Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Targeting antigens to CD180 but not CD40 programs immature and mature B cell subsets to become efficient antigen-presenting cells

doi: 10.4049/jimmunol.1900549

Figure Lengend Snippet: (A) B1-8hi mice were inoculated with 50μg NP-Iso, NP-Iso + αCD180 or NP-αCD180. 24 h later, the expression of MHC II and CD86 was evaluated on NP-BCR− and NP-BCR+ splenic B cells by flow cytometry. Data is the mean ± SEM representative of two independent experiments with n=3-4 per group. (B) B10.A mice were inoculated with 35μg HEL-Iso, HEL-Iso + αCD180 or HEL-αCD180. Three or 24 h later HEL-peptide bound-MHC II I-Ak, represented as peptide bound MHC II over total MHC II expression (HEL-I-Ak/I-Ak MFI) was evaluated on splenic B cell subsets and conventional (c)DCs by flow cytometry. cDCs were defined as B220−, CD3−, NK1.1−, CD11b−/lo, CD11chi. Data is the mean ± SEM representative of two independent experiments per time point with n=3-4 per group. (C) C57BL/6 mice were inoculated with 35μg OVA-Iso, OVA-Iso + αCD180 or OVA-αCD180. Three or 24 h later DCs were isolated from spleen by positive bead selection, B cells were isolated by negative bead enrichment and B cell subsets were sorted by FACS. 1×105 APCs were co-cultured with 1×10^5 CFSE-labeled OT-II T cells. Three days later, T cell proliferation was evaluated as CFSE dilution by flow cytometry. Data is the mean ± SEM combined of three experiments per time point with n=1 (Mock, OVA-Iso) or n=6 (OVA-Iso + αCD180 and OVA-αCD180). * p<0.05, ** p<0.01, ***p<0.001, ****p<0.0001 by 2-way ANOVA with Sidak’s multiple comparison test (A), Kruskal-Wallis test with Dunn’s multiple comparison test (B) or Mann-Whitney test (C).

Article Snippet: Meanwhile, CD4 + T cells (Ly5.1 + ) were isolated from splenocytes from OT-II mice by magnetic bead negative selection (StemCell Technologies) and labeled with CFSE as above.

Techniques: Expressing, Flow Cytometry, Isolation, Selection, Cell Culture, Labeling, Comparison, MANN-WHITNEY

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Targeting antigens to CD180 but not CD40 programs immature and mature B cell subsets to become efficient antigen-presenting cells

doi: 10.4049/jimmunol.1900549

Figure Lengend Snippet: (A,B) C57BL/6 mice were inoculated with 35μg OVA-αCD40 or OVA-αCD180; 24h later DCs were isolated from spleen by positive bead selection, B cells were isolated by negative bead enrichment and B cell subsets were sorted by FACS. 1×105 APCs were co-cultured with 1×105 CFSE-labeled OT-II T cells. Three days later, T cell proliferation was evaluated as CFSE dilution by flow cytometry. The division index (B, left) is the average number of divisions that a cell has undergone. Data is the mean ± SEM combined of three experiments with n=4 (OVA-αCD40) or n=6 (OVA-αCD180). (C) C57BL/6 mice were inoculated with 25μg OVA-αCD40 or OVA-αCD180; 3d later OVA-tetramer-specific cells were isolated from splenocytes by magnetic bead enrichment. Total CD44+, Tetramer+ (left) and Tetramer+, CXCR5+, PD-1+ TFH cells (right) were analyzed by flow cytometry. Data is mean ± SEM combined from three independent experiments, n=12 per group. (D,E) C57BL/6 mice were inoculated with 50μg NP-Iso, NP-αCD40 or NP-αCD180. Once a week, mice were bled and NP-specific IgG (D, left) and the ratio of high affinity NP-specific IgG (NP2) to total NP-specific IgG (NP20) (D, right) was quantitated by ELISA. Nine weeks after inoculation, spleens and bone marrow were harvested and NP-specific IgG secreting antibody secreting cells (ASCs) were evaluated by ELISPOT (E). Data is mean ± SEM combined from two independent experiments, n=8 per group. * p<0.05, ** p<0.01, ***p<0.001 by Mann-Whitney test (A, B, C, D right), Kruskal-Wallis test with Dunn’s multiple comparison test (E) or 2-way ANOVA with Sidak’s multiple comparison test (F left).

Article Snippet: Meanwhile, CD4 + T cells (Ly5.1 + ) were isolated from splenocytes from OT-II mice by magnetic bead negative selection (StemCell Technologies) and labeled with CFSE as above.

Techniques: Isolation, Selection, Cell Culture, Labeling, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Enzyme-linked Immunospot, MANN-WHITNEY, Comparison

Journal: Frontiers in Oncology

Article Title: Combined treatment with epigenetic agents enhances anti-tumor activity of MAGE-D4 peptide-specific T cells by upregulating the MAGE-D4 expression in glioma

doi: 10.3389/fonc.2022.873639

Figure Lengend Snippet: P8-specific T cells inhibited the in vivo growth of U87 cell-derived tumors. (A) Tumor growth was suppressed by the adoptive transfer of P8-specific T cells. (B) Tumors harvested from mice that were inoculated with different groups of T cells and glioma cells. (C) CD8 + T cells infiltrated glioma tissue in vivo . DC, dendritic cell; Tc, T cell; NC, negative control peptide. *P <0.05, **P <0.01, scale bar: 50 μm.

Article Snippet: CD8 + T cells were isolated from PBMCs using CD8 + negative selection MicroBeads EasySepTM Human CD8 + T Cell Isolation Kit (17953, stemcell) according to the manufacturers.

Techniques: In Vivo, Derivative Assay, Adoptive Transfer Assay, Negative Control