Detection of Ribonucleic Acids Improved For Molecular Diagnostics

A new protocol for the detection of short ribonucleic acids (RNA) known as micro-RNA (miRNA) will help isolate sufficient quantities for diagnostic purposes.

The method shows how effectively collecting blood plasma samples, brings a step closer the goal of making miRNA disease diagnostics a reality as the choice of blood collection tube affects quantitation.

A study carried out at the Rosalind Franklin University of Medicine and Science. (Chicago, IL, USA) determined what factors can interfere with the accuracy of miRNA tests The scientists provided clear procedures for the collection and analysis of miRNA, significantly improving their diagnostic accuracy. MicroRNAs regulate processes including fertilization, development, and aging show promise as biomarkers of disease.

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 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. The investigators 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. They also observed that differences in plasma composition among individual donors yield different miRNA measurements.

MicroRNAs can be collected from routinely collected fluids such as blood, saliva, and urine. Over 1,000 miRNAs exist in the human body and deregulation of specific miRNAs is associated with disease. Dominik M. Duelli, PhD, a lead investigator said, "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." The study was published online on December 8, 2011, the Journal of Molecular Diagnostics.

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Rosalind Franklin University of Medicine and Science