In omics research, such as proteomics, metabolomics, etc., blood is a common sample type. However, there is a more abundant class of sample resources behind this common sample type--- FFPE (formalin fixation and paraffin embedding) samples.
Pathology-relevant biomarker studies often require the extraction of nucleic acids from limited amounts of fixed paraffin-embedded (FFPE) tissue. Among them, a major challenge in studying the transcriptome is the low quantity and quality of RNA extracted, which limits our ability to analyze and has a significant impact on the overall study design.
1. Difficulties in FFPE RNA extraction
The difficulty of RNA extraction of FFPE samples is mainly due to the fact that such samples cannot be fixed in time after leaving the body and face the effect of nuclease degradation, and when they are immobilized, they are subjected to wax immersion in a high heat environment, which further promotes the occurrence of RNA degradation, coupled with long-term storage at room temperature and contact with the outside world, which makes RNA degradation widespread. For these reasons, it seems that it is not possible to extract RNA from FFPE samples.
In 1988, after RUPP et al. first isolated RNA from wax blocks, it was found that the fixative solution used to make FFPE samples can make the RNA in the cell and other macromolecular substances, such as proteins, produce extensive cross-links to form a network, so as to avoid the degradation of nucleases and preserve, on the other hand, nucleases are also denatured and inactivated to a certain extent or even inevitably under the action of fixative, and this essence is the basis for the successful extraction of RNA from FFPE samples.
Generally, nucleic acid extraction and purification have three quality inspection indicators:Nucleic acid concentration, nucleic acid purity and nucleic acid integrity, while FFPE samples damage nucleic acids due to *** treatment, which affects the authenticity of downstream application data and increases the difficulty of nucleic acid extraction.
Effect of fixation on DNA, RNA, and proteins.
Also therefore, extractionffpein the samplernaThe following difficulties exist:
1. Nucleic acids are degraded to varying degrees
2. Cross-linking between molecules;
3. The extracted nucleic acid yield is low, the integrity is not high, the quality is poor, and it is difficult to be compatible with downstream qpcr, sequencing and other applications.
The nucleic acid extraction process from FFPE specimens is a complex multi-step operation that requires strict control of the conditions and methods of each link. Although nucleic acid extraction from FFPE specimens is challenging, with the advancement of science and technology, many efficient and reliable nucleic acid extraction methods and kits have emerged, which provide strong support for the wide application of FFPE specimens in biomedical research.
Anyone who knows medicineFFPE Tissue RNA Extraction Kit (Magnetic Bead Method).
This product isolates high-quality RNA from fixed paraffin-embedded (FFPE) tissue specimens and is designed to provide customers with a simple, reliable, and fast method.
Product Advantages:
The process is fast and safe:It does not rely on toxic organic solvents (such as xylene) for dewaxing, which is safe to use and easy to operate.
High quality RNA products:Selective adsorption of nucleic acids by nanoscale superparamagnetic beads with special surface treatment minimizes genomic DNA contamination of purified RNA samples and provides DNase digestion options.
Flexible use scenarios:It can not only support manual extraction, but also efficiently use with liquid workstations or fully automated nucleic acid extractors to carry out fast, large-volume extraction and improve overall efficiency.
Case Study
Depending on whether FFPE RNA includes particularly small fragment RNA, we have two products, product 1 includes small fragment RNA, and product 2 does not include small fragment RNA. As shown in the image above, customers can choose according to their needs.