Quality Control and Assessment

European Molecular Genetics Quality Network (EMQN) Certifications aim to improve the quality of molecular genetic testing. EMQN provides external quality assessment (EQA) schemes and educational resources to laboratories worldwide, with a focus on genetic testing. Their EQA schemes involve the distribution of samples to participating laboratories for analysis, allowing them to assess their performance and compare it with other laboratories. This helps identify areas for improvement and ensures the accuracy and reliability of genetic testing results. EMQN also offers various educational activities, including workshops, conferences, and online courses, to enhance the knowledge and skills of laboratory professionals in the field of molecular genetics. Through their initiatives, EMQN plays a crucial role in promoting quality assurance and standardisation in molecular genetic testing, ultimately benefiting patients and healthcare providers.

EATRIS Certificate of Commitment to Quality (ECCQ) emphasizes the importance of research quality and scientific rigor. The ECCQ pilot was launched as a EATRIS product to increase visibility of certified sites and support communication of quality services to users. This program aims to recognise academic and non-profit institutions committed to upholding high standards in translational research. Institutions undergo a thorough evaluation process, including self-assessment, external audit, and peer review. Successful participants receive the certificate, symbolising their dedication to maintaining excellence in research conduct and infrastructure. The pilot program signifies EATRIS’ commitment to promoting quality and best practices in translational research across Europe.

Enhancing Quality in Preclinical Data (EQIPD) quality framework and toolbox. The EQIPD Quality System is a comprehensive framework designed to enhance the quality and reliability of preclinical research. The EQIPD (Experimental Quality In Project Design) initiative aims to improve experimental design and reporting standards by providing researchers with practical tools and guidelines. The EQIPD Quality System consists of several components, including the EQIPD Quality System Wiki, which serves as a knowledge base for researchers to access resources, templates, and examples related to experimental design, conduct, and reporting. The system also incorporates a training program, quality certification, and an EQIPD Label for projects that meet the specified quality criteria. By implementing the EQIPD Quality System, researchers can enhance the robustness, reproducibility, and translatability of their preclinical studies.

The National Institute of Standards and Technology (NIST) is creating new, economical, qualitative reference materials and data for proteomics comparisons, benchmarking and harmonization. Here we describe a large dataset from shotgun proteomic analysis of RM 8461 Human Liver for Proteomics, a reference material being developed. Consensus identifications using multiple search engines and sample preparations demonstrate a homogeneous and fit-for-purpose material that can be incorporated into automated or manual sample preparation workflows, with the resulting data used to directly assess complete sample-to-data workflows and provide harmonization and benchmarking between laboratories and techniques. Data are available via PRIDE with identifier PXD013608.

MicroRNAs are important negative regulators of protein-coding gene expression and have been studied intensively over the past years. Several measurement platforms have been developed to determine relative miRNA abundance in biological samples using different technologies such as small RNA sequencing, reverse transcription–quantitative PCR (RT-qPCR) and (microarray) hybridization. In this study, we systematically compared 12 commercially available platforms for analysis of microRNA expression. We measured an identical set of 20 standardized positive and negative control samples, including human universal reference RNA, human brain RNA and titrations thereof, human serum samples and synthetic spikes from microRNA family members with varying homology. We developed robust quality metrics to objectively assess platform performance in terms of reproducibility, sensitivity, accuracy, specificity and concordance of differential expression. The results indicate that each method has its strengths and weaknesses, which help to guide informed selection of a quantitative microRNA gene expression platform for particular study goals.

DNA methylation reference dataset for methylation profiling by high throughput sequencing (Methyl-seq). The full article can be found at https://doi.org/10.1186/s13059-021-02529-2

Universal Human Reference RNA is an RNA extract used as a positive control in QuantiGene assays. The Universal Human Reference RNA is composed of total RNA from 10 human cell lines (Adenocarcinoma, mammary gland; Melanoma; Hepatoblastoma, liver; Liposarcoma; Adenocarcinoma, cervix; Histiocytic lymphoma, macrophage, histocyte; Embryonal carcinoma, testis; Lymphoblastic leukemia, T lymphoblast; Glioblastoma, brain; Plasmacytoma; myeloma; B lymphocyte). Equal quantities of DNase-treated total RNA from each cell line were pooled to make the universal control.

Human Brain Total RNA is a total RNA sample extracted from neural tissue for use as a positive control in QuantiGene assays.

Variation in RNA expression data can be attributed to a variety of factors including the quality of the starting material, the level of cellularity and RNA yield, the platform employed, and the person performing the experiment. To control for these sources of variability, a common set of external RNA controls has been developed by the External RNA Controls Consortium (ERCC), an ad-hoc group of academic, private, and public organizations hosted by the National Institute of Standards and Technology (NIST). The controls consist of a set of unlabeled, polyadenylated transcripts designed to be added to an RNA analysis experiment after sample isolation, in order to measure against defined performance criteria. Up until the design of such universally accepted controls, it has been difficult to execute a thorough investigation of fundamental analytical performance metrics. From the trusted brand of quality RNA reagents, Ambion™ ERCC Spike-In Control Mixes are commercially available, pre-formulated blends of 92 transcripts, derived and traceable from NIST-certified DNA plasmids. The transcripts are designed to be 250 to 2,000 nt in length, which mimic natural eukaryotic mRNAs.

Reference materials are vital to benchmarking the reproducibility of clinical tests and essential for monitoring laboratory performance for clinical proteomics. The reference material utilized for mass spectrometric analysis of the human proteome would ideally contain enough proteins to be suitably representative of the human proteome, as well as exhibit a stable protein composition in different batches of sample regeneration. Previously, The Clinical Proteomic Tumor Analysis Consortium (CPTAC) utilized a PDX-derived comparative reference (CompRef) materials for the longitudinal assessment of proteomic performance; however, inherent drawbacks of PDX-derived material, including extended time needed to grow tumors and high level of expertise needed, have resulted in efforts to identify a new source of CompRef material. In this study, we examined the utility of using a panel of seven cancer cell lines, NCI-7 Cell Line Panel, as a reference material for mass spectrometric analysis of human proteome. Our results showed that not only is the NCI-7 material suitable for benchmarking laboratory sample preparation methods, but also NCI-7 sample generation is highly reproducible at both the global and phosphoprotein levels. In addition, the predicted genomic and experimental coverage of the NCI-7 proteome suggests the NCI-7 material may also have applications as a universal standard proteomic reference.

The Peptide Mix and LC/MS Instrument Performance Monitoring Software includes a 6×5 LC-MS/MS Peptide Reference Mix. This combination provides a comprehensive solution for monitoring and optimising LC/MS instrument performance including sensitivity and dynamic range. Researchers can easily optimise their LC/MS system and ensure reliable and precise results.

Unmethylated lambda DNA and CpG-methylated pUC19 DNA, two essential spike-ins employed in EM-seq library preparation. The unmethylated lambda DNA serves to evaluate conversion efficiency, while the CpG-methylated pUC19 DNA acts as a control to ensure accurate and reliable results. These spike-ins play a crucial role in validating the quality of EM-seq libraries, enabling researchers to achieve robust and precise outcomes in their experiments.

NA12878 is a human genomic DNA sample standardised by the National Institute of Standards and Technology (NIST). It serves as a reference standard for the Genome in a Bottle consortium, which is led by NIST. The donor subject, NA12878’s mother, carries a single bp (G-to-A) transition at nucleotide 681 in exon 5 of the CYP2C19 gene (CYP2C19*2). This genetic change creates an aberrant splice site, resulting in a 40-bp deletion at the beginning of exon 5 (from bp 643 to bp 682). As a consequence, amino acids 215 to 227 are deleted from the mRNA, causing an alteration in the reading frame and the production of a truncated, nonfunctional protein. The truncated protein contains 234 amino acids and lacks the heme-binding region, rendering it catalytically inactive.

Multi-omics reference materials and practical tools to enhance the reproducibility and reliability of multi-omics results (Fudan Quartet samples) provided as part of the Quartet project. These well-characterised resources serve as quality control measures, ensuring accurate data integration in precision medicine studies.

SEQC2: somatic mutation reference dataset, https://pubmed.ncbi.nlm.nih.gov/34753956/

The emergence of MS-based proteomic platforms as a prominent technology utilized in biochemical and biomedical research has increased the need for high-quality MS measurements. To address this need, National Institute of Standards and Technology (NIST) reference material (RM) 8323 yeast protein extract is introduced as a proteomics quality control material for benchmarking the preanalytical and analytical performance of proteomics-based experimental workflows. RM 8323 yeast protein extract is based upon the well-characterized eukaryote Saccharomyces cerevisiae and can be utilized in the design and optimization of proteomics-based methodologies from sample preparation to data analysis. To demonstrate its utility as a proteomics quality control material, we coupled LC-MS/MS measurements of RM 8323 with the NIST MS Quality Control (MSQC) performance metrics to quantitatively assess the LC-MS/MS instrumentation parameters that influence measurement accuracy, repeatability, and reproducibility. Due to the complexity of the yeast proteome, we also demonstrate how NIST RM 8323, along with the NIST MSQC performance metrics, can be used in the evaluation and optimization of proteomics-based sample preparation methods.

This tools have been developed by Hartwig. They are open source but are also part of a clinically accredited diagnostic service for cancer care. This means that the individual tools and the pipeline as a whole are part of a rigorous quality control system and is subjected to external quality assessment rounds.

ISO Guide 31:2015 guideline assists reference material (RM) producers in creating comprehensive documentation for their RMs. It outlines the essential information to include in product information sheets and RM certificates, categorizing them as mandatory, recommended, or optional. This information helps users and stakeholders assess the appropriateness of an RM or Certified Reference Material (CRM). This guideline also sets out the minimum criteria for labeling the RM container.

ISO Guide 80:2014 guidance for the in-house preparation of quality control materials (QCMs). ISO Guide 80 outlines the characteristics and preparation processes of reference materials for quality control. It applies to stable materials used locally and those transported without significant property changes. Laboratory staff preparing in-house quality control materials should follow ISO Guides 34 and 35 for transportation-based supply chains. The preparation of quality control materials requires assessments for homogeneity, stability, and limited characterization. It aims to demonstrate statistical control in a measurement system but does not provide usage guidance. The guide offers general information on preparation and includes case studies for different sectors. Users should have material knowledge and be aware of matrix effects and contamination risks.

Luxembourg Institute of Health (LIH) Proficiency Testing (PT) programme, which works as an external quality assessment tool, allows you to verify and benchmark the performance of your biospecimen processing or testing methods. Once registered, you receive standardised samples produced and shipped by LIH to your laboratory. You use either your routine processing method to extract the samples or your routine testing method to characterise the samples. Following processing or testing, you send LIH the extracted samples (processing) via mail, or the results (testing) via an online platform. LIH analyse the received samples, gather the results from all participants and perform statistical analyses and benchmarking. In the end, each participant receives a personalized report, a certificate and a label of participation.