Next-generation sequencing (NGS) of nucleic acids extracted from formalin-fixed, paraffin-embedded (FFPE) samples is becoming increasingly significant in clinical research. However, issues associated with using FFPE samples for downstream analysis persist.
This article discusses how collaborating with a seasoned team of experts who have access to a large biorepository of meticulously gathered and preserved tissue specimens can be critical to overcoming such challenges.
Formalin-fixed, paraffin-embedded samples, and next-generation sequencing
For decades, formalin-fixing paraffin-embedding has been the preferred method for preserving biological samples for diagnostic research. Formalin is relatively inexpensive and can be used to preserve a wide range of tissue samples using a single technique.1
Long-term storage of FFPE samples is more cost-effective than freezing since it requires less maintenance, labor costs, and space.2
The large number of FFPE tissue archives around the world, along with their clinical data, serve as an essential biobank for prospective, retrospective, and translational clinical studies.
NGS is currently widely used in clinical research to detect molecular changes for prognostic, diagnostic, predictive investigations, and precision medicine.
NGS can evaluate the genomes, epigenomes, and transcriptomes of small samples and has been regarded as a useful technology for studying molecules derived from FFPE tissue blocks. However, nucleic acids recovered from FFPE samples are fragmented and chemically altered during the fixation process.3
As a result, it is critical to identify and address the different challenges faced during the selection, usage, and processing of FFPE tissues for downstream applications.
The successful clinical implementation of FFPE-derived NGS requires an understanding and effective management of the limitations associated with FFPE samples, as well as optimized library preparation.
The need for an experienced team and provider when working with FFPE samples
To analyze the problems researchers face while working with FFPE samples, a study led by Chanel Smith, Director of Laboratory Operations at BioIVT, looked into the predictive value of certain FFPE sample properties for the success of future analyses.
The study, which lasted three years at BioIVT, involved evaluating over 2000 molecular assays. These assays used nucleic acids isolated from FFPE tissue samples, which differed in collection methods and fixation times.4
They investigated whether the success of screening outcomes was connected to any of the following factors:
- The percentage of necrosis – The percentage of the whole sample that was necrotic, as established by a board-certified pathologist.
- The time of fixation – The number of hours a sample was exposed to formalin and/or formaldehyde.
- Recovery type – Whether or not the samples were preserved according to known protocols.
- The age of the sample block – The time from FFPE fixation to nucleic acid isolation.
The samples that produced results were considered successful, whereas the samples unable to generate conclusive results were deemed unsuccessful.4
The study discovered that none of these individual characteristics independently predicted the success of downstream applications, including NGS. Instead, the researchers found that working with an experienced team like BioIVT is critical.
Such teams can provide precise sample selection by considering parameters such as refined sample diagnosis, sample quantity, and tissue composition, as well as tailoring the nucleic acid isolation process to the block type and downstream analysis.
This collaborative approach increases the likelihood of obtaining successful outcomes from FFPE samples.4
Image Credit: Elpisterra/Shutterstock.com
BioIVT – A worldwide provider of biological specimens, in vitro products and services
BioIVT is a global leader in high-quality biological samples, offering a cutting-edge variety of value-added services.
BiolVT’s professional staff, with years of expertise and supporting data, knows how specimen collection and preservation affect downstream applications.
For over 20 years, BiolVT has used the same rigorous standards to gather samples and build its ASTERAND® Human Tissue Repository, which contains over 300,000 high-quality, precisely characterized, validated tissues and related biofluids that are available for in-house screening.
Focus on rigorous quality control
All of BiolVT’s tissue samples have been examined independently by board-certified pathologists, collected under IRB-approved techniques, and de-identified before banking and distribution.
BioIVT is accredited by the College of American Pathologists (CAP). All samples in its ASTERAND® Elite Human Tissue Repository are accompanied by extensive clinical data that has undergone independent, rigorous, and standardized review.
Each tissue sample contains up to 200 matched clinical data points, as well as histology and socio-demographic data. Expert pathologists also checked all paired clinical data points for reliability and thoroughness.
To satisfy the current requirements of biomedical research, BiolVT’s standardized collection approach includes both sick and normal tissues from each donor. BiolVT also isolates tumor cells, PBMCs, epithelial cells, and related fibroblasts from other donors.
Elevating your NGS research with BiolVT’s mutation analysis experience
BioIVT has over ten years of expertise managing mutation screening projects using NGS, PCR, IHC, and FISH.
Every month, over 800 specimens are screened in-house using NGS, and BiolVT now has 10,000+ samples with mutation data in its archive, including specific point mutations, fusions, copy number variations (CNV), and both presence and absence mutations.
BiolVT’s annual data analysis provides information on the frequency of mutations seen in clinical environments as well as within its biobank.
Build your confidence in identifying the cohorts you require with BiolVT’s superior NGS screening capabilities. Its wide library of NGS pre-screened samples and comprehensive quality control data will help you identify the exact samples you need.
BiolVT’s inventory management knowledge, together with a group of professional molecular biologists adept at sampling and library preparation, as well as in-house biostatisticians, all work collectively to precisely validate and meet your unique cohort requirements.
BiolVT also provides customized biomarker analysis for certain cohorts, as well as tailored screening capabilities designed to align with your project’s unique requirements.
BioIVT’s extensive library of pre-screened cohorts and biospecimens can help expedite research workflow and reduce study timelines. Direct access to BiolVT’s in-house experts also ensures the feasibility, delivery, and timeliness of any study, hence increasing the efficacy of the clinical research.
Conclusion
BioIVT contributes significantly to the advancement of scientific knowledge of disease mechanisms and pharmacokinetics, as well as the identification of new therapeutic targets to accelerate medical research.
BioIVT is your trusted partner in elevating your NGS clinical research, thanks to its unique and vast biorepository of readily available specimens, as well as years of FFPE tissue preservation and molecular analysis experience.
References and further reading
- Mathieson, W. and Thomas, G. 2019. Using FFPE tissue in genomic analyses: advantages, disadvantages and the role of biospecimen science. Current Pathobiology Reports. 7, pp.35-40.
- Kokkat, T.J., Patel, M.S., McGarvey, D., LiVolsi, V.A. and Baloch, ZW 2013. Archived formalin-fixed paraffin-embedded (FFPE) blocks: a valuable underexploited resource for extraction of DNA, RNA, and protein. Biopreservation and biobanking. 11(2), pp.101-106.
- Hedegaard, J., Thorsen, K., Lund, M.K., Hein, A.M.K., Hamilton-Dutoit, S.J., Vang, S., Nordentoft, I., Birkenkamp-Demtröder, K., Kruhøffer, M., Hager, H. and Knudsen, B. 2014. Next-generation sequencing of RNA and DNA isolated from paired fresh-frozen and formalin-fixed paraffin-embedded samples of human cancer and normal tissue. PloS one. 9(5), p.e98187.
- Chanel Smith MSA, Heather Manza, MS, Aaron P. Moskalik, PhD, Cathie G. Miller, PhD and Shannon Richey. 2019. Key Determinants for Successful Formalin Fixed Paraffin Embedded Tissue Sample Prep for Downstream Applications. BioIVT, Detroit, MI, USA. Also presented at Molecular Med Tri-Con 2019.
About BioIVT
BioIVT, formerly BioreclamationIVT, is a leading global provider of high-quality biological specimens and value-added services. We specialize in control and disease state samples including human and animal tissues, cell products, blood and other biofluids. Our unmatched portfolio of clinical specimens directly supports precision medicine research and the effort to improve patient outcomes by coupling comprehensive clinical data with donor samples.
Our Research Services team works collaboratively with clients to provide in vitro hepatic modeling solutions. And as the world’s premier supplier of ADME-Tox model systems, including hepatocytes and subcellular fractions, BioIVT enables scientists to better understand the pharmacokinetics and drug metabolism of newly discovered compounds and the effects on disease processes. By combining our technical expertise, exceptional customer service, and unparalleled access to biological specimens, BioIVT serves the research community as a trusted partner in ELEVATING SCIENCE®.
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