A systematic review and meta-analysis of preanalytical factors and methodological differences influencing the measurement of circulating vascular endothelial growth factor

Background: Intraocular treatment with antibodies targeting vascular endothelial growth factor (anti-VEGF) inhibits pathological vessel growth in adults and preterm infants. Recently, concerns regarding the impact of anti-VEGF treatment on systemic VEGF levels in preterm infants have been raised. Earlier studies suggest that preanalytical and methodological parameters impact analytical VEGF concentrations, but we have not found a comprehensive systematic review covering preanalytical procedures and methods for VEGF measurements. Objective: This review aimed to evaluate the most critical factors during sample collection, sample handling, and the analytical methods that influence VEGF levels and therefore should be considered when planning a prospective collection of samples to get reproducible, comparable results. Material and methods: PubMed and Scopus databases were searched 2021/Nov/11. In addition, identification of records via other methods included reference, citation, and Google Scholar searches. Rayyan QCRI was used to handle duplicates and the selection process. Publications reporting preanalytical handling and/or methodological comparisons using human blood samples were included. Exclusion criteria were biological, environmental, genetic, or physiological factors affecting VEGF. The data extraction sheets included bias assessment using the QUADAS-2 tool, evaluating patient selection, index-test, reference standard, and flow and timing. Concentrations of VEGF and results from statistical comparisons of analytical methods and/or preanalytical sample handling and/or different sample systems were extracted. The publications covering preanalytical procedures were further categorized based on the stage of the preanalytical procedure. Meta-analysis was used to visualize VEGF concentrations among healthy individuals. The quality of evidence was rated according to GRADE. Results: We identified 1596 publications, and, after the screening process, 43 were considered eligible for this systematic review. The risk of bias estimation was difficult for 2/4 domains due to non-reported information. Four critical steps in the preanalytical process that impacted VEGF quantification were identified: blood drawing and the handling before, during, and after centrifugation. Sub-categorization of those elements resulted in nine findings, rated from moderate to very low evidence grade. The choice of sample system was the most reported factor. VEGF levels (mean [95% CI]) in serum (n = 906, 20 publications), (252.5 [213.1–291.9] pg/mL), were approximated to ninefold higher than in plasma (n = 1122, 23 publications), (27.8 [23.6–32.1] pg/mL), based on summarized VEGF levels with meta-analysis. Notably, most reported plasma levels were below the calibration range of the used method. Conclusion: When measuring circulating VEGF levels, choice of sample system and sample handling are important factors to consider for ensuring high reproducibility and allowing study comparisons. Protocol: CRD42020192433