General descriptive statistics of sample population

Flow analysis of medium/large STB-EVs in the 10K STB-EV pellet. Apogee beads mix were used to set the flow machine’s light scatter resolution to 0.59-1.3 µm and 0.24-1.3 µm silica beads(A). Figure B shows the application of SSC and FSC PMTVs as determined by apogee beads mix for the analysis of m/lSTB-EVs in the 10K pellet. An investigation gate was created to include only medium/large EVs negative for non-placental markers (C & F). Extracellular vesicles from the investigation gate were further analyzed for staining by bioM and expression of PLAP (D & G). Figure E and H shows that the bioM+ PLAP+ EVs were sensitive to detergent treatment. The percent of B-Maleimide+ PLAP+ from the 0.59-1.3 µm gate is consistent under both SSC conditions.

Results of STB-EV characterization. Figure A,B,C and D displays representative transmission electron microscopy (TEM) images with wide view (A and C), medium/large STB-EVs (B), and Small STB-EVs (D).

Results of STB-EV characterization. The western blot characterization of S STB-EVs and mlSTB-EVs (A). 10K refers to mlSTB-EVs and 150K refers to s STB-EVs. CYT C refers to cytochrome c. Figure 2A and 2B show the NTA results of m/lSTB-EVs (B) and sSTB-EVs (C).

Top ten differentially expressed Proteins between normal and preeclampsia placentas (bold font), medium/large STB-EVs (normal font), small STB-EVs(italics)

List of overlapping differentially carried proteins in the placenta, medium/large STB-EVs and small STB-EVs

Figure showing the results of western blot and densitometric verification of PAPP-A2, COL17A1, FLNB and SR-BI in the placenta (A and D) m/lSTB-EVs (B and E) and sSTB-EVs (C and F). Collagen 17A1 did not show up in the placenta homogenate. * means < 0.05, ** means < 0.01, *** means < 0.001, **** means less than 0.001 and ns means non-significant.

Flow chart illustrating the steps involved in characterizing m/lSTB-EVs and sSTB-EVs obtained via differential ultracentrifugation (10,000 and 150,000 g) of maternal perfusate. Nanoparticle trafficking analysis, transmission electron microscopy, flow cytometry, and western blot were used to characterize the pellets from both spins.

Reagents used for western blot.

Antibodies used for western blot.

The top three functionally enriched gene ontologies are: biological process (GO: BP) Placenta (bold font), medium/large STB-EVs (normal font), and small STB-EVs (italics).

The top three functionally enriched KEGG Pathways in the placenta, medium/large STB-EVs, and small STB-EVs (only 1 KEGG). Placenta (bold font), medium/large STB-EVs (normal font), and small STB-EVs (italics)