Journal: Tissue antigens

Article Title: Direct binding to antigen-coated beads refines the specificity and cross-reactivity of four monoclonal antibodies that recognize polymorphic epitopes of HLA class I molecules

doi: 10.1111/tan.12095

Figure Lengend Snippet: (A) HLA class I allotypes represented by the One Lambda Labscreen and Gen-Probe LifeCodes beadsets. (B) Binding of the monomorphic HLA class I antibody W6/32 to beads from One Lambda LabScreen (grey bars) and Gen-Probe LifeCodes (orange bars). The allotypes shown are those common to both beadsets.

Article Snippet: The specific reference as listed in the current publication is noted to the right. fig ft0 fig mode=article f1 fig/graphic|fig/alternatives/graphic mode="anchored" m1 Open in a separate window Figure 2 caption a7 (A) HLA class I allotypes represented by the One Lambda Labscreen and Gen-Probe LifeCodes beadsets. (B) Binding of the monomorphic HLA class I antibody W6/32 to beads from One Lambda LabScreen (grey bars) and Gen-Probe LifeCodes (orange bars).

Techniques: Binding Assay

Journal: Tissue antigens

Article Title: Direct binding to antigen-coated beads refines the specificity and cross-reactivity of four monoclonal antibodies that recognize polymorphic epitopes of HLA class I molecules

doi: 10.1111/tan.12095

Figure Lengend Snippet: (A) Binding of MA2.1 (1μg/ml) to beads coated with HLA class I allotypes from the One Lambda LabScreen (left panel) and Gen-Probe LifeCodes (right panel) beadsets. (B) Alignment of HLA class I allotypes showing selected residues in the α1 and α2 domains. Residues from allotypes that form the epitope recognized by MA2.1 are shaded in grey. (C) Space-filling model of the binding surface of HLA-A*02 (grey) with associated peptide (cyan). Residues highlighted in yellow fall within the footprint recognized by MA2.1. Residues 62–65 are critical for formation of the epitope recognized by MA2.1 and are highlighted in red.

Article Snippet: The specific reference as listed in the current publication is noted to the right. fig ft0 fig mode=article f1 fig/graphic|fig/alternatives/graphic mode="anchored" m1 Open in a separate window Figure 2 caption a7 (A) HLA class I allotypes represented by the One Lambda Labscreen and Gen-Probe LifeCodes beadsets. (B) Binding of the monomorphic HLA class I antibody W6/32 to beads from One Lambda LabScreen (grey bars) and Gen-Probe LifeCodes (orange bars).

Techniques: Binding Assay

Journal: Tissue antigens

Article Title: Direct binding to antigen-coated beads refines the specificity and cross-reactivity of four monoclonal antibodies that recognize polymorphic epitopes of HLA class I molecules

doi: 10.1111/tan.12095

Figure Lengend Snippet: (A) Binding of PA2.1 (1μg/ml) and BB7.2 (1μg/ml) to beads coated with HLA class I allotypes from the One Lambda LabScreen (left panel) and Gen-Probe LifeCodes (right panel) beadsets. (B) Binding of PA2.1 (50μg/ml) and BB7.2 (50μg/ml) to beads coated with HLA class I allotypes from the One Lambda LabScreen (left panel) and Gen-Probe LifeCodes (right panel) beadsets. (C) Alignment of HLA class I allotypes showing selected residues in the α2 domain. Residues from allotypes that form the epitope recognized by PA2.1 and BB7.2 are shaded in grey. (D) Space-filling model of HLA-A*02 (grey) with associated peptide (cyan). Residues highlighted in yellow fall within the footprint recognized by PA2.1 and BB7.2. Tryptophan at position 107 is considered critical for formation of the epitope recognized by PA2.1 and BB7.2 and is highlighted in red.

Article Snippet: The specific reference as listed in the current publication is noted to the right. fig ft0 fig mode=article f1 fig/graphic|fig/alternatives/graphic mode="anchored" m1 Open in a separate window Figure 2 caption a7 (A) HLA class I allotypes represented by the One Lambda Labscreen and Gen-Probe LifeCodes beadsets. (B) Binding of the monomorphic HLA class I antibody W6/32 to beads from One Lambda LabScreen (grey bars) and Gen-Probe LifeCodes (orange bars).

Techniques: Binding Assay

Journal: Tissue antigens

Article Title: Direct binding to antigen-coated beads refines the specificity and cross-reactivity of four monoclonal antibodies that recognize polymorphic epitopes of HLA class I molecules

doi: 10.1111/tan.12095

Figure Lengend Snippet: (A) Binding of BB7.1 (1μg/ml) to beads coated with HLA class I allotypes from the One Lambda LabScreen (left panel) and Gen-Probe LifeCodes (right panel) beadsets. (B) Alignment of HLA class I allotypes showing selected residues in the α1 and α2 domains. Residues from allotypes that form the epitope recognized by BB7.1 are shaded in grey. (C) Space-filling model of the binding surface of HLA-B*07 (grey) with associated peptide (cyan). Residues highlighted in yellow fall within the footprint recognized by BB7.1. Residues 63–71 in the a1 domain and position 131 in the α2 domain are critical for formation of the epitope recognized by BB7.1 and are highlighted in red.

Article Snippet: The specific reference as listed in the current publication is noted to the right. fig ft0 fig mode=article f1 fig/graphic|fig/alternatives/graphic mode="anchored" m1 Open in a separate window Figure 2 caption a7 (A) HLA class I allotypes represented by the One Lambda Labscreen and Gen-Probe LifeCodes beadsets. (B) Binding of the monomorphic HLA class I antibody W6/32 to beads from One Lambda LabScreen (grey bars) and Gen-Probe LifeCodes (orange bars).

Techniques: Binding Assay

Journal: bioRxiv

Article Title: Immuno-proteomic interrogation of dengue infection reveals novel HLA haplotype-specific MHC-I antigens

doi: 10.1101/471821

Figure Lengend Snippet: a) Length distribution of pMHC repertoire in control (blue) and DENV infected – DENV2+ (orange) Raji cells. b) Summed abundance (normalized to total protein abundance) of tryptic peptides from MHC-I HLA-A, HLA-B, and HLA-C proteins in the proteome of control (blue) and DENV infected – DENV2+ (orange) cells. Error bars represent standard deviations across two biological replicates. c) FACS staining MHC-1 with the pan-MHC-I antibody (clone W6/32) in uninfected control (blue) and DENV infected Raji cells DENV2+ (orange). FACS trace for the isotype control is marked in green.

Article Snippet: Immunoprecipitation of the pre-cleared supernatant was performed using a pan-MHC-I antibody (W6/32, Genentech) coupled to Protein-A sepharose beads, on a rotating platform for 12 hrs at 4°C.

Techniques: Infection, Staining

Journal: Frontiers in Immunology

Article Title: T cell interaction with activated endothelial cells primes for tissue-residency

doi: 10.3389/fimmu.2022.827786

Figure Lengend Snippet: EC-induced CD69 expression on memory CD8+ T cells is partly mediated by synergistic action of IL-15, ICAM-1 and VCAM-1. (A+B+C) HMEC were left unstimulated (resting) or stimulated with 10 ng/mL TNFα and/or IFNγ for 3 days (activated) before addition of FACS-sorted memory CD3+ T cells to the co-culture or HMEC cultured medium. CD69 expression and proliferation were assessed at various time points of the co-culture. (A) Percentage of CD69+ cells (left panel) and median fluorescent intensity (MFI) of CD69 (right panel) within CD8+ T cells after co-culture with TNFα and IFNγ-stimulated HMEC, resting HMEC, their cultured medium (sup), or TNFα and IFNγ alone (cyt only). N=3, mean+SEM. 2-Way-ANOVA with Sidak post-hoc test. */#/ 0 P < 0.05, **/##/ 00 P < 0.01, ***/###/ 000 P < 0.001,****/####/ 0000 P < 0.0001. (B) Percentage of CD69+ cells (left panel) and median fluorescent intensity (MFI) of CD69 (right panel) within CD8+ T cells after culture with activated HMEC cultured medium (sup), transwell co-culture or direct co-culture with activated HMEC. N = 5, median. 2-Way ANOVA with Sidak’s multiple comparison test; *p < 0.05, ns = not significant. (C) Percentage of CD69+ cells (left panel) and median fluorescent intensity (MFI) of CD69 (right panel) within CD8+ T cells after co-culture with TNFα- and/or IFNγ-stimulated HMEC or their cultured medium (sup). N = 3, mean+SEM. 2-Way-ANOVA with Sidak post-hoc test. */#/ 0 P < 0.05, **/##/ 00 P < 0.01, ***/###/ 000 P < 0.001,****/####/ 0000 P < 0.0001. (D+E) Levels of soluble ICAM-1 and VCAM-1 (D) and IL-15 and TGF-β (E) measured in cultured medium of resting or TNFα- and/or IFNγ-stimulated HMEC after 3 days, by multiplex immunoassay. N = 3, mean+SEM. Kruskal-Wallis with Dunn’s post-hoc test versus resting EC. *P < 0.05, **P < 0.01. (F+G) Co-culture of TNFα and IFNγ-stimulated HMEC with FACS-sorted memory CD3+ T cells in the presence of (increasing concentrations) of monoclonal antibodies blocking IL-15, TGF-β, ICAM-1 and/or VCAM. (F) The percentage of CD69+ cells was measured by flow cytometry after 18 hours and normalized to the percentage of CD69+ cells in the condition with isotype control (set to 100). N = 3, median. Kruskal-Wallis with Dunn’s post-hoc test versus isotype. (G) The percentage of CD69+ cells was measured by flow cytometry after 18 and 90 hours and normalized to the condition with isotype control (set to 100). N = 4, median. 2-Way-ANOVA with Sidak post-hoc test versus isotype. *P < 0.05, **P < 0.01. (H) Transwell and direct co-culture of TNFα and IFNγ-stimulated HMEC with FACS-sorted memory CD3+ T cells in the presence of monoclonal antibodies blocking IL-15, ICAM-1 and/or VCAM. Percentage of CD69+ cells within CD8+ T cells was measured by flow cytometry after 90 hours and normalized to the condition with isotype control (set to 100). N = 4, median. 2-Way-ANOVA with Sidak post-hoc test versus isotype. (I) Co-culture of TNFα and IFNγ-stimulated HMEC with FACS-sorted memory CD3+ T cells in the presence of 35 μg/mL monoclonal antibody blocking HLA-ABC or isotype control. The percentage of CD69+ cells and median fluorescent intensity (MFI) of CD69 was measured by flow cytometry after 18 and 90 hours. N = 4. Dotted lines indicate paired measurements. 2-Way-ANOVA with Sidak post-hoc test versus isotype. *P < 0.05, ***P < 0.001.

Article Snippet: MHC-I interactions were blocked with an antibody against HLA-ABC (W6/32, 35 μg/mL, Bioceros) .

Techniques: Expressing, Co-Culture Assay, Cell Culture, Comparison, Multiplex Assay, Bioprocessing, Blocking Assay, Flow Cytometry, Control