**Conflict of interest**

over a decade now, the majority of methodologies used for miRNA extraction and profiling

The major points affecting the miRNA extraction and application are choice of coagulant, plasma preparation and biological condition. Among the three most widely used anticoagulants (EDTA, citrate, and heparin), EDTA is shown to be the least interfering chemical in the subsequent miRNA profiling, while heparin and citrate interfere with enzymes used for various PCR [23]. Circulating miRNAs are either encapsulated in vesicles or found in complexes with proteins and lipoproteins, therefore are considered to be relatively stable. However, it is recommended that plasma preparation is done within 2 h of phlebotomy since blood cells start to release miRNAs into the collected sample causing changes in miRNA profile. Even mild haemolysis is also considered as an interfering factor associated with miRNA contamination from blood cells. Visual identification of minor haemolysis is difficult, therefore a simple measurement of absorbance can be an accurate and time saving solution. Prepared plasma samples could be quickly tested for haemolysis by measuring the absorbance peak of free haemoglobin at 414 nm [24]. Plasma samples with A<sup>414</sup> > 0.18 show signs of miRNA released from erythrocytes which might interfere with the overall profile of circulating miRNAs [24]. It is worth noting that lipaemia affects the A414 measurement; therefore, it is recommended to perform a second measurement at A385 to detect the presence of lipaemia and use it to calculate a lipaemia-independent haemolysis score which could be adopted as a pre-analytical quality control [25]. Circulating miRNAs are often found in association with lipoproteins, therefore biological conditions, such as fasting, might have an impact on miRNA profiling [23]. The use of miRNA biomarkers in clinical setting is only starting therefore to standardise the pre-analytical procedure of sampling

and sample preparation it is worth to have as little environmental variables as possible.

A medical laboratory or clinical laboratory are laboratories for the examination of materials derived from the human body with a purpose of providing necessary information for diagnosis, prevention and treatment of disease as well as for follow up evaluation of health of a patient during the treatment. The 'International Organization for Standardization' (ISO) is an 'International Authority' for setting up standard guidelines for various organisations and laboratories. Each quality manual is based on internationally accepted standards and provides guidance for public health and clinical laboratories on writing policies and procedures that support a quality management system. It comprises a main document providing information and written procedures for laboratory quality (standard operating procedures, forms, and processes). It is worth to remember always to address quality manual before starting testing,

The chapter was supported by project "Age-related remodelling of aorta and dilatative pathology of ascending aorta: search for epigenetic biomarkers", project number: SEN-05/2016.

**2.9. Quality manual for laboratory testing**

14 Plasma Medicine - Concepts and Clinical Applications

sampling or consulting patients [24].

**Acknowledgements**

come from a scientific research and are not approved for clinical use yet.

There is no conflict of interests.
