Isolating exosomal RNA with the Norgen Biotek kit

December 30, 2013

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Newcastle University
Academic Haematology
Research Associate

Company:

Norgen Biotek

Product Name:

Urine Exosome RNA Isolation Kit

Catalog Number:

47200

I am involved in a study to identify microRNA biomarkers for graft versus host disease (GvHD), a potential complication of haematopoetic stem cell transplantation. It would be of great value to identify biomarkers for the onset or severity of GvHD that can be detected in bodily fluids that require minimally invasive sample collection, such as urine. I have used the Norgen Biotek Urine Exosome RNA Isolation kit to successfully isolate exosomal RNA from urine samples, taken from GvHD patients.

Experimental Design and Results Summary

Application

RNA Isolation

Starting Material

Urine

Protocol Overview

The protocol is relatively straight-forward and simple to follow. First, the sample is centrifuged at two different speeds to remove cells, debris and residual bacterial cells. The unique Norgen Biotek kit resin is then added to the sample, which binds exosomes. The sample is then subjected to lysis and wash steps via a column-based method, much similar to other column-based RNA extraction kits on the market. Finally, the total RNA is eluted from the column ready for downstream applications.

Tips

After the final elution step, residual resin can be present in the sample. This will sink to the bottom of the sample tube upon centrifugation and thus, can be removed from the final eluate by pipetting the RNA into a fresh tube

Results Summary

The resulting RNA eluted using this kit is of high quality and can be readily assessed by downstream q-RT-PCR. Yields assessed in our laboratory were higher when directly compared to the ultracentrifugation method of exosomal recovery. The protocol is also faster and does not rely on the specialist equipment required for ultracentrifugation.

Additional Notes

None

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Summary

The Good

Rapid protocol and effective RNA recovery

The Bad

Centrifugation steps prior to column-based RNA extraction are time consuming

The Bottom Line

Efficient kit for the extraction of RNA from urine exosomes that is more effective and less time consuming than the traditional ultracentrifugation method

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