cryo tem micrographs  (JEOL)

Journal: Biomaterials Research

Article Title: Autotaxin-Scavenging Nanoliposomes for Prolonged Colon Retention and Autophagy-Mediated Mucosal Immune Restoration in Colitis

doi: 10.34133/bmr.0345

Figure Lengend Snippet: Characterization and ATX-scavenging activity of AS-Lipo@R. (A) Cryo-TEM images of Lipo and AS-Lipo (scale bar, 100 nm). (B) Particle size and zeta potential of Lipo and AS-Lipo measured by DLS. (C) Colloidal stability of Lipo and AS-Lipo was evaluated by monitoring changes in particle size and zeta potential over 72 h in PBS using DLS. (D) Concentration-dependent inhibition of recombinant mouse ATX activity as a function of BMP-22 concentration in free BMP-22, Lipo, and AS-Lipo, evaluated using a choline release assay. (E) ATX binding to the surface of Lipo and AS-Lipo as a function of ATX concentration, quantified by ELISA. (F) HPLC chromatograms showing rapamycin (R) encapsulation in AS-Lipo@R, as indicated by the disappearance of the free R peak. (G) Cumulative release profiles of rapamycin from Lipo@R and AS-Lipo@R in PBS over 72 h measured by HPLC. (H) Fluorescence imaging showing colocalization of DiO-labeled Lipo or AS-Lipo (green), Alexa Fluor 647 (AF647)-labeled ATX (red), and LysoTracker (purple) in RAW 264.7 macrophages. AS-Lipo-bound ATX is internalized and colocalizes with lysosomes, indicating lysosomal degradation. Right panels show fluorescence intensity profiles along the indicated lines, quantifying the colocalization (scale bar, 100 and 50 μm). Statistical significance was determined by one-way ANOVA with Tukey’s post hoc test ( n = 3). * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001.

Article Snippet: Cryo-TEM micrographs were acquired using a JEM-2100F electron microscope (JEOL, Japan).

Techniques: Activity Assay, Zeta Potential Analyzer, Concentration Assay, Inhibition, Recombinant, Release Assay, Binding Assay, Enzyme-linked Immunosorbent Assay, Encapsulation, Fluorescence, Imaging, Labeling

Journal: Acta Pharmaceutica Sinica. B

Article Title: A rationally designed cancer vaccine based on NIR-II fluorescence image-guided light-triggered remote control of antigen cross-presentation and autophagy

doi: 10.1016/j.apsb.2022.11.027

Figure Lengend Snippet: NIR-II fluorescence imaging of BPBBT-labeled liposomes (BPBBT-Lipo) for lymph node delivery. (A–C) Size distribution of BPBBT-Lipo in different sizes measured by dynamic light scattering (DLS). Insets: the cryo-transmission electron micrographs (TEMs) of their respective BPBBT-Lipo, scale bar, 25 nm of (A), 100 nm of (B), 200 nm of (C). (D) Fluorescence emission spectra of BPBBT-Lipo (10 μmol/L) in water excited at 830 nm. (E) Live NIR-II fluorescence imaging of the lymphatic drainage of BPBBT-Lipo in different sizes at various timepoints following s.c. injection in the footpad of mice. Left, photographs; Right, NIR-II images, scale bar = 1 cm. The course of the lymphatic drainage of OVA-BPBBT-Lipo 150 following the injection (0–2 h) was also recorded . (F) Quantification of average fluorescence intensity of BPBBT-Lipo in the paws or the PLNs from Supporting Information Fig. S2 , respectively ( n = 3). (G) The distribution of BPBBT-Lipo 150 loading FITC-labeled OVA (OVA-FITC-BPBBT-Lipo 150 ) in different cells of the PLNs after the s.c. administration for 1.5 h ( n = 4). (H) Intravital fluorescence microscopic images of OVA-BPBBT-Lipo 150 in DCs in the PLN at different timepoints following the s.c. injection of the liposomes . Green, the NIR-II fluorescence of OVA-BPBBT-Lipo 150 . Red, DCs in the PLN of mice following the intravenous injection of APC-labeled anti-mouse CD11c antibody at 1 h before the administration of the liposomes, scale bar = 50 μm. Data are expressed as mean ± SD.

Article Snippet: The morphology of liposomes was observed via cryo-transmission electron micrograph (Cryo-TEM, FEI Tecnai G2 F20, 200 kV, FEI, Hillsboro, OR, USA).

Techniques: Fluorescence, Imaging, Labeling, Transmission Assay, Injection

Journal: Acta Pharmaceutica Sinica. B

Article Title: A rationally designed cancer vaccine based on NIR-II fluorescence image-guided light-triggered remote control of antigen cross-presentation and autophagy

doi: 10.1016/j.apsb.2022.11.027

Figure Lengend Snippet: The enhanced MHC I cross-presentation pathway through photothermally induced endolysosomal escape. (A) Cryo-TEM of OVA-BPBBT-Lipo 150 , scale bar = 100 nm. (B) Cellular uptake of OVA-FITC or OVA-FITC-loaded liposomes (OVA-FITC-Lipo 150 ) in DC2.4 cells was analyzed by flow cytometry ( n = 3). (C) Fluorescence micrograph of liposomes loading both Cy5-labeled OVA (OVA-Cy5) and BPBBT (OVA-Cy5-BPBBT-Lipo 150 ) in DC2.4 cells. The cells treated with free OVA-Cy5 plus BPBBT-Lipo 150 (OVA-Cy5 + BPBBT-Lipo 150 ) were used as control, scale bar = 20 μm. (D) Fluorescence micrograph of liposomes loading OVA-FITC and BPBBT (OVA-FITC-BPBBT-Lipo 150 ) in the PLNs of mice at 1.5 h post-injection, scale bar = 50 μm. (E) Flow cytometric analysis of the SIINFEKL presentation by DCs in the PLNs of mice after injection with various formulations for 24 h ( n = 3). In the OVA + BPBBT-Lipo 150 group, the mice were s.c. injected with free OVA and BPBBT-Lipo 150 , respectively. (F) Temperature–time curve of H 2 O, OVA-Lipo 150 or OVA-BPBBT-Lipo 150 (50 μmol/L of BPBBT) in water irradiated with laser (808 nm, 1 W/cm 2 ) for 10 min, followed by another 10 min of cooling down. Δ T represents the increase of the temperature. (G) TEM of DC2.4 cells incubated with OVA-BPBBT-Lipo 150 for 2 h with or without laser irradiation (808 nm, 1 W/cm 2 , 5 min). Yellow arrows, intact endolysosomes; red arrows, endolysosomes with ruptured membrane, scale bar = 500 nm. (H) Immunofluorescence micrographs of the intracellular distribution of OVA-FITC in the PLNs after the s.c. injection of OVA-FITC-BPBBT-Lipo 150 for 1.5 h with or without laser treatment. In the laser-treated group, the PLNs of mice were visualized with NIR-II fluorescence imaging under the anesthetic condition at 1.5 h post-injection. Then, the PLNs were irradiated with the 808 nm laser (1 W/cm 2 , 10 min) under the imaging guidance, scale bar = 5 μm. (I) Flow cytometric analysis of the SIINFEKL presentation by DCs in the PLNs after the injection with various formulations for 24 h ( n = 3). The procedure of laser treatment was the same as (H). Data are expressed as mean ± SD. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, ∗∗∗∗ P < 0.0001 vs . indicated.

Article Snippet: The morphology of liposomes was observed via cryo-transmission electron micrograph (Cryo-TEM, FEI Tecnai G2 F20, 200 kV, FEI, Hillsboro, OR, USA).

Techniques: Flow Cytometry, Fluorescence, Labeling, Injection, Irradiation, Incubation, Immunofluorescence, Imaging