The type of cells and fluorescent dyes influenced on the non-specific binding of fluorescently labeled miRNAs to cell surfaces.
The percentage of fluorescence-positive cells in each group of K562 cells (A) and Hela cells (B) treated by Cy5-RS and FAM-RS with and without RNAiMAX was analyzed using flow cytometry and shown on figure. (C) K562 cells or Hela cells were treated by FAM-RS with and without RNAiMAX. The images were scanned by LSCM without nuclear staining using DAPI. The FAM fluorescence image, transmitted light image and merged image of them were shown on figure. The total fluorescence intensity per cell (TFIPC) was also calculated in each group of K562 cells (D) and Hela cells (E) treated by Cy5-RS and FAM-RS with and without RNAiMAX. All statistical analysis was performed by ANOVA. ▲: P<0.05 compared with the groups of K562 or Hela cells transfected by Cy5-RS with and without RNAiMAX. ★: P<0.05 compared with K562 cells transfected by FAM-RS with and without RNAiMAX. △: P<0.05 compared with Hela cells transfected by FAM-RS and RNAiMAX. The structural formula and hydrophilic curve of Cy5 (F) and FAM (G) showed that Cy5 was strongly hydrophobic with LogD 3.81 and FAM was strongly hydrophilic with LogD -5.53 at pH = 7.0.