Table 1.
Current strategies to evaluate internalization of complexes into MSCs including uptake and inhibition assays
| NAs | Fluorescent probe for labeling | Endocytic inhibitors | Analytical methods | Ref |
|---|---|---|---|---|
| miRNA (miR-218) | Alexa Fluor 647 (red, miRNA), Lysotracker Green (green, lysosomes), and Hoechst 33342 (UV, cell nucleus) | NO | Confocal microscopy | [182] |
| pDNA (pBMP-2) | FITC (green, PEI) and DAPI (UV, cell nucleus) | NO | Confocal microscopy | [5] |
| pDNA (pGFP) | Cy3 (red, pDNA), WGA-Alexa Fluor (green, cytoplasm) and DAPI (UV, cell nucleus) | NO | Cy3 positive by FCM and confocal microscopy | [86] |
| miRNA (miR-133a) | Dy547 (red, miRNA) | NO | Fluorescence plate reader | [206] |
| pDNA (pBMP-2) | YOYO-1 (red, pDNA) and DAPI (UV, cell nucleus) | NO | Confocal microscopy | [177] |
| pDNA (pGFP and pluc) | YOYO-1 (red, pDNA) | Chlorpromazine, wortmannin, genistein, Me-β-CD, baf-A1, nocodazol, and aphidicolin | Confocal microscopy (internalization) and luminescence plate reader (pluc inhibition) | [187] |
| pDNA (pBMP-2) | Alex Fluor 594 conjugated dextran (red, MCMs) and pGFP (green) | Chlorpromazine, Me-β-CD, and amiloride | Fluorescence microscopy (internalization and pGFP inhibition) | [190] |
| pDNA (pBMP-2) | NO | Sucrose, LY294002, Me-β-CD and amiloride | ELISA (pBMP-2 inhibition) | [110] |
| siRNA (siRUNX2) | TAMRA (red, siRNA) | NO | TAMRA positive by FCM and fluorescence microscopy | [185] |
| pDNA (pBMP-2 and pTGF-β3) | Cy3 (red, pDNA), Alexa 488 Phalloidin (green, actin filaments) and DAPI (UV, cell nucleus) | NO | Confocal microscopy | [19] |
| miRNA (antmiR-138) | Cy3 (red, miRNA) and DAPI (UV, cell nucleus) | NO | Fluorescence microscopy | [184] |
| siRNA (siNoggin) | Cy3 (red, siRNA) | NO | Fluorescence microscopy | [118] |
| pDNA (pGFP and pluc) | Rhodamine B (red, peptide) and Hoechst 33342 (UV, cell nucleus) | NO | Fluorescence microscopy | [189] |
| pDNA (pGFP) | Rhodamine B (red, polyamide), Lysotracker Green DND-26 (green, endosomes) and DAPI (UV, cell nucleus) | Chlorpromazine, genistein, Me-β-CD, and amiloride | Confocal microscopy (internalization) and FCM (pGFP inhibition) | [113] |
| pDNA (pOSX-GFP) | Texas red (red, PEI) and DAPI (UV, cell nucleus) | NO | Confocal microscopy | [174] |
| miRNA | Dy547 (red, miRNA) and Calcein-AM (green, cytoplasm) | NO | Fluorescence microscopy | [207] |
| miRNA (antmiR-138) | Cy3 (red, miRNA) and Hoechst 33342 (UV, cell nucleus) | NO | Fluorescence microscopy | [117] |
| miRNA (miR335-5p) | Cy5 (red, pDNA), Atto 565 dye (pink-red, MNPs), FluoReporter Oregon Green® (green, PEI), and DAPI (UV, cell nucleus) | NO | Confocal microscopy | [66] |
| miRNA (antmiR-138) | Cy3 (red, miRNA) and DIO (green, cell membrane) | NO | Confocal microscopy | [116] |
| pDNA (pTGF-β1) | NO | Chlorpromazine | Luminescence plate reader (pluc) | [135] |
| pDNA (psox9) | Cy5 (red, pDNA), RITC (red, NPs) and FITC (green, PEI-PLGA nanoparticles) | NO | Confocal microscopy | [27] |
Abbreviations: miRNA micro-RNA, UV ultraviolet, pDNA plasmid DNA, BMP-2 bone morphogenic protein-2, FITC fluorescein isothiocyanate, PEI polyethyleneimine, DAPI 4', 6-diamidino-2-phenylindol, OSX osterix, GFP green fluorescent protein, Cy3 cyanine 3, luc luciferase, Me-β-CD methyl-β-cyclodextrin, baf-A1 bafilomycin A1, MCM mineral-coated microparticles, RUNX2 runt-related transcription factor 2, Cy5 cyanine 5, TGF-β1 and -β3 transforming growth factor-β1 and -β3, sox9 sex-determining region Y-type high mobility group box 9, RITC rhodamine isothiocyanate, NPs nanoparticles, PLGA poly (DL-lactic-co-glycolic acid)