What are FFPE Samples

What is Formalin Fixed Paraffin Embedded Tissue?

Formalin fixed paraffin embedded or FFPE tissues are valuable for both therapeutic applications and research. FFPE is a specific technique used to prepare and preserve tissue specimens utilized in research, examination, diagnostics, and drug development. Tissues are first collected from both diseased and non-diseased donors. The tissue specimen is first preserved through a process called formalin fixing. This step helps to preserve the vital structures and protein within the tissue. It is then embedded into a paraffin wax block and sliced into the required slices, mounted on a microscopic slide, and examined.

 

The FFPE Process

The process starts by a specimen being selected and then excised from a donor or patient. Samples can also be obtained from other animals such as snakes, mice, or many others. After excision, the tissue is immersed for approximately eighteen to twenty-four hours in a 10% neutral buffered formalin. The tissue is then dehydrated using increasing concentrates of ethanol. Next, the tissue is embedded into paraffin to become FFPE blocks. The methods utilized are dependent on the requirements of the researcher or physician who is requesting the FFPE samples. Specifications about how the issue is cut, size, and purpose of the tissue are all important. Once the procedure is complete a certified pathologist will  evaluate the quality of the specimen.

 

Storage of FFPE Tissue

FFPE samples can be stored in hospitals, biobanks, and research centers. Storage facilities often keep records of how the tissue was collected, the preservation procedures, and demographic information (such as, but not limited too: the origin, duration, age, ethnicity, gender, and stage of disease) of the donor. The demographic information is an important factor in research and in clinical trials. FFPE samples that are properly preserved are very valuable and can be stored at room temperature for a long period of time.

 

Applications

FFPE samples are important as they are often used in:

a)       Immunohistochemistry

The sectioned FFPE specimens are mounted on a slide, bathed in a solution containing antibodies, and then stained so that they can be more clearly seen. This method is important for physicians and researchers looking for pathology in the tissue such as Alzheimer’s or cancer.

b)      Oncology

FFPE samples are vital in the field of oncology as tumor tissues have characteristic morphologies allowing researchers to look for certain proteins. These proteins are then used to help in the assessment of treatment and diagnosis.

c)       Hematology

In the study of blood and its disorders, FFPE samples are important in determining the anomalies and discovery of cures. The specimens can be used in studies related to tissue regeneration, genetics, and toxicology.

d)      Immunology

FFPE samples from a donor with autoimmune disease helps in determining the cause and development of therapy for the condition.

 

Complications or Limitations

One of the possible limitations of the fixation process using formalin is the potential denaturation of the proteins that are present in the tissue making them undetectable to antibodies. To compensate for this issue, antigen retrieval techniques were developed. The antigen retrieval technique specifically recovers DNA, RNA, and proteins from FFPE samples. For this method to work, the quality of FFPE samples are critical. There is also the issue that there is no standard procedure to be used in the preanalytical processing such as fixation and DNA isolation. This means that minor differences such as the different use of instruments, sample handling, and methodology can result in variation that affects the quality of DNA and study results. Some of the factors that have been found to affect study results from FFPE samples are:

  1. Inaccurate logging of fixation protocol
  2. Variation in fixation time
  3. Temperature during fixation
  4. Storage conditions of FFPE samples

 

Quality Control

To ensure the highest quality of FFPE samples, those who collect and store these samples should:

  1. Follow ethical and legal standards.
  2. Keep a clear and accurate record of donors.
  3. Provide information regarding the sampling and collection process
  4. Be supervised by a licensed pathologist during the collection of samples
  5. Have a complete chain of custody
  6. Work only with a carefully selected network of distributors that consistently provide high quality and accurate samples

Fresh Frozen Tissue

Fresh frozen tissues are specimens that are preserved using liquid nitrogen through a method known as “flash freezing”. These specimens are then stored in a freezer that is set at a temperature of less than -80 degrees Celsius. Fresh frozen tissue has different applications than  FFPE samples as they can be used in native morphology studies or molecular analysis as well.

 

FFPE Samples Vs. Fresh Frozen Tissue

FFPE and fresh frozen tissue have their pros and cons. They are two different types of samples that have different uses dependent on the requirements of the research or clinical study. 

  1. FFPE blocks are very hard and can be easily stored at room temperature for decades without the need of special equipment making the type of tissue sample very cost efficient.
  2. There is a large archive of FFPE samples available for researchers due to the easiness associated with it storage.
  3. FFPE specimens have been used for decades making it incredibly familiar to pathologists.
  4. Fresh frozen tissue is much more suitable for the analysis of native proteins, polymerase chain reaction, and next generation DNA sequencing.
  5. Fresh frozen tissue ensures the preservation of DNA, RNA, and native proteins.
  6. Fresh frozen tissues require specialized equipment for storage. This means mechanical failure, power outages, and carelessness can affect the quality of the samples.

 

References:

1)      Ward T. The importance of proper formalin fixation of FFPE samples. Personalis. Accessed 6/19/2018. https://www.personalis.com/importance-proper-formalin-fixation-ffpe-specimens/

2)      Doiron L. 5 quality control rules for cancer tissue banks. Folio Conversant. Accessed 6/19/2018. http://www.conversantbio.com/blog/bid/339034/5-Quality-Control-Rules-for-Cancer-Tissue-Banks

3)      FFPE vs frozen tissue samples. BioChain. Accessed 6/19/2018. https://www.biochain.com/general/ffpe-vs-frozen-tissue-samples/

4)      What is FFPE tissue and what are its uses. BioChain. Accessed 6/19/2018. https://www.biochain.com/general/what-is-ffpe-tissue/