Journal: bioRxiv

Article Title: Tissue-wide profiling of human lungs reveals spatial sequestration of macrophages in tuberculosis

doi: 10.1101/2025.04.11.648467

Figure Lengend Snippet: (A) Heatmap showing the relative abundance of CD4 + (top) and CD8 + (bottom) T cell sub-populations in each lesion type. Rows are ordered based on the similarity in marker expression patterns. N, the border of necrotic lesions. (B) Representative OPAL multi-plex image illustrating a germinal center-like structure in a B3-type lesion. The image with merged DAPI, CD20, BCL6, and PNAd is shown. Arrows indicate PNAd + high endothelial venules and arrow heads indicate BCL6 + B cells. (C) Paired analysis comparing the spatial association relationships between B cells and Tfh (C10) or proinflammatory CD4 + T cells (C08) in all B3-type lesions. Each line indicates a lesion. The level of statistical significance is determined using Wilcoxon signed-rank test, **** P < 0.0001. (D) Cell type pheno-map of a representative B3-type lesion with cell type indicated. (E) Scatter plots showing the spatial centrality index (immCPI) of Tfh (C10), proinflammatory CD4 + T cells (C08), and proinflammatory CD8 + T cells (C11) in all B3-type lesions ( n = 110). The level of statistical significance is determined using a Mann-Whitney U test, **** P < 0.0001. (F) Heatmap showing the median value of pair-wised spatial association relationships between CD4 + and CD8 + T cell subsets in B3 (left) and T (right) type lesions. (G) Column scatter plot showing spatial relationship between macrophages and CD4 + and CD8 + T cell subtypes in T-type lesions. (H) Heatmap indicating statistical significance from post hoc multi-group comparison results in (G) using a One-way ANOVA test, the level of significance was set at P = 0.05.

Article Snippet: Next, tissue sections were incubated for 1 hour at room temperature with primary antibodies against CD20 (1:100, L26, Abcam), CD4 (1:100, EPR6855, Abcam), and overnight under 4°C with BCL6 (1:100, EPR11410-43, Abcam), and PNAd (1:200, MECA-79, Novus Biologicals).

Techniques: Marker, Expressing, MANN-WHITNEY, Comparison

Journal: Scientific Reports

Article Title: Anti-IL-6 eluting immunomodulatory biomaterials prolong skin allograft survival

doi: 10.1038/s41598-019-42349-w

Figure Lengend Snippet: Local release of anti-IL-6 from IMB suppresses LN fibrosis. The skin allografts were harvested at 7 days post-transplantation. ( A-i ) Lymphatic vessel expansion (Lyve-1 + ) and HEV elongation (MECA79 + ) were similar between all groups. ( A-ii,iii ) Dense staining of collagen I and PDPN was seen in DLNs harvested from control and GelMA group compared to those from GelMA/anti-IL-6 group. (representative images from 4 different mice per group).

Article Snippet: Samples were cut into 5 μm-thick sections and stained with Lyve-1 (Abcam, #ab14917; 1:300), CD11b (Abcam, #ab8878; 1:250), CD11c (Biolegend, #101206; 1:100), CD169 (Biolegend, #142406; 1:100), CD3 (Abcam, #ab16669; 1:250), MECA79 (Novus Biologicals, #NB100-77673; 1:200), Collagen I (Novus Biologicals, #NB600-408; 1:200), and PDPN (R&D System, #AF3244; 1:200) antibodies.

Techniques: Transplantation Assay, Staining, Control

Journal: The Journal of Experimental Medicine

Article Title: Lymphocyte–HEV Interactions in Lymph Nodes of a Sulfotransferase-deficient Mouse

doi: 10.1084/jem.20030057

Figure Lengend Snippet: Intravascular staining with MECA79 and effect on in vivo homing. (A–D) MECA79 mAb was injected intravenously into HEC-GlcNAc6ST−/− or +/+ mice. 30 min later peripheral lymph nodes were dissected out and processed for immunofluorescence microscopy. Bright field images of +/+ (A) and −/− (C) lymph nodes. Intravascular staining with MECA79 of (B) +/+ and (D) −/− HEV. Magnification 200×. (E) MECA79 mAb or PBS was injected intravenously into HEC-GlcNAc6ST−/− or +/+ mice and the homing of fluorescently labeled cells was determined and normalized to produce a homing index as described in Materials and Methods. Consistent with our previous findings , homing in the HEC-GlcNAc6ST−/− mice was 59% relative to homing in wild-type.

Article Snippet: MECA79 reactivity was determined using biotinylated MECA79 (Ligocyte Pharmaceuticals).

Techniques: Staining, In Vivo, Injection, Immunofluorescence, Microscopy, Labeling

Journal: The Journal of Experimental Medicine

Article Title: Lymphocyte–HEV Interactions in Lymph Nodes of a Sulfotransferase-deficient Mouse

doi: 10.1084/jem.20030057

Figure Lengend Snippet: Effect of MECA79 mAb on lymphocyte rolling. (A) MECA79 mAb or isotype control rat IgM was injected via a femoral artery catheter into HEC-GlcNAc6ST−/− or +/+ mice. 15 min later, fluorescently labeled lymphocytes were injected. Data are based on 2 HEC-GlcNAc6ST−/− mice injected with rat IgM (1st-2; 2nd-5; 3rd-8; 4th-5; 5th-2 venules), two +/+ mice injected with rat IgM (1st-1; 2nd-2; 3rd-3; 4th-3; 5th-3 venules), two HEC-GlcNAc6ST−/− mice injected with MECA79 (1st-3; 2nd-6; 3rd-8; 4th-8; 5th-7 venules), and 2 +/+ mice injected with MECA79 (1st-2; 2nd-5; 3rd-8; 4th-8; 5th-5 venules). Bars represent mean ± SEM. (B) Fluorescently labeled L-selectin−/− lymphocytes were injected into HEC-GlcNAc6ST−/− mice treated with MECA79. Data are based on 4 HEC-GlcNAc6ST−/− mice injected with MECA79 (1st-3; 2nd-6; 3rd-8; 4th-9; 5th-7 venules) and one HEC-GlcNAc6ST−/− mouse injected with MECA79 and L-selectin−/− lymphocytes (1st-1; 2nd-3; 3rd-5; 4th-3; 5th-2 venules).

Article Snippet: MECA79 reactivity was determined using biotinylated MECA79 (Ligocyte Pharmaceuticals).

Techniques: Control, Injection, Labeling

Journal: The Journal of Experimental Medicine

Article Title: Lymphocyte–HEV Interactions in Lymph Nodes of a Sulfotransferase-deficient Mouse

doi: 10.1084/jem.20030057

Figure Lengend Snippet: 35 Sulfate incorporation and MECA79 reactivity of GlyCAM-1 and CD34. (A and D) Lymph nodes from HEC-GlcNAc6ST−/− and +/+ mice were incubated with Na 35 SO 4 . Immunoprecipitated GlyCAM-1 and CD34 were separated by SDS-PAGE and transferred to PVDF for Western blotting. (A) GlyCAM-1 was detected with an antipeptide Ab (CAMO5). (D) CD34 was detected with an affinity-purified polyclonal Ab. (B and E) 35 S-labeled immunoprecipitated ligands GlyCAM-1 (B) and CD34 (E) detected by autoradiography. (C and F) MECA79 reactivity of GlyCAM-1 (C) and CD34 (F) detected by Western blotting. (G) Relative sulfate incorporation and MECA79 reactivity of GlyCAM-1 and CD34 from HEC-GlcNAc6ST−/− mice. 35 S incorporation was measured by performing densitometry on autoradiographs as shown in B and E and normalized for protein based on Western blots performed in parallel with protein-specific Abs as in A and D. MECA79 reactivity was measured by performing densitometry on Western blots shown in C and F and normalized based on protein-specific Western blots performed in parallel. Data are the means ± SEMs of three independent preparations.

Article Snippet: MECA79 reactivity was determined using biotinylated MECA79 (Ligocyte Pharmaceuticals).

Techniques: Incubation, Immunoprecipitation, SDS Page, Western Blot, Affinity Purification, Labeling, Autoradiography

Journal: The Journal of Experimental Medicine

Article Title: Lymphocyte–HEV Interactions in Lymph Nodes of a Sulfotransferase-deficient Mouse

doi: 10.1084/jem.20030057

Figure Lengend Snippet: ELISA analyses of GlyCAM-1 from HEC-GlcNAc6ST−/− mice. Equivalent amounts of HEC-GlcNAc6ST−/− and +/+ GlyCAM-1 were coated on ELISA plates. (A) Protein was quantified using an anti–GlyCAM-1 Ab. (B) Relative MECA79 reactivity. L-selectin–IgM (C), P-selectin–IgM (D), or E-selectin–IgM (E) chimeric proteins or biotinylated-AAL (F) were allowed to bind and were subsequently detected with an alkaline phosphatase-conjugated secondary Ab. EDTA (selectins) or fucose (AAL) was used to show specificity of the respective interactions. Plots are representative of three experiments performed on two independent preparations of GlyCAM-1.

Article Snippet: MECA79 reactivity was determined using biotinylated MECA79 (Ligocyte Pharmaceuticals).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: The Journal of Experimental Medicine

Article Title: Lymphocyte–HEV Interactions in Lymph Nodes of a Sulfotransferase-deficient Mouse

doi: 10.1084/jem.20030057

Figure Lengend Snippet: Jurkat cell rolling on GlyCAM-1. (A) Equivalent amounts of HEC-GlcNAc6ST−/− and +/+ GlyCAM-1 were coated on dishes. Jurkat cells were perfused through the flow chamber and the number of rolling cells was determined for each shear stress. Values represent the mean ± SD. Addition of EDTA, mAb DREG56, or sialidase treatment of −/− and +/+ GlyCAM-1 inhibited >95% of rolling. The experiment shown is representative of three independent experiments each performed in duplicate using two different fields of view. (B) The velocity of 40 rolling cells was determined for each shear stress. Values represent the mean ± SD. (C) GlyCAM-1 from HEC-GlcNAc6ST−/− and +/+ mice was coated such that nearly equivalent numbers of rolling cells were observed at wall shear stress of 1–2 dyn/cm 2 . Dishes were treated with varying concentrations of MECA79 mAb and rolling of Jurkat T cells was determined. Data was normalized based on the number of cells that rolled in the absence of mAb. Data shown is for a shear stress of 1 dyn/cm 2 . (D) Varying concentrations of GlyCAM-1 from HEC-GlcNAc6ST+/+ and −/− mice were coated yielding two different levels of rolling for each form of GlyCAM-1. The effect of 10 μg/ml of MECA79 on rolling was determined for each type and concentration of GlyCAM-1. Values represent the mean ± SD for shear stress of 1 dyn/cm 2 .

Article Snippet: MECA79 reactivity was determined using biotinylated MECA79 (Ligocyte Pharmaceuticals).

Techniques: Shear, Concentration Assay