PicoPure™ RNA Isolation Kit, 40 samples - FAQs

The PicoPure RNA Isolation Kit can be used with as little as a single cell to samples with up to 100 µg of RNA.

Find additional tips, troubleshooting help, and resources within our Nucleic Acid Purification and Analysis Support Center.

The PicoPure RNA Isolation Kit is not recommended for isolation of RNA smaller than 100 nucleotides.

Formalin-fixed paraffin-embedded (FFPE) samples introduce unique challenges for gene expression profiling and gene expression quantitation, including chemical modification and fragmentation of RNA molecules. The initial paraffin blot preparation and storage significant affect the RNA quality. Archival FFPE samples present additional challenges due to increased RNA degradation over time. As a result, not all FFPE samples contain high quality RNA. 

In addition, a typical Bioanalyzer profile of RNA from an FFPE sample can look fragmented and degraded and as a result, that approach may not be useful in determining the transcribability of your RNA. Therefore, a tissue scrape test is recommended. You can use the entire adjacent section to isolate RNA and to access the RNA quantity and quality for intact RNA. Two sets of β-actin gene-specific primers designed to amplify short segments of DNA within the β-actin target sequence from the 5’ and 3’ of the β-actin gene can be used to estimate the amount of RNA in a given sample in relation to β-actin content using real-time PCR. The RNA quantity derived from the 3' primer set is used to quantitate the RNA in the FFPE tissue scrape. The ratio of the RNA yield obtained from both sets of PCR primers is the 3'⁄5' ratio and is used as an indication of RNA quality. If there is less product yield of the 5’ β-actin primers, it indicates that there is less full length of RNA.

The dissection and embedding steps are critical for the RNA quality for LCM samples. We suggest using the entire adjacent section or dissecting a large area of the section (>1000 cells) for RNA isolation. Before proceeding to LCM microdissection, we recommend assessing the quality of RNA isolated from the adjacent section. With good amount of RNA, one can assess the RNA quantity and quality using the Qubit RNA HS Assay Kit and the Agilent Bioanalyzer. This will help to make sure that good quality RNA is obtained from a low number of LCM cells.

The Arcturus PicoPure RNA Isolation Kit removes total cellular RNA from pico-scale samples including frozen sections, alcohol fixed-, or fresh samples. Please note, samples fixed with formaldehyde are not recommended for use with this kit.

Some other kits have been used for RNA isolation from LCM cells. These kits include the RecoverAll Total Nucleic Acid Isolation Kit for Formalin-Fixed Paraffin-Embedded (FFPE) (Cat. No. AM1975), RecoverAll Multi-Sample RNA/DNA Isolation Workflow (Cat. No. A26135), and the RNAqueous- Micro Total RNA Isolation Kit (Cat. No. AM19310).

The Arcturus PicoPure RNA Isolation Kit (Cat. Nos. KIT0204, KIT0214) is specifically designed to consistently recover high-quality total RNA from fewer than 10 cells, even from a single cell. This kit is designed to provide consistent, efficient RNA recovery without sacrificing quality. Small volume elution allows you to maximize your recovery of RNA from small numbers of cells for use with your gene expression analysis experiments.

For LCM cells from paraffin sections, we recommend using the Arcturus Paradise Plus RNA Extraction and Isolation Kit (Cat. No. KIT0312I) that unlocks RNA from formalin-fixed, paraffin-embedded (FFPE) tissues. As little as 5 ng of the isolated fixed RNA can then be amplified using reagents and methods that have been specially optimized to overcome the challenges of cross-linked FFPE templates.

We also recommend the Arcturus Paradise PLUS QC Kit (Cat. Nos. KIT0303, KIT0313). This kit offers a simple solution for evaluating the quality of RNA from formalin-fixed paraffin-embedded (FFPE) tissues before proceeding with LCM and downstream molecular analysis. The reagents provided in the kit are optimized to enable efficient extraction, isolation, and reverse transcription of total RNA from FFPE tissue scrapes. The RNA quantity and quality is then measured using quantitative real-time PCR with a housekeeping gene like beta actin.