Our exRNAQC study clarifies the impact of blood collection and processing variables on extracellular RNA (exRNA), or cell-free RNA (cfRNA), analysis. We evaluated pre-analytical factors at an unprecedented scale, both for small and long RNAs. Our goal was three-fold: raising awareness of the impact of these factors, providing performance metrics for evaluating newly developed blood collection tubes and RNA purification methods, and establishing standardized guidelines for high-quality sample collection and profiling – addressing a critical gap in the research community.

The spark: a whiteboard discussion

The journey began in 2017 with a simple but pressing question: how should we collect and process blood samples to refine and enhance our exRNA analyses? We had observed that pre-analytical variables substantially impacted our blood-based liquid biopsy biomarker analyses in the childhood cancer neuroblastoma – leaving us puzzled about the best strategy to obtain high-quality exRNA data. As we searched the literature for answers, it became evident that our lab was not alone in facing this challenge. Many research teams were struggling with the same issue, yet no consensus existed on best practices for blood sample collection and processing. What started as an internal discussion soon turned into an ambitious research effort – one that would grow far beyond our initial scope.

In total, we evaluated ten different blood collection tubes, eight RNA purification kits, and three time intervals between blood collection and processing. To do so, we ended up collecting more than 1.6 liters of blood and generating over 11 billion sequencing reads to compare 456 extracellular transcriptomes. Each time, we evaluated multiple performance metrics, such as the number of genes, RNA concentration, RNA purification efficiency, and replicate variability. Our findings showed that not only the individual choice of kit, tube, and time to processing matters, but also the specific combination of these factors. Using the insights from our study, we were also able to provide recommendations for both end users and manufacturers of kits and tubes.

A collaborative endeavor

Scientific breakthroughs rarely happen in isolation, and this study was no exception. The complexity of the problem meant that no single researcher could tackle it alone. What followed was a large-scale effort initiated by the OncoRNALab (Ghent University, Belgium), with Prof. Jo Vandesompele and Prof. Pieter Mestdagh bringing together researchers from diverse backgrounds under the exRNAQC Consortium. Several stakeholders, including Illumina, Qiagen, Promega, and Roche, recognized the importance of our study and therefore decided to sponsor reagents for evaluation. In addition, our study was funded by multiple Belgian and European non-profit funding agencies. Without this support, successful completion of the project would not have been possible in an academic research setting.

From securing funding and optimizing methodologies to conducting experiments and developing data analysis pipelines, every aspect of this project was fueled by teamwork. The study far exceeded what we initially envisioned, not only in the number of pre-analytical variables being tested and compared, but also in the sheer number of researchers who joined forces throughout the study. While changes in team composition and dynamics also come with challenges, the study is a great example of what can be accomplished when uniting behind a common goal – and how much we can learn from each other along the way.

Rethinking scientific recognition

In academia, publications are often seen as markers of productivity, with career progression tied to authorship positions. Yet, for a project as deeply collaborative as this, we resisted the pressure to follow the traditional linear co-author model, where recognition is concentrated at the first and last author positions. Instead, we embraced a different philosophy – one that values shared knowledge over individual accolades.

By using the CRediT (Contributor Roles Taxonomy) system, we ensured that every team member’s contributions were properly acknowledged, reflecting the collaborative nature of our work. As such, this publication illustrates that science advances not through prestige but through openness, teamwork, and collective effort.

Looking ahead

While this publication marks the end of one chapter, it is just the beginning of another. Our hope is that the insights gained from this study will help standardize the exRNA field, reducing variability and improving the reliability of liquid biopsy-based analyses. We also hope it serves as an inspiration for future large-scale research collaborations.

So, to everyone who contributed to this journey – whether by running experiments, analyzing data, providing insights, or supporting the project in any way – this achievement belongs to all of us.

Here’s to teamwork, perseverance, and the shared pursuit of scientific progress!

Original post can be found here.