Journal: Nature Communications

Article Title: An in-situ peptide-antibody self-assembly to block CD47 and CD24 signaling enhances macrophage-mediated phagocytosis and anti-tumor immune responses

doi: 10.1038/s41467-024-49825-6

Figure Lengend Snippet: a Schematic illustration of tumor immune quiescent microenvironment, in situ assembly of PAC-SABIs on the cancer cell surface, and blockage of CD47 and CD24 phagocytic checkpoints by PAC-SABIs. Figure was created with BioRender.com and released under a Creative Commons Attribution-NonCommercial-NoDerivs 4.0 international license. b Time-dependent CLSM images of 4T1 cells treated with NBD-labeled PAC-SABIs. Scale bar: 20 μm. Three independent experiments were performed. c Time-dependent CLSM images of PAN02 cells treated with NBD-labeled PAC-SABIs. Scale bar: 20 μm. Three independent experiments were performed. d Merged bright field and fluorescence images of 4T1 cell treated with NBD-labeled PAC-SABIs for 120 min. Scale bar: 20 μm. Three independent experiments were performed. e CLSM images of 4T1 cell treated with Dil dye (red) and NBD-labeled PAC-SABIs for 120 min. Scale bar: 10 μm. Three independent experiments were performed. f , g Fluorescence distribution of NBD-labeled PAC-SABIs on 4T1 cell. Three independent experiments were performed. h Time-dependent SEM images of 4T1 cells treated with PAC-SABIs. The red arrows point to PAC-SABIs on cell membrane. Scale bar: 2 μm. Three independent experiments were performed. i Time-dependent SEM images of PAN02 cells treated with PAC-SABIs. The red arrows point to the PAC-SABIs on cell membrane. Scale bar: 2 μm. Three independent experiments were performed. j CLSM images of 3D 4T1 spheroids treated with Cy5.5 PAC-SABIs, Calcein-AM, and Hoechst 33342. Scale bar: 50 μm. Three independent experiments were performed. k CLSM images of 3D 4T1 spheroids along the z -axis position. Scale bar: 50 μm. Three independent experiments were performed.

Article Snippet: Treatment: anti-mouse CD24 antibody (BioXcell, #BE0360, clone M1/69), anti-human CD24 antibody (Novus Biologics, #NB100-64861, clone SN3), anti-mouse CD47 antibody (BioXcell, #BE0270, clone MIAP301), anti-mouse PD-1 antibody (BioXcell, #BE0273, clone 29F.1A12TM), anti-CSF1R antibody (BioXcell, #BE0213, clone AFS98).

Techniques: In Situ, Labeling, Fluorescence, Membrane

Journal: Nature Communications

Article Title: An in-situ peptide-antibody self-assembly to block CD47 and CD24 signaling enhances macrophage-mediated phagocytosis and anti-tumor immune responses

doi: 10.1038/s41467-024-49825-6

Figure Lengend Snippet: a Phagocytosis images of pHrodo-red + over time. Scale bar: 50 μm. Three independent experiments were performed. b Phagocytosis of 4T1 cells, in the presence of IgG control or PAC-SABIs. The error bars represent the mean ± SD ( n = 3 independent experiments). c Representative 3D CLSM image reconstruction of in vitro phagocytosis of 4T1 cells (pHrodo-red + , red) by BMDMs and RAW264.7 cells (Calcein-AM, green). Three independent experiments were performed. d Representative flow cytometry plots depicting BMDM phagocytosis of 4T1 cells treated with IgG control, anti-CD24 mAb, SAMIs and PAC-SABIs. e Quantitative analysis of BMDM flow cytometry results. The error bars represent the mean ± SD ( n = 3 independent experiments; ** p < 0.01, *** p < 0.001; the p value was analyzed by a two-tailed unpaired Student’s t test). f Representative flow cytometry plots depicting RAW264.7 cell phagocytosis of 4T1 cells treated with IgG control, anti-CD24 mAb, SAMIs and PAC-SABIs. g Quantitative analysis of RAW264.7 cell flow cytometry results. The error bars represent the mean ± SD ( n = 3 independent experiments; ** p < 0.01; the p value was analyzed by a two-tailed unpaired Student’s t test). Source data are provided as a Source Data file.

Article Snippet: Treatment: anti-mouse CD24 antibody (BioXcell, #BE0360, clone M1/69), anti-human CD24 antibody (Novus Biologics, #NB100-64861, clone SN3), anti-mouse CD47 antibody (BioXcell, #BE0270, clone MIAP301), anti-mouse PD-1 antibody (BioXcell, #BE0273, clone 29F.1A12TM), anti-CSF1R antibody (BioXcell, #BE0213, clone AFS98).

Techniques: Control, In Vitro, Flow Cytometry, Two Tailed Test

Journal: ImmunoTargets and Therapy

Article Title: Exploring the Association Between Immune Cell Phenotypes and Osteoporosis Mediated by Inflammatory Cytokines: Insights from GWAS and Single-Cell Transcriptomics

doi: 10.2147/ITT.S510102

Figure Lengend Snippet: Single-Cell RNA Sequencing Analysis of Immune Cell Characteristics in a Postmenopausal Osteoporosis Patient. ( a ) Immune cells were clustered based on unique molecular markers. ( b ) B cells were extracted and further annotated, identifying the target cell population: IgD+ CD24+ B cells. ( c ) Highly expressed genes in the IgD+ CD24+ B cell population were screened, followed by gene set enrichment analysis.

Article Snippet: The cell pellet was resuspended in 100 μL of PBS containing 1% BSA (Solarbio) and stained with CD24 (FITC Anti-Mouse CD24 Antibody, E-AB-F1179C, Elabscience) and IgD (PE Anti-Mouse IgD Antibody, E-AB-F1189D, Elabscience) at 4°C for 30 min. After staining, 1 mL of PBS containing 1% BSA was added, followed by centrifugation at 300g for 5 min.

Techniques: RNA Sequencing

Journal: ImmunoTargets and Therapy

Article Title: Exploring the Association Between Immune Cell Phenotypes and Osteoporosis Mediated by Inflammatory Cytokines: Insights from GWAS and Single-Cell Transcriptomics

doi: 10.2147/ITT.S510102

Figure Lengend Snippet: IgD+CD24+ B cells and IL-17C play critical roles in osteoporosis. ( a )The micro CT results in osteoporosis group and control group. ( b and c ) The proportion of IgD+CD24+ B cells in lymphocytes of mice in osteoporosis group and control group. Horizontal axis: CD24, Vertical axis: IgD; ( d ) ELISA was used to measure IL-17C levels in the serum obtained from mouse eyeball blood; ( e ) Western blot was performed to detect IL-17C levels in mouse bone tissue. ( f ) Immunohistochemistry (IHC) was used to assess IL-17C levels in bone tissue.

Article Snippet: The cell pellet was resuspended in 100 μL of PBS containing 1% BSA (Solarbio) and stained with CD24 (FITC Anti-Mouse CD24 Antibody, E-AB-F1179C, Elabscience) and IgD (PE Anti-Mouse IgD Antibody, E-AB-F1189D, Elabscience) at 4°C for 30 min. After staining, 1 mL of PBS containing 1% BSA was added, followed by centrifugation at 300g for 5 min.

Techniques: Micro-CT, Control, Enzyme-linked Immunosorbent Assay, Western Blot, Immunohistochemistry

Journal: Cancer Cell

Article Title: Single-cell analysis defines a pancreatic fibroblast lineage that supports anti-tumor immunity

doi: 10.1016/j.ccell.2021.06.017

Figure Lengend Snippet:

Article Snippet: Anti-mouse CD24 (150Nd) clone M1/69 , Fluidigm , 3150009B.

Techniques: Blocking Assay, Virus, Recombinant, Plasmid Preparation, Saline, Lysis, Staining, Reverse Transcription, In Vivo, Electroporation, Mass Cytometry, Conjugation Assay, Illumina Sequencing, Library Quantification, In Vitro, Quantitative RT-PCR, Software, Real-time Polymerase Chain Reaction, Cytometry, Microscopy, Imaging, Spectrophotometry

Journal: Stem Cell Research & Therapy

Article Title: Lumican from DESC spheroids enhances odontoblastic differentiation of SCAPs

doi: 10.1186/s13287-025-04849-7

Figure Lengend Snippet: Characterization of SCAPs and DESCs. a Microscopic image of the apical buds from the lower incisors of 5-7-day-old C57BL/6 mice. b Primary mixed cell population. c Purified SCAPs exhibiting spindle-shaped morphology. d Multidirectional differentiation potential of SCAPs. Alcian Blue, Alizarin Red, and Oil Red O staining confirmed glycosaminoglycan deposition, mineralized nodule formation, and lipid droplet accumulation, indicative of chondrogenic, osteogenic, and adipogenic differentiation, respectively. e , f Flow cytometry analysis revealed CD24, CD133, and CD90 positivity, and CD45 negativity in SCAPs ( n = 3). The raw dot plots are presented in Supplementary Fig. S3. g Purified DESCs displaying cobblestone-like morphology. h Immunofluorescence imaging demonstrated positive K14 expression in DESCs (K14, green; DAPI, blue). i , j Flow cytometry analysis revealed CD29 and CD49f positivity, and CD90 negativity in DESCs ( n = 3). The raw dot plots are presented in Supplementary Fig. S3

Article Snippet: For SCAP analysis, cells were labeled with allophycocyanin (APC)-conjugated CD24 antibody (APC-65121, Proteintech, China, 1:300) as well as FITC-conjugated CD133 (11-1331-82, Thermo Fisher Scientific, USA, 1:500), CD90, and CD45 (MCD4501, Thermo Fisher Scientific, USA, 1:500) antibodies.

Techniques: Purification, Staining, Flow Cytometry, Immunofluorescence, Imaging, Expressing

Journal: Microbiology Spectrum

Article Title: Epitope mapping targeting the K205R protein of African swine fever virus using nanobody as a novel tool

doi: 10.1128/spectrum.01750-24

Figure Lengend Snippet: Production and identification of Nb-HRP fusion proteins, expressed and secreted, against ASFV K205R protein. The pCAGGS-HA-Nb1-, -Nb14-, -Nb35-, and -Nb82-HRP recombinant plasmids were transfected into HEK293T cells for 60 h to produce Nb-HRP fusion proteins. The HEK293T cells were fixed or collected 60 h after transfection, and the expression of Nb-HRP fusions was analyzed using ( A ) immunofluorescence assay and ( B ) western blotting with anti-HA monoclonal antibody as the primary antibody. ( C ) Cell culture supernatants were collected 60 h after transfection or not, and detection of HRP activity in the Nb-HRP fusions secreted into the cell culture supernatants was conducted using direct ELISA. Data are mean ± SD values of three independent results. * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001. Nb, nanobody; HRP, horseradish peroxidase.

Article Snippet: In a separate step, the PVDF membranes were blocked and subsequently incubated with primary antibodies for 2 h at RT, specifically, mouse anti-HA monoclonal antibody (mAb; 1:2,000; Proteintech Group, Inc.) and anti-α-tubulin (1:5,000; Proteintech Group, Inc.).

Techniques: Recombinant, Transfection, Expressing, Immunofluorescence, Western Blot, Cell Culture, Activity Assay, Direct ELISA

Journal: Microbiology Spectrum

Article Title: Epitope mapping targeting the K205R protein of African swine fever virus using nanobody as a novel tool

doi: 10.1128/spectrum.01750-24

Figure Lengend Snippet: Four Nbs, namely Nb1, Nb14, Nb35, and Nb82, bind to both prokaryotically and eukaryotically expressed K205R proteins. ( A ) SDS-PAGE analysis of prokaryotically expressed ASFV-K205R protein. ( B ) Nb1, Nb14, Nb35, and Nb82’s reaction with ASFV-K205R protein expressed with E. coli was determined using western blotting, with ASFV-DP96R used as a negative control. ( C ) Four Nbs specifically reacted with the ASFV-K205R protein, as determined by direct ELISA. Data are mean ± SD values of three independent results. **** P < 0.0001. ( D ) Titration of cell supernatant containing Nb1-, Nb14-, Nb35-, and Nb82-HRP fusion proteins binding to the ASFV-K205R protein. ( E ) Affinity of Nb1-, Nb14-, Nb35-, and Nb82-His protein toward ASFV-K205R protein. ( F ) Western blot analysis of eukaryotically expressed ASFV-K205R protein using anti-HA monoclonal antibody for detection. Four Nbs specifically reacted with the eukaryotically expressed ASFV-K205R protein, as determined using ( G ) western blotting and ( H ) immunofluorescence assay. Nb, nanobody; ASFV, African swine fever virus.

Article Snippet: In a separate step, the PVDF membranes were blocked and subsequently incubated with primary antibodies for 2 h at RT, specifically, mouse anti-HA monoclonal antibody (mAb; 1:2,000; Proteintech Group, Inc.) and anti-α-tubulin (1:5,000; Proteintech Group, Inc.).

Techniques: SDS Page, Western Blot, Negative Control, Direct ELISA, Titration, Binding Assay, Immunofluorescence, Virus

Journal: Microbiology Spectrum

Article Title: Epitope mapping targeting the K205R protein of African swine fever virus using nanobody as a novel tool

doi: 10.1128/spectrum.01750-24

Figure Lengend Snippet: Fine mapping of the epitopes in the ASFV K205R protein recognized by K205R-Nb35. ( A ) Schematic diagram of different truncated ASFV K205R protein peptides produced in a eukaryotic expression system. ( B ) Western blot analysis of the eukaryotically expressed ASFV-K205R protein with anti-HA monoclonal antibody for detection. K205R-Nb35 reacted with different eukaryotically expressed truncated K205R protein peptides, as determined by using ( C ) western blotting and ( D ) immunofluorescence assay. ( E ) Schematic diagram of the synthesis of P13, P14, P15 truncated and mutant K205R peptides. ( F ) K205R-Nb35 reacted with different truncated and mutant K205R peptides, as revealed by peptide-based ELISA. ( G ) Schematic diagram of the synthesis of different single amino acid mutant K205R peptides. ( H ) Peptide-based ELISA detection of K205R-Nb35 reacted with P16, P17, P18, and P19. Data are mean ± SD values of three independent results. **** P < 0.0001; ns, no significant. African swine fever virus; Nb, nanobody.

Article Snippet: In a separate step, the PVDF membranes were blocked and subsequently incubated with primary antibodies for 2 h at RT, specifically, mouse anti-HA monoclonal antibody (mAb; 1:2,000; Proteintech Group, Inc.) and anti-α-tubulin (1:5,000; Proteintech Group, Inc.).

Techniques: Produced, Expressing, Western Blot, Immunofluorescence, Mutagenesis, Peptide ELISA, Virus