stylized logo and the Beckman Coulter product and service marks mentioned herein are trademarks or registered trademarks of Beckman Coulter, Inc. in the United States and other counters. All other trademarks are the property of their respective owners.



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