Constitution of biobank of human tissues requires careful handling and storage of biological material, to guarantee the quality of samples. Tissue preparation is also critical for further applications such as transcriptomic profiling. In this study, our aim was to evaluate the impact of different disruption techniques (FastPrep-24 instrument, GentleMACS dissociator, and syringe/needle) and homogenizing buffers (RLT versus QIAzol) on RNA purity and quality of metabolic tissues (adipose tissues, liver and skeletal muscle) present in the COMET Biobank. For all homogenization methods used and tissue types, the A260/280 ratios reached values ≥ 1.8, which are in the range of what is found in human tissues and cell lines, while the A260/230 ratios were however ≤ 1.8, with the lowest value obtained with GentleMACS Dissociator. In addition, GentleMACS Dissociator combined with QIAzol reagent gave the highest RIN value and 28S/18S ratio for all tissues tested, except for muscle. Performing RT-qPCR, Ct values for different housekeeping genes can be influenced by extraction methods and RNA quality of samples. In conclusion, we have demonstrated that different disruption techniques and homogenizing buffers impact the purity and some quality markers of RNA, and can also impact quantification of mRNAs by RT-qPCR in human metabolic tissues.

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Background The objective of the COMET (COllection of MEtabolic Tissues) biobank project is to create a high-quality collection of insulin-sensitive tissues (liver, muscle, adipose tissues, and epiploic artery) and blood sample derivatives (plasma, serum, DNA and RNA), collected from 270 grade 2–3 obese patients undergoing bariatric surgery. Relevant data on patient such as clinical/biological characteristics and sample handling are also collected. For this, our aim was to establish a Quality Management System (QMS) to meet the reliability and quality requirements necessary for its scientific exploitation. Materials and methods The COMET QMS includes: (1) Quality Assurance to standardize all stages of the biobanking process, (2) Quality Controls on samples from the first patients included in order to validate the sample management process and ensure reproducible quality; and 3) “in process” Quality Controls to ensure the reliability of the storage procedures and the stability of the samples over time. Results For serum and plasma, several corrective actions, such as temperature handling and centrifugation conditions, were made to the protocol and led to improvement of the volume and quality of samples. Regarding DNA, all samples evaluated achieved a satisfactory level of purity and integrity and most of them yielded the required DNA quantity. All frozen tissue samples had RNAs of good purity. RNA quality was confirmed by RIN, achieving values in most cases over 7 and efficient amplification of housekeeping genes by RT-qPCR, with no significant differences among samples from the same tissue type. In the “in process” Quality Controls, DNA, RNA, and histological integrity of tissues showed no differences among samples after different preservation times. Conclusion Quality Control results have made it possible to validate the entire biobank process and confirm the utility of implementing QMS to guarantee the quality of a biospecimen collection.