Forum for Science, Industry and Business

Sponsored by:     3M 
Search our Site:

 

New protocols improve detection of microRNAs for diagnosis

07.12.2011
MicroRNAs (miRNAs) that regulate processes including fertilization, development, and aging show promise as biomarkers of disease.

They can be collected from routinely collected fluids such as blood, saliva, and urine. However, a number of factors can interfere with the accuracy of miRNA tests. In a study published online today in the Journal of Molecular Diagnostics, a group of researchers provide clear procedures for the collection and analysis of miRNA, significantly improving their diagnostic accuracy.

"Our study demonstrates that inherent differences in biological samples and the methods used to collect and analyze them can dramatically affect the detection and quantitation of microRNAs," reports lead investigator Dominik M. Duelli, PhD, Department of Cellular and Molecular Pharmacology, Chicago Medical School at Rosalind Franklin University of Medicine and Science. "We developed measures to overcome the interfering activities and improved the sensitivity of miRNA detection up to 30-fold."

Over 1,000 miRNAs exist in the human body. Deregulation of specific miRNAs is associated with disease. Measuring the amount of miRNAs in body fluids can aid in the diagnosis of disease or conditions such as pregnancy. Dr. Duelli and his colleagues quantified two miRNAs: miR-16, which acts as a tumor suppressor and is deregulated or lost in some cancers, including breast cancer; and miR-223, which has been implicated in pregnancy and other conditions, as well as in some malignant diseases.

"A fundamental challenge to making microRNA diagnostics broadly available has been the inability to isolate enough high quality material to analyze. Our paper outlines ways of effectively collecting blood plasma samples, thus bringing us one step closer to the goal of making [miRNA] disease diagnostics a reality," adds co-investigator Sarah Linnstaedt, PhD, of Duke University.

The authors found that the choice of blood collection tube affects quantitation. Traditional green-top heparin tubes interfered nearly completely with miRNA detection. Grey-top tubes containing the anticoagulant sodium fluoride and potassium oxalate (NaF/KOx) provided the best results. Although miR-16 is about 500 times more abundant in blood plasma than miR-223, the results for both were similar, indicating that the differences in detection resulting from the choice of collection method apply to other miRNAs. Furthermore, collection of miR-223 in serum yielded more variable results, signifying that for some miRNAs, analysis of blood in plasma form is preferred.

The study indicated that natural components of blood plasma co-purify with miRNAs, interfering with their detection. The authors identified extra steps in purification, and the ideal dilution level, to reduce the interference. "Although counterintuitive, by reducing the starting material, inhibitors were presumed to be diluted below a threshold of interference. Careful titration of starting material yields more accurate miRNA quantitation," explains Dr. Duelli. In another approach, the authors avoided the problem of contamination by combining an enzyme that overcomes plasma inhibitors with standard enzymes to increase the sensitivity of miRNA detection by about 30-fold.

Finally, the authors observed that differences in plasma composition among individual donors yield different miRNA measurements. "These results raise the possibility that factors including diet, exercise, circadian rhythms, and seasons, which alter the blood chemistry, might affect miRNA detection and quantitation," says Dr. Duelli.

"The implications of this work are that without consideration of the variables we have identified, miRNA quantitation of human samples may not be reliable for the purpose of biomarker development. We provide approaches that enable faithful quantitation of miRNA abundance in body fluid," concludes Dr. Duelli.

The article is "Plasma Components Affect Accuracy of Circulating Cancer-Related MicroRNA Quantitation," by D-J. Kim, S. Linnstaedt, J. Palma, J. Cheol Park, E. Ntrivalas, J.Y.H. Kwak-Kim, A. Gilman-Sachs, K. Beaman, M.L. Hastings, J.N. Martin, and D.M. Duelli (doi: 10.1016/j.jmoldx.2011.09.002). Published online ahead of its issue, the study will appear in the Journal of Molecular Diagnostics, Volume 14, Issue 1 (January 2012) published by Elsevier.

David Sampson | EurekAlert!
Further information:
http://www.elsevier.com

All articles from Life Sciences >>>

The most recent press releases about innovation >>>

Die letzten 5 Focus-News des innovations-reports im Überblick:

Im Focus: Making lightweight construction suitable for series production

More and more automobile companies are focusing on body parts made of carbon fiber reinforced plastics (CFRP). However, manufacturing and repair costs must be further reduced in order to make CFRP more economical in use. Together with the Volkswagen AG and five other partners in the project HolQueSt 3D, the Laser Zentrum Hannover e.V. (LZH) has developed laser processes for the automatic trimming, drilling and repair of three-dimensional components.

Automated manufacturing processes are the basis for ultimately establishing the series production of CFRP components. In the project HolQueSt 3D, the LZH has...

Im Focus: Wonder material? Novel nanotube structure strengthens thin films for flexible electronics

Reflecting the structure of composites found in nature and the ancient world, researchers at the University of Illinois at Urbana-Champaign have synthesized thin carbon nanotube (CNT) textiles that exhibit both high electrical conductivity and a level of toughness that is about fifty times higher than copper films, currently used in electronics.

"The structural robustness of thin metal films has significant importance for the reliable operation of smart skin and flexible electronics including...

Im Focus: Deep inside Galaxy M87

The nearby, giant radio galaxy M87 hosts a supermassive black hole (BH) and is well-known for its bright jet dominating the spectrum over ten orders of magnitude in frequency. Due to its proximity, jet prominence, and the large black hole mass, M87 is the best laboratory for investigating the formation, acceleration, and collimation of relativistic jets. A research team led by Silke Britzen from the Max Planck Institute for Radio Astronomy in Bonn, Germany, has found strong indication for turbulent processes connecting the accretion disk and the jet of that galaxy providing insights into the longstanding problem of the origin of astrophysical jets.

Supermassive black holes form some of the most enigmatic phenomena in astrophysics. Their enormous energy output is supposed to be generated by the...

Im Focus: A Quantum Low Pass for Photons

Physicists in Garching observe novel quantum effect that limits the number of emitted photons.

The probability to find a certain number of photons inside a laser pulse usually corresponds to a classical distribution of independent events, the so-called...

Im Focus: Microprocessors based on a layer of just three atoms

Microprocessors based on atomically thin materials hold the promise of the evolution of traditional processors as well as new applications in the field of flexible electronics. Now, a TU Wien research team led by Thomas Müller has made a breakthrough in this field as part of an ongoing research project.

Two-dimensional materials, or 2D materials for short, are extremely versatile, although – or often more precisely because – they are made up of just one or a...

All Focus news of the innovation-report >>>

Anzeige

Anzeige

Event News

Expert meeting “Health Business Connect” will connect international medical technology companies

20.04.2017 | Event News

Wenn der Computer das Gehirn austrickst

18.04.2017 | Event News

7th International Conference on Crystalline Silicon Photovoltaics in Freiburg on April 3-5, 2017

03.04.2017 | Event News

 
Latest News

DGIST develops 20 times faster biosensor

24.04.2017 | Physics and Astronomy

Nanoimprinted hyperlens array: Paving the way for practical super-resolution imaging

24.04.2017 | Materials Sciences

Atomic-level motion may drive bacteria's ability to evade immune system defenses

24.04.2017 | Life Sciences

VideoLinks
B2B-VideoLinks
More VideoLinks >>>