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. 2021 Oct 14;24(11):103282. doi: 10.1016/j.isci.2021.103282

Figure 2.

Figure 2

Generation and characterization of XP-GFP and XP-GFP-DNAJB6 sEVs

(A) XP-GFP and XP-GFP-DNAJB6 constructs used in this study. XP-tagged proteins are directed into sEVs.

(B) Representative images of donor cells expressing XP-GFP or XP-GFP-DNAJB6. Wild-type cells serve as control. XP-tagged proteins colocalize with CD9, an MVB marker protein. The insets show enlarged view of boxed areas. DAPI was used for nuclear staining.

(C) Size distribution of sEVs measured by NTA.

(D) Western blot analysis of whole-cell and purified sEV lysates (20 μg protein per sample per lane). XP-tagged GFP and GFP-DNAJB6 proteins are enriched in sEV fractions as compared with respective donor cells. WT sEVs isolated from untransfected cells serve as a control. Endogenous DNAJB6b is present in cell lysates and at low amounts in sEV fractions. Note the high amount of DNAJB6 present in XP-GFP-DNAJB6 sEV fraction. SEV marker Lamp2b is present in all sEV samples at similar levels. The endoplasmic reticulum marker protein calnexin is nearly absent in all sEV fractions. XP: XPack; sEV: purified sEV lysate; Cell: whole-cell lysate. Scale bar: 10 μm.