Journal: Signal Transduction and Targeted Therapy

Article Title: Enhanced potency of an IgM-like nanobody targeting conserved epitope in SARS-CoV-2 spike N-terminal domain

doi: 10.1038/s41392-024-01847-8

Figure Lengend Snippet: Interpretation of N235/MN235-mediated neutralization mechanism. a The overall features of N235 bound to the Omicron sub-variant BA.1 S trimer. The NTD/N235 complex was superimposed onto the S trimer (PDB code: 7wz1), which contains one “up” RBD and two “down” RBDs. The S monomer that clashed with N235 is colored in slate. b Comparison of mean fluorescence intensity (MFI). The GFP-fused S proteins were transiently expressed on the surface of BHK-21 cells and stained with His-tagged N235 or CV3-13 Fab in 10, 30, and 100 μg/mL, respectively. The complex proteins were then incubated with RBD-targeting antibody CB6, human ACE2 (hACE2), S2-targeting antibody 76E1, and NTD-targeting antibody 4A8. Experiments were performed twice, and one representative was displayed. c Comparisons of MFI. The GFP-fused S proteins were transiently expressed on the surface of BHK-21 cells and stained with RBD-targeting antibody CB6, S2-targeting antibody 76E1, and NTD-targeting antibody 4A8 before incubation with His-tagged N235 or CV3-13 Fab in 10, 30, and 100 μg/mL, respectively. The vertical axis represents the level of S1 shedding, calculated by dividing the MFI of cells with surface-expressed S treated with CB6, hACE2, 4A8 and 76E1 by the MFI of the 76E1-treated group. Cells were gated based on the FSC-A and SSC-A (P1) as shown in Supplementary Fig. . d Western blot analysis of S1 subunits. The S proteins were transiently expressed on the surface of 293T cells and incubated with His-tagged N235 or CV3-13 Fab in 1, 10, and 100 μg/mL, respectively. PBS was negative control. The supernatants were collected after 1 h and detected via Western blot assay using an anti-SARS-CoV-2 S1 polyclonal antibody. Experiments were performed twice, and one representative was displayed. e Possible mechanism of N235-mediated neutralization: N235 binding triggers shedding off S1 subunits from S trimers, rending viruses non-infectious

Article Snippet: Immunoblotting was performed using an anti-SARS-CoV-2 S1 polyclonal antibody (1:2000 dilution; Sino Biological, Cat#40150-T62) followed by incubation with horseradish peroxidase (HRP)-conjugated goat anti-rabbit secondary antibody (1:10000 dilution; Gene-Protein Link, Cat# P12S12S).

Techniques: Neutralization, Variant Assay, Comparison, Fluorescence, Staining, Incubation, Western Blot, Negative Control, Binding Assay

Journal: Signal Transduction and Targeted Therapy

Article Title: Enhanced potency of an IgM-like nanobody targeting conserved epitope in SARS-CoV-2 spike N-terminal domain

doi: 10.1038/s41392-024-01847-8

Figure Lengend Snippet: The prophylactic efficacy of MN235 and N235 against SARS-CoV-2 pseudovirus in vivo. a 6–8 week-old female BALB/c mice were infected by i.n. with Ad5‑hACE2. Five days later, mice were administered N235, MN235 and control antibody by i.n. before challenged with luciferase-expressing SARS-CoV-2 Omicron XBB and BA.1 pseudovirus ( n = 4 per group). Mice were imaged 24 h after pseudovirus infection. b The bioluminescence signal in the nasal passage was quantified. Data are presented as mean ± s.d. Student’s t -test was used to analyze differences between groups. * P < 0.05

Article Snippet: Immunoblotting was performed using an anti-SARS-CoV-2 S1 polyclonal antibody (1:2000 dilution; Sino Biological, Cat#40150-T62) followed by incubation with horseradish peroxidase (HRP)-conjugated goat anti-rabbit secondary antibody (1:10000 dilution; Gene-Protein Link, Cat# P12S12S).

Techniques: In Vivo, Infection, Luciferase, Expressing

Journal: Open Medicine

Article Title: Cholesterol induces inflammation and reduces glucose utilization

doi: 10.1515/med-2023-0701

Figure Lengend Snippet: Primer sequence list of RT-qPCR

Article Snippet: The cells were stained with anti-glucose-6-phosphatase catalytic subunit 2 (G6PC2) antibody (catalog number: CSB-PA873624LA01HU, 1:100; CUSABIO) overnight at 4°C and stained an hour later at 37°C in the dark with Alexa Fluor® 488-labeled goat anti-rabbit IgG (catalog number: GB25303, 1:100; Servicebio).

Techniques: Sequencing

Journal: Open Medicine

Article Title: Cholesterol induces inflammation and reduces glucose utilization

doi: 10.1515/med-2023-0701

Figure Lengend Snippet: Antibody information for western blotting and immunohistochemistry

Article Snippet: The cells were stained with anti-glucose-6-phosphatase catalytic subunit 2 (G6PC2) antibody (catalog number: CSB-PA873624LA01HU, 1:100; CUSABIO) overnight at 4°C and stained an hour later at 37°C in the dark with Alexa Fluor® 488-labeled goat anti-rabbit IgG (catalog number: GB25303, 1:100; Servicebio).

Techniques: Western Blot

Journal: Open Medicine

Article Title: Cholesterol induces inflammation and reduces glucose utilization

doi: 10.1515/med-2023-0701

Figure Lengend Snippet: Effect of cholesterol on glucose utilization by beta-TC-6 cells. (a) After beta-TC-6 cells were treated with cholesterol, the glucose content in the cell culture supernatant was detected. (b) After the beta-TC-6 cells were treated with cholesterol, the G6PC2 mRNA level was detected using RT-qPCR. (c) After the beta-TC-6 cells were treated with cholesterol, the G6PC2 protein level was detected using an immunofluorescence experiment. (d) Statistical analysis of green fluorescence in (c). “MOCK” indicates the control group, * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001.

Article Snippet: The cells were stained with anti-glucose-6-phosphatase catalytic subunit 2 (G6PC2) antibody (catalog number: CSB-PA873624LA01HU, 1:100; CUSABIO) overnight at 4°C and stained an hour later at 37°C in the dark with Alexa Fluor® 488-labeled goat anti-rabbit IgG (catalog number: GB25303, 1:100; Servicebio).

Techniques: Cell Culture, Quantitative RT-PCR, Immunofluorescence, Fluorescence, Control

Journal: Open Medicine

Article Title: Cholesterol induces inflammation and reduces glucose utilization

doi: 10.1515/med-2023-0701

Figure Lengend Snippet: Effects of cholesterol on mice pancreas. (a) After the mice were treated with cholesterol, the lesions of the pancreatic tissue were observed using hematoxylin–eosin staining. (b) After the mice were treated with cholesterol, the insulin content in their blood was detected by ELISA. (c) After the mice were treated with cholesterol, the glucose content in their blood was detected. (d) Immunofluorescence was used to observe the expression of the G6PC2 protein in mice pancreas after they were treated with cholesterol. (e) Statistical analysis of green fluorescence in (d). “MOCK” indicates the control group, * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001.

Article Snippet: The cells were stained with anti-glucose-6-phosphatase catalytic subunit 2 (G6PC2) antibody (catalog number: CSB-PA873624LA01HU, 1:100; CUSABIO) overnight at 4°C and stained an hour later at 37°C in the dark with Alexa Fluor® 488-labeled goat anti-rabbit IgG (catalog number: GB25303, 1:100; Servicebio).

Techniques: Staining, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Expressing, Fluorescence, Control

Journal: Open Medicine

Article Title: Cholesterol induces inflammation and reduces glucose utilization

doi: 10.1515/med-2023-0701

Figure Lengend Snippet: Effects of cholesterol on the inflammatory response and the expression of endoplasmic reticulum-related molecules in mice pancreas. (a) After the mice were treated with cholesterol, G6PC2, GRP78, NLRP3, casp1, and IL-1β mRNA levels were detected using RT-qPCR. (b) After the mice were treated with cholesterol, GRP78, G6PC2, NLRP3, pro-casp1, and p20 (cleaved casp1) protein levels were detected by western blot, using GAPDH as an internal reference. (c) Statistical analysis of the western blot experiments in (b). (d) After the mice were treated with cholesterol, GRP78, NLRP3, casp1, and IL-1β protein levels were detected via immunohistochemistry. (e) Statistical graph of immunohistochemistry analysis in (d). “MOCK” indicates the control group, * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001.

Article Snippet: The cells were stained with anti-glucose-6-phosphatase catalytic subunit 2 (G6PC2) antibody (catalog number: CSB-PA873624LA01HU, 1:100; CUSABIO) overnight at 4°C and stained an hour later at 37°C in the dark with Alexa Fluor® 488-labeled goat anti-rabbit IgG (catalog number: GB25303, 1:100; Servicebio).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Immunohistochemistry, Control

Journal: iScience

Article Title: Calpain-mediated cleavage generates a ZBTB18 N-terminal product that regulates HIF1A signaling and glioblastoma metabolism

doi: 10.1016/j.isci.2022.104625

Figure Lengend Snippet:

Article Snippet: Rabbit polyclonal anti-CAPN2 , Cell Signaling Technology , Cat#2539S; RRID: AB_2069843.

Techniques: Virus, Recombinant, Purification, Mutagenesis, Caspase-Glo Assay, Reverse Transcription, Sequencing, Extraction, Gene Expression, Western Blot, Software, Microscopy