The process of formalin-fixed paraffin-embedding (FFPE) emerged in the late 19th century as a valuable histological method for stabilizing and preserving the morphology and cellular details of tissue samples. In 2017 the human tissues segment was estimated to account for the second largest share of the global biobanking market by sample type. This is attributable in large part to the expanded use of these tissues as molecular biomarkers of disease, primarily in oncology, using immunohistochemical and genetic-based assay systems.
Given the significant potential of FFPE tissues, this material must be collected, documented and preserved properly in order to gain its full potential when using it for protein and nucleic acid-based analyses. In this blog we’ll review the importance of standardization in establishing a FFPE tissue block collection.
Using FFPE tissues reliably for protein and nucleic-acid based assays, including immunohistochemistry techniques, expression analysis, and next gen sequencing is problematic because of the diversity found in the clinic and hospital environments when harvesting these tissues for their primary use in histology and cytology. Well defined, standardized procedures for documenting, collecting and maximizing the quality of the tissue samples must be developed across hospitals and clinics with the knowledge that in addition to their initial primary use (histology and cytology) these samples will also be used for an ever expanding range of genetic and protein analyses requiring the isolation of RNA, DNA and proteins.
The first steps in standardizing sample collection require documentation on sample obtainment (including pre- and post-sample removal, detailing storage conditions immediately after removal until retrieved for processing). For instance:
- Was the sample obtained by needle biopsy, or removal of a larger portion such as tumor resection, full or partial organ/tissue removal?
- For both sample removal techniques, how long was the tissue stored, at what temperature, were any enzyme inhibitors added to slow/stop nucleic acid degradation?
- How was the larger tissue specimen processed prior to fixation?
- How much time elapsed between tissue removal and fixation?
For the fixation process, tissue thickness is key to perfusion of the fixative as incorrect thickness can cause under- as well as over- fixation. Next, the fixed tissue is embedded in a three step process; each step in the process is critical (dehydration, clearing, impregnation). At the final step, the tissue is embedded in a paraffin block and readied for sectioning. Throughout the process, it is important to document the process and detail information on all reagents used, including lot numbers and expiration dates. Using optimized methods for FFPE tissue block creation can improve the isolation of RNA and DNA for use in genetic assays, and will require a greater focus on establishing standards beginning from sample collection to methods for nucleic acid isolation.
Standardization plays a pivotal role across all functions of a biobank. By incorporating standardized processes into your biobanking operations you can increase efficiencies and reduce waste. It’s key to ensure that all collection sites involved have an agreed upon and easily understandable standard protocol that indicates what information is collected, how it is collected, and how the sample is collected/labeled. It’s equally important to ensure that the material collection process aligns with the data analysis parameters and downstream research goals.
Many study designers often rely on each site to determine how they will record data. This can place a heavy burden on the teams so it’s important to make collection as easy as possible. These sites are often not dedicated to one study, but rather are participating in this collection because of the patient’s enrollment. They may be participating in many studies at the same time as an addition to their normal job routine.
While there are many benefits of standardization, it is not always easy to implement. Information on best practices are often scattered across multiple channels. It can also be challenging to find agreement among your teams based on their interpretation of those best practices or their past experiences. It’s important to involve site-leads to gain their buy-in on the proposed process and employ good communication. A few additional tools that can help support standardization include:
- Training plans
- Bulleted protocols
- Pre-printed labels
- Pre-produced kits
- Approved consumables
When developing the standard, consideration should be given to:
- The ultimate potential size of the collection (how big should the unique identifier/accession number be?)
- HIPAA regulation:
- How to format the unique identifier, simple randomization or complex embedded information?
- Who will maintain the connection between participant/patient and the sample?
- Under what use are the samples initially being collected? Will they require a 10 year waiting period?
- Which slides, labels, or cassettes will be used?
- Which techniques are used in sample collection?
- Providing kitted solutions, bulk supplies, or just pre-printed labels? Providing bulk supplies is often the best solution for dedicated sites as it allows for pathologists to manage their stocks while also allowing for uniformity across all collectors.
While it is often easier said than done to incorporate standardization when developing an FFPE block collection, the benefits to the downstream research are well worth the upfront effort. To learn more about how Fisher Clinical Services can help support your upcoming projects, contact us today.