Journal: Nanomaterials

Article Title: Exosomal Surface Protein Detection with Quantum Dots and Immunomagnetic Capture for Cancer Detection

doi: 10.3390/nano11071853

Figure Lengend Snippet: Schematic of the QD-based EXO assay ( a ), and characterization of EXOs ( b ), and QDs ( c – e ). EXOs were captured from biofluids with MB via CD81 monoclonal antibodies. Targeted surface cancer marker was recognized with primary antibody and then detected with secondary antibody-conjugated QD655. Signals were measured with fluorescence spectroscopy to quantify the QDs and correspondingly the surface protein markers on EXOs. ( b ) SEM image of plasma exosomes from a BC patient. ( c ) Absorption spectrum and ( d ) emission spectrum of IgG-QD655. ( e ) DLS characterization of the hydrodynamic size of IgG-QD655 and MB.

Article Snippet: Anti-rabbit CD81 antibody was purchased from Boster Biological Technology (Pleasanton, CA, USA).

Techniques: Bioprocessing, Marker, Fluorescence, Spectroscopy, Clinical Proteomics

Journal: Frontiers in Immunology

Article Title: Gemcitabine-cisplatin chemotherapy plus anti-PD-L1 therapy reinvigorates antitumor immune response by reprogramming the intrahepatic cholangiocarcinoma microenvironment

doi: 10.3389/fimmu.2025.1666393

Figure Lengend Snippet: Differential analysis of CD8 + T cells between untreated and GCP-treated ICC samples. (A) UMAP plots showing all T cells labeled in different colors according to cell type annotation. (B) Volcano plot showing the differentially expressed genes of intra-tumoral CD8 + T cells between pretreated and posttreated ICC samples. (C) Immunofluorescence images showing the infiltration of CD8 + CD81 + T cells in treatment-naïve ICC samples from the TMA cohort (n=89). Kaplan–Meier survival curves for OS (D) and RFS (E) of 89 ICC patients grouped by infiltration levels of CD8 + CD81 + T cells. P values were determined via log-rank test. (F) Boxplot showing infiltration levels of CD8 + CD81 + T cells in GCP-treated tumors revealed by multiplex immunofluorescence in GCP cohort. (G) Violin plots showing the expression scores of tissue resident, exhausted, and co-stimulatory gene signatures in tumor-infiltrating CD8 + T cells between pretreated and posttreated samples. The P values were calculated by Wilcox test. OS, overall survival; RFS, recurrence-free survival. ****p < 0.0001.

Article Snippet: After that, the slides were incubated with CD81 primary antibody (1:200 dilution, A01281-2, BOSTER) and Alexa Fluor 488 donkey anti-rabbit antibody as described above.

Techniques: Labeling, Immunofluorescence, Multiplex Assay, Expressing

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Constitutive SoxS Expression in a Fluoroquinolone-Resistant Strain with a Truncated SoxR Protein and Identification of a New Member of the marA-soxS-rob Regulon, mdtG ▿

doi: 10.1128/AAC.00944-09

Figure Lengend Snippet: Western blot analysis of PS5 and NorE5 using antibodies against AcrB and TolC.

Article Snippet: The membranes were blocked using 1× PBS containing Tween 20 diluted 1/2,000 (PBS-T) and 5% skim milk for 1 h at room temperature, followed by an overnight incubation at 4°C with the primary antibodies against the AcrB and TolC proteins (Antibody Bcn, Barcelona, Spain) diluted 1/500 in PBS-T.

Techniques: Western Blot

Journal: Viruses

Article Title: Single MVA-SARS-2-ST/N Vaccination Rapidly Protects K18-hACE2 Mice against a Lethal SARS-CoV-2 Challenge Infection

doi: 10.3390/v16030417

Figure Lengend Snippet: Construction and in vitro characterization of MVA-N and MVA-ST/N. ( A ) Schematic diagram of the MVA genome with the major deletion sites I to VI. MVA-SARS-2-N (MVA-N): The intergenomic region 069R and 070L of the MVA genome was targeted by inserting the gene sequence encoding the N-protein of SARS-CoV-2 from the virus isolate Wuhan HU-1 under the control of the vaccina virus promoter PmH5. Repetitive sequences served to remove the GFP-marker gene by intragenomic homologous recombination (marker gene deletion) to generate MVA-N. MVA-SARS-2-ST/N (MVA-ST/N): The site of deletion III was targeted by inserting the gene sequence encoding the stabilized S-protein (ST) of SARS-CoV-2 under transcriptional control of the vaccinia virus promoter PmH5. Repetitive sequences served to remove the mCherry marker gene by intragenomic homologous recombination (marker gene deletion) to generate MVA-SARS-2-ST (MVA-ST) . The N-protein of SARS-CoV-2 was inserted into MVA-ST as described for MVA-N to generate MVA-ST/N. ( B ) Multiple-step growth analysis of recombinant MVA-N and MVA-ST/N. CEF or DF-1 cells as well as human HaCat cells were infected at a multiplicity of infection (MOI) of 0.1 with MVA, MVA-N, or MVA-ST/N and samples were collected at the indicated time points. Samples were titrated on CEF monolayers, and plaque-forming units (PFUs) were determined. Differences between the groups were analyzed, determining the area under the curve (AUC) prior to analysis by Kruskal–Wallis Test. ( C , D ) Synthesis of the SARS-2-ST and SARS-2-N protein by MVA-N and MVA-ST/N. ( C ) Western blot analysis of SARS-2-S2 and SARS-2-N in lysates of MVA-ST/N-infected cells indicating synthesis of S- and N-protein and Western blot analysis of SARS-2-N in lysates of MVA-N-infected cells indicating synthesis of N-protein. DF-1 cells were infected with an MOI of 5 with mock control, MVA, MVA-N, or MVA-ST/N and collected 0, 4, 8, 12, 24, and 48 h post infection (hpi). Polypeptides in cell lysates were detected using a monoclonal antibody against SARS-2-S2 and SARS-2-N. ( D ) Western blot analysis of SARS-2-S2 and SARS-2-N in lysates of MVA-N- and MVA-ST/N-infected cells indicating synthesis of S- and N-protein. Analysis of Hsp90 served as positive control. DF-1 cells were infected with an MOI of 5 with mock control, MVA, MVA-N, or MVA-ST/N and collected 24 h post infection (hpi). Polypeptides in cell lysates were detected using a monoclonal antibody against SARS-2-S2 and SARS-2-N. ( E ) Double immunofluorescent staining for SARS-2-N and SARS-2-S2 in MVA-N- and MVA-ST/N-infected Vero cells (MOI = 0.1; 17 hpi). Non-infected Vero cells (mock) and cells infected with non-recombinant MVA (MVA) served as controls. Permeabilized infected cells were probed with a monoclonal antibody directed against SARS-2-N or the S2-domain of SARS-CoV-2. Polyclonal goat anti-mouse antibody or polyclonal goat anti-rabbit antibody served to visualize red and green fluorescence for N-specific (green) and S-specific (red) fluorescence staining. Cell nuclei were counterstained with DAPI (blue). The yellowish color in the merge image indicates the overlapping of the red (S-protein) and green (N-protein) colors. Scale bar: 50 μm.

Article Snippet: The blots were blocked in a phosphate buffered saline buffer (PBS) containing 5% non-fat dried milk powder (PanReac AppliChem, Darmstadt, Germany) and 0.1% Tween20 (Sigma-Aldrich, Taufkirchen, Germany) and were incubated overnight with primary antibodies targeting the S2-domain of the S-protein (1A9, 1:4000; GeneTex, Irvine, CA, USA) or the N-protein (HL249, 1:4000; GeneTex, Irvine, CA, USA) of SARS-CoV-2.

Techniques: In Vitro, Sequencing, Virus, Control, Marker, Homologous Recombination, Recombinant, Infection, Western Blot, Positive Control, Staining, Fluorescence