The Yin and Yang of microRNA Assay Methods.
The Yin and Yang of microRNA Assay Methods.
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microRNA 测定方法的阴阳。
DOI：
10.2174/2211536605666160725130028
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发表时间：
2017
期刊：
影响因子：
--
通讯作者：
J. Issartel
中科院分区：
文献类型：
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作者：
Audrey Guttin;Hélène Ipas;M. Barbado;C. Mouret;E. Garçion;J. Issartel
客观化
生物样品中的microRNA检测可以通过不同的方法进行，主要依靠杂交过程、qPCR或RNA测序。为了检测和验证肿瘤样本中的microRNA生物标志物，我们对不同方法获得的定量结果的一致性提出了质疑。
方法
我们使用不同的microRNA分析方法，特别是与AffymetrixTM阵列杂交、qPCR和基于Taqman microRNA分析(Life TechnologiesTM)的数字液滴qPCR(BioradTM)，测量了几种生物样本中的microRNA浓度，例如培养的肿瘤细胞或肿瘤组织(冰冻组织或FFPE样本)。我们还将我们的结果与通过不同技术方法获得并在文献中可用的其他数据进行了比较。
结果
我们发现，用不同方法测定的样品中microRNA含量的一致性很差。技术平台和microRNA检测方案可能是造成观察到的不一致的原因。
结论
在为临床目的或基础研究分析microRNA时，似乎有必要记住所有microRNA检测方法的具体缺陷，例如我们在这里披露的那些方法。显然，只有在严格使用相同的技术方法和试剂进行microRNA评估时，才能获得有效的样本间比较和有意义的多中心研究。
OBJECTIVE
microRNA assessments in biological samples can be performed by different methods that mainly rely on hybridization process, qPCR or RNA sequencing. With the aim to detect and validate microRNA biomarkers in tumor samples, we challenged the consistency of the quantitative results obtained with the different methods.
METHODS
We measured microRNA concentrations in several biological samples such as cultured tumor cells or tumor tissues (frozen tissues or FFPE samples) using different microRNA assay methods, in particular hybridization to AffymetrixTM arrays, qPCR and digital droplet qPCR (BioradTM) based on Taqman microRNA assays (Life TechnologiesTM). We also compared our results to other data that have been obtained with different technical approaches and available in the literature.
RESULTS
We found poor consistency for the microRNA amounts measured in the samples assayed by the different methods. Both technical platforms and microRNA assays protocols may be responsible for the observed inconsistencies.
CONCLUSION
When assaying microRNAs for clinical purpose or fundamental researches it seems necessary to keep in mind the specific pitfalls of all the microRNA detection methods such as those we disclose here. Obviously, valid inter sample comparisons and meaningful multicenter studies can only be obtained when microRNA assessments are strictly performed with identical technical approaches and reagents.