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Figure 1. Characterization of APS-Lat-EXO and its ability to induce latent HIV-1 reactivation in J-Lat 10.6 cells. (A) Representative transmission electron microscopy images showing the morphology of isolated exosomes (scale bar, 100 nm); (B) Nanoparticle tracking analysis demonstrating the size distribution of APS-Lat-EXO, with a major peak around 105 nm; (C) Detection of exosomal surface markers (CD9-FITC, CD63-PE, CD81-APC) by Amnis imaging flow cytometry; (D) Expression of exosomal marker proteins (CD9, CD63, and CD81) confirmed by Western blotting; (E) Uptake of PKH-26-labeled exosomes by J-Lat 10.6 cells, visualized using Amnis imaging flow cytometry; (F) Representative fluorescence microscopy images (20×) showing GFP expression in J-Lat 10.6 cells following treatment; (G and H) Flow cytometric quantification of GFP-positive cell populations, with boxed areas indicating GFP⁺ cells; (I) Summary analysis of the proportion of GFP-expressing cells; (J) HIV-1 p24 levels in culture supernatants measured by ELISA (100-fold dilution); (K and L) Relative expression levels of Gag and LTR transcripts in cells and culture supernatants assessed by RT-qPCR. APS was applied at 800 μg/mL, while APS-Lat-EXO and Lat-EXO were used at 15 μg/mL. PHA (positive control) was used at 5 μg/mL. Data are presented as mean ± SD from three independent experiments (n = 3). Statistical significance was determined using Tukey’s multiple comparisons test. ^*P < 0.05; ^**P < 0.01; ^***P < 0.001; ^****P < 0.0001. APS: Astragalus polysaccharide; EXO: exosome; APS-Lat-EXO: APS-modified latency-reversing exosome; HIV-1: human immunodeficiency virus type 1; J-Lat: Jurkat latency model; TEM: transmission electron microscopy; NTA: nanoparticle tracking analysis; FITC: fluorescein isothiocyanate; PE: phycoerythrin; APC: allophycocyanin; Amnis IFC: Amnis imaging flow cytometry; GFP: green fluorescent protein; ELISA: enzyme-linked immunosorbent assay; RT-qPCR: reverse transcription quantitative polymerase chain reaction; PHA: phytohemagglutinin; SD: standard deviation; CD9: cluster of differentiation 9; CD63: cluster of differentiation 63; CD81: cluster of differentiation 81; PKH26: Paul Karl Horan 26; p24: HIV-1 capsid protein p24; Gag: group-specific antigen; LTR: long terminal repeat; BF: bright field.