Semiquantitative analysis of carbon quantum dot-labeled exosomes based on carrier ampholyte-free paper-based...

In this paper, a semiquantitative analysis method based on carrier ampholyte-free paper-based isoelectric focusing (CAF-PAD-IEF) was developed for the isolation and detection of carbon quantum dots (CQDs) labeled exosomes. A stable pH gradient was established on the glass fiber paper by the migration of H⁺ and OH^-. Three pH gradients of pH 3-9, 3-7 and 6-9 were established by changing the species and pH of anolytes and catholytes. The pH gradients were evaluated with phycocyanin (PC), ovalbumin and bovine hemoglobin (BHb) as standard proteins, and egg white was used as model sample too. The linear correlation coefficient (R²) of the pH gradient (pH 3-9) reached 0.9966. Exosomes obtained by ultracentrifugation from urine were used as the standard to realize the semiquantitative detection of exosomes. The urine exosomes were labeled with CQDs and then successfully focused based on their isoelectric points (pIs). The exosome concentration was determined semiquantitatively according to the fluorescence intensity of the focused bands. The standard curve was drawn based on the fluorescence intensity peak areas of the focused bands with the R² value of 0.9881. This method was applied to the determination of exosome concentration in fetal bovine serum (FBS) and concentrated urine with the relative errors of -4.17 % and -8.70 % respectively compared with the values determined by nanoparticle tracking analysis (NTA), showing great potential in quick semiquantitative determination of exosomes in real samples.