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. 2019 Aug 2;20(15):3789. doi: 10.3390/ijms20153789

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© 2019 by the authors.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

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Comparative analysis of lectin-specific glycosignatures in Mel202-derived ectosomes. Reactions with biotinylated (A) PHA-L and PHA-E, (C) SNA and MAA, and (E) AAA and GNA, where lectins are shown in the upper panels and controls with blocking solution of competitive sugars or CH₃COOH (as indicated in Supplementary Data 1) are shown in the lower panels. Seventy micrograms of protein were loaded per lane. Detection was performed via nitro-blue tetrazolium and 5-bromo-4-chloro-3′-indolyphosphate (NBT/BCIP) staining. H—the whole cell extract, M—membrane fraction, E—ectosomes. Densitometric analysis of (B) PHA-L- and PHA-E-, (D) SNA- and MAA-, and (F) AAA- and GNA-positive bands. The intensity of detected bands was expressed in arbitrary units (AU) as a peak area on a respective densitogram. Each bar represents the intensity of corresponding bands in the whole cell extract, membrane fraction, and ectosomes from Mel202 cells. * denotes bands that were enriched in ectosomes, relative to the membrane fraction. The values from control lectin blots with blocking solutions were subtracted beforehand.

Comparative analysis of lectin-specific glycosignatures in Mel202-derived ectosomes. Reactions with biotinylated (A) PHA-L and PHA-E, (C) SNA and MAA, and (E) AAA and GNA, where lectins are shown in the upper panels and controls with blocking solution of competitive sugars or CH₃COOH (as indicated in Supplementary Data 1) are shown in the lower panels. Seventy micrograms of protein were loaded per lane. Detection was performed via nitro-blue tetrazolium and 5-bromo-4-chloro-3′-indolyphosphate (NBT/BCIP) staining. H—the whole cell extract, M—membrane fraction, E—ectosomes. Densitometric analysis of (B) PHA-L- and PHA-E-, (D) SNA- and MAA-, and (F) AAA- and GNA-positive bands. The intensity of detected bands was expressed in arbitrary units (AU) as a peak area on a respective densitogram. Each bar represents the intensity of corresponding bands in the whole cell extract, membrane fraction, and ectosomes from Mel202 cells. * denotes bands that were enriched in ectosomes, relative to the membrane fraction. The values from control lectin blots with blocking solutions were subtracted beforehand.