Fig 2: Study design and Urinary extracellular vesicles (uEV) quality control(A) Study design for the next generation sequencing of uEV mRNAs to assess urine collections, quality and reproducibility (technical comparisons), origins of uEV transcripts (uEV on Tissue map), and discovery of candidate non-invasive markers for diabetic kidney disease (DKD marker candidates).(B–D) Transmission electron micrographs showing uEV of typical morphology and variable sizes; filamentous structures compatible with Tamm-Horsfall protein filaments are visible in the in-set B1.(E) Western blotting showing the presence of uEV-enriched markers CD9 and PDX in uEV preparations.(F) Representative electropherograms of RNA isolated from uEV and analyzed with bioanalyzer using Agilent Pico kit. An RNA peak between 25 and 200 nt was detected in all samples and with this characteristic shape passed quality control.(G) Examples showing signs of nucleic acid degradation (arrows) in uEV RNA samples and not passing quality control. 24 h (24h); albumin excretion rate (AER); blood pressure (BP); body-mass index (BMI); estimated glomerular filtration rate (eGFR); waist-to-hip ratio (WHR); Genotype-Tissue Expression (GTEx) project; glycated hemoglobin (HbA1C); Lymph Node Carcinoma of the prostate (LnCaP), macroalbuminuria (Macro); microalbuminuria (Micro); non-diabetic control (Control); normoalbuminuria (Normo); messenger RNA sequencing (mRNAseq); Podocalyxin (PDX), type 1 or type 2 diabetes (T1D, T2D); urinary extracellular vesicles (uEV); principal component analysis (PCA); single-nucleus RNA sequencing (snRNA-seq).