ISOLATE II RNA Micro Kit is specially designed for the fast and efficient isolation of extremely pure total RNA from very small samples. Typical sample material comprises small numbers of cells (up to 5 x 105) and tissue (up to 5 mg) such as laser-captured cells, microdissected cryosections, fine needle aspirates, biopsy samples, pellets of cultured cells and flow cytometer sorted cells. The protocol is easy to follow on a step-by-step basis. The cells are lysed with an optimized lysis buffer containing chaotropic ions, which simultaneously inactivates RNases thus preventing degradation of the released RNA. The lysate is then applied to a spin column to selectively remove genomic DNA (RNase-free DNase I is supplied with the kit). There is no need to perform a separate DNase I digestion step. The RNA is then bound to the silica membrane. The novel design allows elution of RNA in as little as 5–30µl. Simple wash steps remove the remaining cell debris. Pure, highly concentrated RNA is eluted in the final step with RNase-free water. Carrier RNA- poly(-A) RNA is included in the kit, which typically does not interfere with subsequent RT-PCR, but allows for optimal performance with the smallest samples. The online product manual has protocols for purifying total RNA from microdissected cryosections, laser- captured cells and cultured cells, purifying total RNA from tissue, DNase I treatment of purified RNA in solution and clean-up and concentration of RNA from reaction mixtures. The isolated RNA shows excellent performance in downstream applications such as reverse transcription, primer extension, real-time PCR, Northern blot analysis, microarrays and RNA protection assays.

Features

• Rapid protocol: 40 min/12 preps
• Perfect for small samples: biopsy or single cells
• High yield and purity (A260/A280 ratio 1.9 - 2.1)
• Excellent RNA recovery and integrity (RIN >9)
• Extra filters (shredders) and DNase I included

Applications

• Isolation of RNA from small amounts of samples: Single cells, Pellets of cultured cells, Biopsy materials, Cryosections, Laser-captured cells, Small amounts of tissues, Flow sorted cells
• Isolated RNA is ready for downstream applications such as reverse transcription and real-time PCR

Components: ISOLATE II Filters, ISOLATE II RNA Micro Columns, Collection Tubes, Buffer RLY Wash Buffer, RW1 Wash Buffer, RW2 Membrane Desalting Buffer, MEM Reaction Buffer for DNase I, RDN DNase I, RNase-free (lyophilized) Carrier RNA Reducing Agent, TCEP, RNase-free water

Storage Conditions: ISOLATE II RNA Micro Kit contents should be stored dry at room temperature. Under these conditions, the kit is stable for 12 months.