Journal: eLife

Article Title: IFNγ induces epigenetic programming of human T-bethi B cells and promotes TLR7/8 and IL-21 induced differentiation

doi: 10.7554/elife.41641

Figure Lengend Snippet: Figure 2. ASC development from BN precursors is enhanced in Th1 containing co-cultures. Cartoon (a) depicting day 6 paired co-cultures containing Th1 (Be1 co-cultures) or Th2 (Be2 co-cultures) effectors generated from the same HD, BN cells from a second allogeneic HD and exogenous IL-21 and IL-2. Flow cytometric analysis showing T-bet expression (b) on gated HD B cells (non-ASCs) from Be1 and Be2 co-cultures. Phenotyping (c) of day 6 B cell-gated Be1 cells showing T-bet expression in combination with other surface markers. (d–g) ASC development in HD day 6 paired Be1 and Be2 co-cultures showing representative flow plots (d) and frequencies (e) of CD38hiCD27+ ASCs in CD19+/lo-gated B lineage cells. Frequencies of IgM+ (f) or IgG+ (g) ASCs in day 6 paired Be1 and Be2 co-cultures. See Figure 2—figure supplement 1 for BN isolation strategy and characterization of polarized Th1 and Th2 effectors. See Figure 2—figure supplement 2 for gating strategy to identify IgG+ and IgM+ ASCs. See Figure 2—figure supplement 3 for proliferation analysis of B cells in paired day 6 HD Be1 and Be2 co-cultures. Analyses in (b–c) are from representative co-cultures (n > 30). Experiments (e–g) performed on 15 (e), 8 (f) or 6 (g) independent paired Be1 and Be2 co-cultures. Statistical analyses were performed using a non- parametric Wilcoxon paired t test (e) or paired Student’s t test (f–g). P values *<0.05, **<0.01, ****<0.0001. DOI: https://doi.org/10.7554/eLife.41641.005 The following figure supplements are available for figure 2:

Article Snippet: DOI: https://doi.org/10.7554/eLife.41641 23 of 36 Continued Reagent type (species) or resource Designation Source or reference Identifiers Additional information Antibody PercP/Cy5.5 Mouse Anti-Human CD4 (clone OKT4) eBioscience 45-0048-42 (1:200) Antibody BV510 Mouse Anti-Human CD4 (clone OKT4) Biolegend 317444 (1:100) Antibody Fitc Mouse Anti-Human CD11c (clone Bu15) Biolegend 337214 (1:200) Antibody PE Mouse Anti-Human CD11c (clone Bu15) Biolegend 337205 (1:400) Antibody PercP/Cy5.5 Mouse Anti-Human CD14 (clone HCD14) Biolegend 325621 (1:200) Antibody Fitc Mouse Anti-Human CD19 (clone LT19) Miltenyi 302256 (1:100) Antibody PE Mouse Anti-Human CD19 (clone HIB19) Biolegend 302208 (1:200) Antibody PercP/Cy5.5 Mouse Anti-Human CD19 (clone HIB19) Biolegend 302230 (1:100) Antibody APC Mouse Anti-Human CD19 (clone HIB19) BD Pharmingen 555415 (1:200) Antibody APC-H7 Mouse Anti-Human CD19 (clone HIB19) BD Pharmingen 560727 (1:100) Antibody BV421 Mouse Anti-Human CD19 (clone HIB19) Biolegend 302234 (1:200) Antibody V500 Mouse Anti-Human CD19 (clone HIB19) BD Horizon 561121 (1:100) Antibody PercP/Cy5.5 Mouse Anti-Human CD21 (clone Bu32) Biolegend 354908 (1:100) Antibody Fitc Mouse Anti-Human CD23 (clone M-L23.4) Miltenyi 130-099-365 (1:100) Antibody PE Mouse Anti-Human CD23 (clone EBVCS-5) Biolegend 338507 (1:200) Antibody APC Mouse Anti-Human CD23 (clone M-L233) BD Pharmingen 558690 (1:200) Antibody Fitc Mouse Anti-Human CD27 (clone M-T271) Biolegend 356404 (1:100) Antibody PercP/Cy5.5 Mouse Anti-Human CD27 (clone M-T271) Biolegend 356408 (1:100) Antibody APC Mouse Anti-Human CD27 (clone M-T271) Biolegend 356410 (1:200) Continued on next page Zumaquero et al. eLife 2019;8:e41641.

Techniques: Generated, Expressing, Isolation

Journal: Arteriosclerosis, thrombosis, and vascular biology

Article Title: Naive CD8 + T Cells Expressing CD95 Increase Human Cardiovascular Disease Severity

doi: 10.1161/ATVBAHA.120.315106

Figure Lengend Snippet: CyTOF Antibody List.

Article Snippet: CD19 , HIB19 , 142Nd , Fluidigm , Fluidigm.

Techniques: Conjugation Assay

Journal: PLoS Pathogens

Article Title: Inhibition of polo-like kinase 1 (PLK1) facilitates reactivation of gamma-herpesviruses and their elimination

doi: 10.1371/journal.ppat.1009764

Figure Lengend Snippet: (A) Total RNAs were extracted from the fixed diffuse large B-cell lymphoma (DLBCL) tissue samples of HIV-negative (HIV-, n = 3) and HIV-positive (HIV+, n = 5) individuals. mRNA level of PLK1 in these tissue samples was measured by RT-qPCR and normalized to GAPDH. (B) A PE-conjugated CD19 antibody was used for B cell enrichment from PBMCs of HIV-infected, aviremic patients (HIV+, n = 5) or healthy donors (HIV-, n = 5). Purity of isolated CD19 + B cells was confirmed by flow cytometry analysis. (C) mRNA level of PLK1 in above isolated B cells (B) was measured by RT-qPCR and normalized to GAPDH. (D) Protein level of intracellular PLK1 in above isolated B cells (B) was measured by immunofluorescence. Percentage of PLK1-expressing cells was determined by flow cytometry analysis. (F) HIV-1 IIIB infected Jurkat cells were co-cultured with BJAB cells that were pre-stained with CFSE. Protein level of intracellular PLK1 in CFSE-labeled BJAB cells was measured by immunofluorescence. Percentage of PLK1-expressing cells was determined by flow cytometry analysis. (G-I) Protein level of PLK1 in BJAB (G), Akata-BX1 (H), or TREx BCBL1-Rta (I) cells treated with recombinant Nef (rNef) protein at the increasing dose or DMSO was measured by immunoblotting. GAPDH was used as loading control. Results were calculated from n = 3 independent experiments and presented as mean ± SD (* p<0.05; two-tailed paired Student t-test).

Article Snippet: Portion of isolated B cells was incubated with CD19 antibody (1/200 of stock, Milteny) for 30 mins to determine the purity using BD Accuri C6 Plus with corresponding optical filters.

Techniques: Quantitative RT-PCR, Infection, Isolation, Flow Cytometry, Immunofluorescence, Expressing, Cell Culture, Staining, Labeling, Recombinant, Western Blot, Control, Two Tailed Test

Journal: Advanced Science

Article Title: Bispecific Nanosystems Enable Multieffector Immune Cell Retargeting for Hematologic Malignancy Therapy

doi: 10.1002/advs.202509103

Figure Lengend Snippet: A) This diagram depicts the dual‐targeting strategy of biHSNPs in facilitating tumor cell elimination. Each biHSNP features two functional arms: one specifically binds to a tumor‐associated antigen on lymphoma cells (CD19/CD20), while the other engages and activates immune effector cells (CD3/CD16). By bridging tumor cells and immune cells, this approach enhances immune‐mediated tumor cell lysis, leveraging the immune system's inherent ability to identify and eliminate abnormal cells for precise and efficient cancer cell killing. B) Schematic illustration of the T cell activation and tumor cell death mechanism mediated by biHSNPs.

Article Snippet: FITC anti‐human CD19 antibody[CB19] (catalog No.: E‐AB‐F1004C, species reactivity: human), FITC anti‐human CD20 antibody[BCA/B20] (Catalog No.: E‐AB‐F1045C, species reactivity: human) and FITC anti‐human CD3 antibody[OKT‐3] (catalog No.: E‐AB‐F1001C, species reactivity: human) were obtained from Elabscience Biotechnology Co.,Ltd. (China, Wuhan).

Techniques: Functional Assay, Lysis, Activation Assay

Journal: Advanced Science

Article Title: Bispecific Nanosystems Enable Multieffector Immune Cell Retargeting for Hematologic Malignancy Therapy

doi: 10.1002/advs.202509103

Figure Lengend Snippet: Schematic illustration of the chemical synthesis and characterization of antibody‐conjugated hollow silica nanoparticles. A) Synthetic pathway of PEGylated hollow silica nanoparticles. B) Fourier transform‐infrared spectroscopy (FT‐IR) characterization of HSNP@PEG (2). C) Proton nuclear magnetic resonance ( 1 H NMR) characterization of HSNP@PEG. D) DLS and TEM images of HSNP@PEG. E) DLS and TEM images of HSNP@PEG@NH 2 (3). F) Fluorescence spectrum of HSNP@PEG@NH 2 @FITC (9.62 µ m ), confirming successful FITC conjugation. G) DLS and TEM images of HSNP@PEG@NH 2 @FITC (4). H–K) DLS and TEM results for monospecific nanoparticles (HSNP αCD3 , HSNP αCD16 , HSNP αCD19 , HSNP αCD20 ). L–O) DLS and TEM results for bispecific nanoparticles (HSNP αCD3 + αCD19 , HSNP αCD3 + αCD20 , HSNP αCD16 + αCD19 , HSNP αCD16 + αCD20 ). All scale bars in the TEM images represent 100 nm. HSNP αCD3 , HSNP αCD16 , HSNP αCD19 and HSNP αCD20 : Monospecific HSNPs conjugated with a single type of antibody (anti‐CD3, anti‐CD16, anti‐CD19, or anti‐CD20, respectively). HSNP αCD3 + αCD19 , HSNP αCD3 + αCD20 , HSNP αCD16 + αCD19 and HSNP αCD16 + αCD20 : Bispecific HSNPs conjugated with two different types of antibodies (e.g., anti‐CD3 and anti‐CD19). Antibody types are indicated using lowercase Greek letters (e.g., αCD3).

Article Snippet: FITC anti‐human CD19 antibody[CB19] (catalog No.: E‐AB‐F1004C, species reactivity: human), FITC anti‐human CD20 antibody[BCA/B20] (Catalog No.: E‐AB‐F1045C, species reactivity: human) and FITC anti‐human CD3 antibody[OKT‐3] (catalog No.: E‐AB‐F1001C, species reactivity: human) were obtained from Elabscience Biotechnology Co.,Ltd. (China, Wuhan).

Techniques: Fourier Transform Infrared Spectroscopy, Spectroscopy, Nuclear Magnetic Resonance, Fluorescence, Conjugation Assay

Journal: Advanced Science

Article Title: Bispecific Nanosystems Enable Multieffector Immune Cell Retargeting for Hematologic Malignancy Therapy

doi: 10.1002/advs.202509103

Figure Lengend Snippet: Binding specificity and cell–cell linkage induced by biHSNPs. Target cells were incubated with FITC‐labeled, antibody‐coated HSNPs at 4 °C for 30 min, followed by flow cytometry analysis to confirm binding specificity. A–C) Depict cell–cell linkage induced by biHSNPs: A) Schematic illustration of cell‐cell complex formation mediated by biHSNPs. B) Confocal microscopy images showing stable junctional complexes between immune effector cells and tumor cells facilitated by HSNP αCD3 + αCD19 and HSNP αCD3 + αCD20 . C) Flow cytometry analysis of cell–cell complexes between pre‐stained Jurkat (green, CellTracker Green CMFDA) and Raji cells (red, CellTracker Red CMTPX), demonstrating effective linkage by biHSNPs. D) Verification of biHSNPs binding to CD3 + cells using PBMCs as the CD3 + cell line. E) Verification of biHSNPs binding to CD19 + /CD20 + cells using Raji cells as the target. F) Verification of biHSNPs binding to CD16 + cells using PBMCs as the CD16 + cell line. G) Verification of biHSNPs binding to CD19 + /CD20 + cells using Raji cells as the target.

Article Snippet: FITC anti‐human CD19 antibody[CB19] (catalog No.: E‐AB‐F1004C, species reactivity: human), FITC anti‐human CD20 antibody[BCA/B20] (Catalog No.: E‐AB‐F1045C, species reactivity: human) and FITC anti‐human CD3 antibody[OKT‐3] (catalog No.: E‐AB‐F1001C, species reactivity: human) were obtained from Elabscience Biotechnology Co.,Ltd. (China, Wuhan).

Techniques: Binding Assay, Incubation, Labeling, Flow Cytometry, Confocal Microscopy, Staining

Journal: Advanced Science

Article Title: Bispecific Nanosystems Enable Multieffector Immune Cell Retargeting for Hematologic Malignancy Therapy

doi: 10.1002/advs.202509103

Figure Lengend Snippet: A) Cytotoxicity and cytokine release assays of biHSNPs in luciferase‐transfected Raji cells. B) In vitro cytotoxicity of HSNP@PEG, HSNP αCD3 , HSNP αCD19 , and bispecific HSNP αCD3 + αCD19 . C) In vitro cytotoxicity of HSNP@PEG, HSNP αCD3 , HSNP αCD20 , and bispecific HSNP αCD3 + αCD20 . D,E) Cytokine secretion levels (IFN‐γ, IL‐2, and TNF‐α) for HSNP αCD3 , HSNP αCD19 , and HSNP αCD3 + αCD19 ; and HSNP αCD20 and HSNP αCD3 + αCD20 . F) In vitro cytotoxicity of HSNP@PEG, HSNP αCD16 , HSNP αCD19 , and bispecific HSNP αCD16 + αCD19 . G) In vitro cytotoxicity of HSNP@PEG, HSNP αCD16 , HSNP αCD20 , and bispecific HSNP αCD16 + αCD20 . H,I) Cytokine secretion levels (IFN‐γ, IL‐2, and TNF‐α) for HSNP αCD16 , HSNP αCD19 , HSNP αCD16 + αCD19 , HSNP αCD20 , and HSNP αCD16 + αCD20 . J) Cytotoxicity of combined bispecific nanoparticle treatments: HSNP αCD3 + αCD19 with HSNP αCD16 + αCD19 , and HSNP αCD3 + αCD20 with HSNP αCD16 + αCD20 . K) In vitro cytotoxicity of HSNP αCD3 + αCD19 combined with HSNP αCD16 + αCD19 , and HSNP αCD3 + αCD20 combined with HSNP αCD16 + αCD20 . Data are presented as mean ± SD ( n ≥ 3), Statistical differences in B–I) were analyzed by One‐way ANOVA and K) were analyzed by Student's t‐test and the statistical significance is indicated as ** P ≤ 0.01 and *** P ≤ 0.001.

Article Snippet: FITC anti‐human CD19 antibody[CB19] (catalog No.: E‐AB‐F1004C, species reactivity: human), FITC anti‐human CD20 antibody[BCA/B20] (Catalog No.: E‐AB‐F1045C, species reactivity: human) and FITC anti‐human CD3 antibody[OKT‐3] (catalog No.: E‐AB‐F1001C, species reactivity: human) were obtained from Elabscience Biotechnology Co.,Ltd. (China, Wuhan).

Techniques: Luciferase, Transfection, In Vitro

Journal: Advanced Science

Article Title: Bispecific Nanosystems Enable Multieffector Immune Cell Retargeting for Hematologic Malignancy Therapy

doi: 10.1002/advs.202509103

Figure Lengend Snippet: Enhanced T‐cell activation mediated by biHSNPs. A) Representative Quantitative analysis of CD19 + and CD8 + cell populations based on IHC. D,F) Representative immunofluorescence (IF) images showing the distribution of CD19 + Raji cells and CD8 + T cells. E–G) Quantitative analysis of CD19 + and CD8 + cells based on IF staining. H–J) Representative flow cytometry plots illustrating; I) Ki67 + expression in Raji cells and K) CD107a expression in CD8 + T cells across treatment groups. Statistical differences are analyzed by using One‐way ANOV and statistical significance is indicated as * P ≤ 0.05, ** P ≤ 0.01 and **** P ≤ 0.0001.

Article Snippet: FITC anti‐human CD19 antibody[CB19] (catalog No.: E‐AB‐F1004C, species reactivity: human), FITC anti‐human CD20 antibody[BCA/B20] (Catalog No.: E‐AB‐F1045C, species reactivity: human) and FITC anti‐human CD3 antibody[OKT‐3] (catalog No.: E‐AB‐F1001C, species reactivity: human) were obtained from Elabscience Biotechnology Co.,Ltd. (China, Wuhan).

Techniques: Activation Assay, Immunofluorescence, Staining, Flow Cytometry, Expressing