PAXgene Blood miRNA Kit
1. Can the PAXgene Blood miRNA Kit be used for diagnostic or prognostic procedures?
No. The product is ‘For Research Use.’ Not for use in diagnostic procedures. No claim or representation is intended to provide information for the diagnosis, prevention, or treatment of a disease.
2. How great is the enrichment of miRNA when using the dedicated PAXgene Blood miRNA Kit compared to the PAXgene Blood RNA Kit?
If the PAXgene Blood miRNA Kit is used, an enrichment of small RNAs as part of the total RNA is achieved. The enrichment may vary between different miRNA species and samples, but on average an enrichment of about three or more CT-values can be expected.
3. How many samples can be processed with the PAXgene Blood miRNA Kit?
The kit can be used for 50 RNA isolations.
4. Which sample types can be processed using the PAXgene Blood miRNA protocols?
2.5 ml whole human blood collected in PAXgene Blood RNA Tubes.
5. Do the samples require pretreatment before starting the manual protocol or before loading onto the QIAcube instrument?
Yes. Samples have to be incubated at least 2 h at room temperature (18-22°C). Incubation of the PAXgene Blood RNA Tube overnight may increase yields slightly in some cases. Room temperature storage for up to three days has no effect on the RNA profile. During the preparation of the RNA the manual pretreatment consist of a centrifugation and a pellet resuspenion step (see thePAXgene Blood miRNA Handbook).
6. Do you have stability data for RNA isolated with PAXgene miRNA Kit?
Stability data are not yet available, but a stability study was started.
7. What are the expected yields of RNA from PAXgene Blood RNA Tubes isolated with the PAXgene Blood miRNA kit?
Typical yields of RNA isolated from 2.5 ml healthy, human, whole blood (4.8 x 106 –1.1 x 107 leukocytes/ml) are ≥3 μg for >95% of the samples processed (seeTechnical Note). Since yields are highly donor-dependent, individual yields may vary.
8. How should the samples be stored before purification of RNA?
Room temperature storage of the filled PAXgene Blood RNA Tubes for up to three days has no effect on the RNA profile. For longer storage they should be stored at –20°C or –70°C.
9. What is the maximum number of samples for the QIAcube variant of the PAXgene Blood miRNA protocol for a single run?
Up to 12 samples can be processed in a single run.
10. Why is the final heat denaturation step so important?
Denaturation of the eluate is essential for maximum efficiency in downstream applications, such as RT-PCR, other amplification reactions, or cDNA synthesis. It is not necessary to denature samples more than once; samples remain denatured after freezing and thawing.
11. Is it possible to isolate genomic DNA or virus nucleic acids with PAXgene Blood miRNA protocol?
No. The PAXgene Blood RNA System has been optimized for cellular RNA only. PreAnalytiX offers a dedicated system for collection and isolation of genomic DNA from human whole blood (see PAXgene Blood DNA System).
12. Do eluates generated with the PAXgene Blood miRNA Kit contain residual genomic DNA?
Typically, the amount of genomic DNA (gDNA) in eluates of the PAXgene Blood miRNA Kit is very low as long as the kit is stored and used according to the instructions of the PAXgene Blood miRNA Kit Handbook. For downstream applications that are especially sensitive to gDNA contamination, a specialized protocol for the QIAcube helps reduce gDNA content further and can be downloaded from the QIAcube product page at www.qiagen.com.