This blog was written by guest writers Paraj Mandrekar (Technical Services Scientist 3) and Michelle Mandrekar, (Research Scientist 4).
Here are some designer’s notes comparing the Maxwell® RSC Blood DNA and the Maxwell® RSC simplyRNA kit chemistries for nucleic acid extraction.
The Maxwell RSC Blood DNA Kit and Maxwell RSC simplyRNA Blood Kit were both developed from the same non-silica-based purification chemistry and use the same underlying paramagnetic particle. This chemistry is characterized by an extreme binding capacity (the capacity of nucleic acid that can be bound on the particle), leading to both chemistries being capable of isolating large amounts of nucleic acid volumes and then eluting into relatively small volumes (50 µL). It is not unusual with either chemistry to have isolates that exceed 100 ng/µL. Although the chemistries have several similarities, there are some important distinctions between how the two chemistries were designed that influence which kit you choose for your nucleic acid extraction.
This is part 3 of a three-part series on FFPE sample processing. Part 1 (link) Part 2 (link)
I would like to automate FFPE processing, but I am worried about sample cross contamination, how can I minimize my risks?
As a gold standard for oncology research, hundreds of millions of FFPE samples are collected and banked worldwide. These samples provide a rich source of data for identification of biomarkers in the search for early detection assays for cancer as well as diagnostics that could help direct treatment decisions and monitor treatment.
Formalin-Fixed Paraffin embedded (FFPE) samples are being used in increasing numbers of molecular assays. In my last blog I discussed some of the pre-analytical variables that can affect results obtained when using FFPE samples. Laboratories can increase the quality of downstream results by controlling variables where possible. While exacting control over the sample acquisition and fixation process can improve results, quality testing of incoming samples is a crucial step in assuring optimal results. There are numerous methods that can be used to evaluate the quality of samples and they can provide different information that can be used to assess sample integrity and suitability for different applications.
Formalin Fixed Paraffin Embedded samples (FFPE) have been a mainstay of the pathology lab for over 100 years. Initially FFPE blocks were sectioned, stained with simple dyes and used for studying morphology, but now a variety of biomolecules can be analyzed in these samples. Over the past 10 years we have discovered that there is a treasure trove of genomics data waiting to be unearthed in FFPE tissue. While viral RNAs and miRNA were some of the first molecules found to be present and accessible for analysis starting in the 1990s, improvements to DNA and RNA extraction methods have demonstrated that PCR, qPCR, SNP genotyping, Exome and WGS are possible. This has resulted scientific publications of DNA and RNA data generated from FFPE samples starting in 2006, and today we see immense amounts of data generated from FFPE—with nearly 2000 citations in 2018 reporting sequencing of FFPE samples.
Depending on the type of project, prospective or
retrospective, the genomics scientist has an opportunity to affect the
probability of success by better understanding the fixation process. The
challenge with FFPE is the host of variables that have the potential to
negatively affect downstream assays.
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