anti human cd177 antibody  (Miltenyi Biotec)

Journal: Advanced Science

Article Title: CD177 Deficiency Defines a Stable Subtype of Human Neutrophil Granulocytes with Tumor Promoting Activity

doi: 10.1002/advs.76236

Figure Lengend Snippet: CD177 marks a stable and durable neutrophil subpopulation. (a) Exemplary flow cytometry plots showing differential CD177 expression on neutrophils (CD66b + ) in three individuals. (b) Percentage of CD177‐expressing neutrophils out of total neutrophil count in 11 healthy donors measured consecutively over multiple days, up to 160 days apart, values represented as mean±SD, with individual values color‐coded for the day of measurement. (c) Percentage of CD177 + neutrophils out of total neutrophil count in 4 healthy donors, measured over the course of 24 h at different timepoints. (d) Proportion of CD177 + neutrophils found in blood or sputum of 5 healthy individuals 2 h after irritation of the oral cavity using a 10% Tabasco‐saline solution. (e) Proportion of circulating CD177 + neutrophils in 33 ischemic stroke patients at different timepoints post hospital admission, measured via flow cytometry of peripheral blood. Grey bars depict patients with changes ≥18% over time; data presented as mean±SD for individual values at indicated time points.

Article Snippet: Cells were placed on ice, centrifuged for 5 min at 300xg at 4°C, resuspended in LIVE/DEAD Fixable Violet Dead Cell Stain (Invitrogen; Cat. # L34955 ) in PBS, and incubated on ice for 15 min. A FITC‐conjugated anti‐human CD177 antibody (Miltenyi Biotec; Cat. # 130‐126‐380) was added for 15 min at 4°C in PBS containing 0.5% BSA and 2 mM EDTA (AppliChem; Cat. # A4892) before fixation with 2% formaldehyde in PBS for 15 min on ice.

Techniques: Flow Cytometry, Expressing, Saline

Journal: Advanced Science

Article Title: CD177 Deficiency Defines a Stable Subtype of Human Neutrophil Granulocytes with Tumor Promoting Activity

doi: 10.1002/advs.76236

Figure Lengend Snippet: CD177 status remains unchanged upon neutrophil activation. (a) Exemplary flow cytometry plots comparing CD177 and CD62L expression patterns on neutrophils of one individual before and after stimulation with 50 ng/mL fMLP for 1 h. (b) Percentage of CD62L expression on neutrophils upon triggering with indicated stimuli for 1 h, analyzed by CD177 surface expression, data presented as mean±SD and individual values, n = 10. Compared to unstimulated all but ATP, LTB4, IL‐6, CytC, & IL 17 significantly reduced CD62L expression on the cell‐surface (* = p ≤ 0.0001). (c) Proportion of CD177 + neutrophils out of total neutrophil count from healthy individuals upon 1 h stimulation with the indicated stimuli, data presented as mean±SD with individual values color‐coded by donor (n = 10).

Article Snippet: Cells were placed on ice, centrifuged for 5 min at 300xg at 4°C, resuspended in LIVE/DEAD Fixable Violet Dead Cell Stain (Invitrogen; Cat. # L34955 ) in PBS, and incubated on ice for 15 min. A FITC‐conjugated anti‐human CD177 antibody (Miltenyi Biotec; Cat. # 130‐126‐380) was added for 15 min at 4°C in PBS containing 0.5% BSA and 2 mM EDTA (AppliChem; Cat. # A4892) before fixation with 2% formaldehyde in PBS for 15 min on ice.

Techniques: Activation Assay, Flow Cytometry, Expressing

Journal: Advanced Science

Article Title: CD177 Deficiency Defines a Stable Subtype of Human Neutrophil Granulocytes with Tumor Promoting Activity

doi: 10.1002/advs.76236

Figure Lengend Snippet: CD177 mRNA expression occurs in immature neutrophils. (a) Absolute transcript numbers of CD177 and established neutrophil markers CXCR2, CXCR4, and CXCL8 normalized to GAPDH, in CD177 sorted neutrophils. Values given as mean±SD, including individual values; (*: p ≤ 0.05), n = 9. (b) Gene signatures (mRNA expression) from different clusters of bone marrow neutrophils, following the patterns of maturation state, calculated from single‐cell RNA sequencing data as described in Montaldo et al. 2022 (18). (c) Single‐cell RNA sequencing of bone marrow neutrophils and their precursors, UMAP and clustering were calculated via unbiased highly variable genes and kmers, respectively (d) Corresponding expression of CD177 mRNA in individual cells within identified developmental neutrophil stages, n = 12 924. (e) Quantification of the amount of CD177 + expressing vs. CD177 − cells in the identified developmental stages shown in b and c. (f) Flow cytometric analysis of the CD177 expression pattern on neutrophils of two patients with acute lymphoblastic leukemia 7 days before (dpt ‐7) (middle panel) and 28 days after (dpt 28) (right panel) hematopoietic stem cell transplantation. The CD177 expression pattern of the respective donor is shown in the left panel for comparison. The patients showed a clinically confirmed donor chimerism of >99% on dpt 28.

Article Snippet: Cells were placed on ice, centrifuged for 5 min at 300xg at 4°C, resuspended in LIVE/DEAD Fixable Violet Dead Cell Stain (Invitrogen; Cat. # L34955 ) in PBS, and incubated on ice for 15 min. A FITC‐conjugated anti‐human CD177 antibody (Miltenyi Biotec; Cat. # 130‐126‐380) was added for 15 min at 4°C in PBS containing 0.5% BSA and 2 mM EDTA (AppliChem; Cat. # A4892) before fixation with 2% formaldehyde in PBS for 15 min on ice.

Techniques: Expressing, Single Cell, RNA Sequencing, Transplantation Assay, Comparison

Journal: Advanced Science

Article Title: CD177 Deficiency Defines a Stable Subtype of Human Neutrophil Granulocytes with Tumor Promoting Activity

doi: 10.1002/advs.76236

Figure Lengend Snippet: CD177 protein expression is developmentally fixed and absent from mature CD177 − neutrophils. (a) Confocal imaging of neutrophils isolated from peripheral blood of a healthy donor, stained for CD66b (cyan), CD177 (magenta), and DAPI (grey) as nuclear counterstain; scale bar is 30 µm. (b) May‐Grünwald‐staining of FACS‐sorted and cytospun CD177 − and CD177 + neutrophils used as input data for multidimensional feature comparison; scale bar is 10 µm. (c) UMAP dimension reduction of accessed shape features derived from >4000 sorted CD177 − and CD177 + neutrophils, including representative images of resulting clusters.

Article Snippet: Cells were placed on ice, centrifuged for 5 min at 300xg at 4°C, resuspended in LIVE/DEAD Fixable Violet Dead Cell Stain (Invitrogen; Cat. # L34955 ) in PBS, and incubated on ice for 15 min. A FITC‐conjugated anti‐human CD177 antibody (Miltenyi Biotec; Cat. # 130‐126‐380) was added for 15 min at 4°C in PBS containing 0.5% BSA and 2 mM EDTA (AppliChem; Cat. # A4892) before fixation with 2% formaldehyde in PBS for 15 min on ice.

Techniques: Expressing, Imaging, Isolation, Staining, Comparison, Derivative Assay

Journal: Advanced Science

Article Title: CD177 Deficiency Defines a Stable Subtype of Human Neutrophil Granulocytes with Tumor Promoting Activity

doi: 10.1002/advs.76236

Figure Lengend Snippet: Integrated proteomic‐lipidomic profiling reveals distinct molecular signatures in CD177‐defined neutrophils. (a) PCA analysis of whole proteomics data of FACS‐sorted CD177 − and CD177 + neutrophils from 8 healthy donors (labeled D1‐D8). (b) Heatmap showing expression patterns of N1(black)/N2(red)‐associated proteins found in FACS‐sorted CD177 − and CD177 + neutrophils from 8 healthy donors (D1‐D8). (c) REACTOME pathway analysis of neutrophil proteomes of sorted CD177 − and CD177 + neutrophils (n = 8). (d) PCA analysis of whole lipidomic data of MACS‐sorted CD177 − and CD177 + neutrophils from 3 healthy donors. (e) Heatmap showing lipids found in MACS‐sorted CD177 − and CD177 + neutrophils from 3 healthy donors. (f) Heatmap depicting expression patterns of proteins associated with fatty acid oxidation (top) and arachidonic acid metabolism (bottom) in FACS‐sorted CD177 − and CD177 + neutrophils from 8 healthy donors (D1‐D8).

Article Snippet: Cells were placed on ice, centrifuged for 5 min at 300xg at 4°C, resuspended in LIVE/DEAD Fixable Violet Dead Cell Stain (Invitrogen; Cat. # L34955 ) in PBS, and incubated on ice for 15 min. A FITC‐conjugated anti‐human CD177 antibody (Miltenyi Biotec; Cat. # 130‐126‐380) was added for 15 min at 4°C in PBS containing 0.5% BSA and 2 mM EDTA (AppliChem; Cat. # A4892) before fixation with 2% formaldehyde in PBS for 15 min on ice.

Techniques: Labeling, Expressing

Journal: Advanced Science

Article Title: CD177 Deficiency Defines a Stable Subtype of Human Neutrophil Granulocytes with Tumor Promoting Activity

doi: 10.1002/advs.76236

Figure Lengend Snippet: CD177 delineates an immunoregulatory neutrophil subtype in head‐and‐neck cancer. (a) Proportion of CD177 expressing tumor‐associated neutrophils (TAN) in head‐and‐neck cancer (HNC) tissue separated by tumor progression stage (early T1/2, late T3/4), data given as mean+SD (* = p ≤ 0.05), n(early) = 9; n(late) = 17. (b) Kaplan‐Meier curves, showing long‐term survival of HNC patients with >10% CD177 + TANs found within the tumor stroma (magenta) against patients with <10% CD177 + (blue) separated by early stages (T1/T2; top) and late stages (T3/T4; bottom). Significant differences according to the Log‐rank test were found for early‐stage tumors. (c) Microscopy images of UT‐SCC‐50 cells cocultured with the indicated neutrophil subpopulation for 48 h and quantification of cell‐free area. (d) Microscopy images of UT‐SCC‐50 cells cocultured with T cells and indicated autologous CD177‐FACS‐sorted neutrophils for 48 h and quantification of cell free area.

Article Snippet: Cells were placed on ice, centrifuged for 5 min at 300xg at 4°C, resuspended in LIVE/DEAD Fixable Violet Dead Cell Stain (Invitrogen; Cat. # L34955 ) in PBS, and incubated on ice for 15 min. A FITC‐conjugated anti‐human CD177 antibody (Miltenyi Biotec; Cat. # 130‐126‐380) was added for 15 min at 4°C in PBS containing 0.5% BSA and 2 mM EDTA (AppliChem; Cat. # A4892) before fixation with 2% formaldehyde in PBS for 15 min on ice.

Techniques: Expressing, Microscopy

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: CD177 + neutrophils alleviate acute pancreatitis-associated lung injury by protecting colonic barrier and restoring gut microbiota

doi: 10.1007/s00018-026-06116-1

Figure Lengend Snippet: Ablation of Cd177 in mice deteriorates pancreatic damage and neutrophil infiltration in the context of AP. ( a ) Representative images of pancreas sections obtained from mice as indicated, with H&E stained. The first row represents the WT sham and KO sham groups (×100). The second row represents WT AP and KO AP groups (×100). The third row represents the enlarged pictures in the second row (×400). The arrows indicate neutrophils. ( b ) Changes in pancreatic edema, acinar necrosis, inflammatory cell infiltration, and total pancreatic pathology scores measured between WT sham and KO sham groups ( n = 10 each). ( c ) Changes in serum amylase and lipase levels measured between WT sham and KO sham groups at 24 h after AP or sham surgery ( n = 5 each). ( d ) Representative images of pancreas sections obtained from the indicated mice and stained for MPO. ( e ) Changes in MPO IHC staining scores measured between WT sham and KO sham groups ( n = 5 each). WT, wild type; Sham, sham operation; KO, Cd177 knock-out; AP, acute pancreatitis. Data are presented as mean ± SEM (standard error of mean). * P < 0.05, ** P < 0.01, and *** P < 0.001 as indicated

Article Snippet: In addition, Cd177 flox/flox ; MRP8 Cre (conditional knock-out, cKO) mice were obtained from Shanghai Model Organisms Company (Shanghai, China) using Cre-loxP system (Supplementary Fig. c-d).

Techniques: Staining, Immunohistochemistry, Knock-Out

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: CD177 + neutrophils alleviate acute pancreatitis-associated lung injury by protecting colonic barrier and restoring gut microbiota

doi: 10.1007/s00018-026-06116-1

Figure Lengend Snippet: Ablation of Cd177 aggravates lung injury and neutrophil infiltration in the context of AP. ( a ) Representative images of lung sections obtained from mice as indicated, with H&E stained. The first row represents the WT sham and KO sham groups (×100). The second row represents WT AP and KO AP groups (×100). The third row represents the enlarged pictures in the second row (×400). ( b ) Changes in alveolar edema, inflammatory cell infiltration, and total lung pathology scores measured in the WT sham versus KO sham groups ( n = 10 each). ( c ) Changes in MPO IHC staining scores measured the lungs of the WT sham versus KO sham groups ( n = 5 each). ( d ) Representative images of lung sections obtained from the indicated mice and stained for MPO. WT, wild-type; Sham, sham operation; KO, knock-out; AP, acute pancreatitis. Data are presented as mean ± SEM. * P < 0.05, and ** P < 0.01 as indicated

Article Snippet: In addition, Cd177 flox/flox ; MRP8 Cre (conditional knock-out, cKO) mice were obtained from Shanghai Model Organisms Company (Shanghai, China) using Cre-loxP system (Supplementary Fig. c-d).

Techniques: Staining, Immunohistochemistry, Knock-Out

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: CD177 + neutrophils alleviate acute pancreatitis-associated lung injury by protecting colonic barrier and restoring gut microbiota

doi: 10.1007/s00018-026-06116-1

Figure Lengend Snippet: Ablation of Cd177 impairs the integrity of the colonic mucosal barrier in the context of AP. ( a ) Representative images of colon sections obtained from the indicated mice, with H&E stained. ( b ) Representative images of colon sections obtained from the indicated mice and stained for MPO. ( c ) Changes in MPO IHC staining scores measured in the colon in the indicated groups ( n = 5 each). ( d ) Intestinal histological scores ( n = 5 each). ( e ) Western blot analysis (top) and changes in (bottom) of E-cadherin and Occludin proteins in colon tissues obtained from the indicated groups ( n = 4 each). WT, wild-type; Sham, sham operation; KO, knock-out; AP, acute pancreatitis. Data are presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001 as indicated

Article Snippet: In addition, Cd177 flox/flox ; MRP8 Cre (conditional knock-out, cKO) mice were obtained from Shanghai Model Organisms Company (Shanghai, China) using Cre-loxP system (Supplementary Fig. c-d).

Techniques: Staining, Immunohistochemistry, Western Blot, Knock-Out

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: CD177 + neutrophils alleviate acute pancreatitis-associated lung injury by protecting colonic barrier and restoring gut microbiota

doi: 10.1007/s00018-026-06116-1

Figure Lengend Snippet: Ablation of Cd177 in neutrophils aggravates pancreatic damage and neutrophil infiltration in mice with AP. ( a ) Representative images of pancreas sections obtained from the indicated mice and stained with H&E. ( b ) Changes in pancreatic edema, acinar necrosis, inflammatory cell infiltration, and total pancreatic pathology scores measured in mice as indicated. ( c ) Changes in serum amylase and lipase levels measured in mice as indicated. ( d ) Representative images of pancreas sections obtained from the indicated mice and stained for MPO. ( e ) Changes in MPO IHC staining scores measured in indicated mice. n = 6 each. Sham, sham operation; AP, acute pancreatitis. Data are presented as mean ± SEM. ns, not statistically significant; * P < 0.05, ** P < 0.01, and *** P < 0.001 as indicated

Article Snippet: In addition, Cd177 flox/flox ; MRP8 Cre (conditional knock-out, cKO) mice were obtained from Shanghai Model Organisms Company (Shanghai, China) using Cre-loxP system (Supplementary Fig. c-d).

Techniques: Staining, Immunohistochemistry

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: CD177 + neutrophils alleviate acute pancreatitis-associated lung injury by protecting colonic barrier and restoring gut microbiota

doi: 10.1007/s00018-026-06116-1

Figure Lengend Snippet: Ablation of Cd177 in mouse neutrophils aggravates lung injury and neutrophil infiltration in mice with AP, but ABX treatment alleviates these changes. ( a ) Representative images of lung sections obtained from the indicated mice and stained with H&E. ( b ) Changes in alveolar edema, inflammatory cell infiltration, and total lung pathology scores measured in mice as indicated. ( c ) Changes in MPO IHC staining scores measured in mice as indicated. ( d ) Representative images of lung sections of mice as indicated, with MPO stained. n = 6 each. Sham, sham operation; AP, acute pancreatitis. Data are presented as mean ± SEM. ns, not statistically significant, * P < 0.05, ** P < 0.01, and *** P < 0.001 as indicated

Article Snippet: In addition, Cd177 flox/flox ; MRP8 Cre (conditional knock-out, cKO) mice were obtained from Shanghai Model Organisms Company (Shanghai, China) using Cre-loxP system (Supplementary Fig. c-d).

Techniques: Staining, Immunohistochemistry

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: CD177 + neutrophils alleviate acute pancreatitis-associated lung injury by protecting colonic barrier and restoring gut microbiota

doi: 10.1007/s00018-026-06116-1

Figure Lengend Snippet: Cd177 cKO impairs the colonic mucosal barrier in mice with AP. ( a ) Representative images of colon sections obtained from the indicated mice and stained with H&E. n = 6 each. ( b ) Intestinal histological scores. n = 4 each. ( c ) The mRNA levels of colonic barrier proteins in colon tissues obtained from the indicated groups. n = 4 each. ( d ) Western blot analysis (top) and changes in (bottom) of E-cadherin and Occludin proteins in colon tissues obtained from the indicated groups. n = 4 each. ( e ) Representative images of colon sections obtained from the indicated mice and stained for MPO. n = 6 each. (f) Changes in MPO IHC staining scores measured in mice as indicated. n = 6 each. WT, wild-type; Sham, sham operation; KO, knock-out; cKO, conditional knock-out; AP, acute pancreatitis. Data are presented as mean ± SEM. ns, not statistically significant, * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001 as indicated

Article Snippet: In addition, Cd177 flox/flox ; MRP8 Cre (conditional knock-out, cKO) mice were obtained from Shanghai Model Organisms Company (Shanghai, China) using Cre-loxP system (Supplementary Fig. c-d).

Techniques: Staining, Western Blot, Immunohistochemistry, Knock-Out

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: CD177 + neutrophils alleviate acute pancreatitis-associated lung injury by protecting colonic barrier and restoring gut microbiota

doi: 10.1007/s00018-026-06116-1

Figure Lengend Snippet: Ablation of Cd177 affects the change in gut microbiota species in AP mice. ( a ) Differences in gut microbiota species at the genus level. ( b ) Differences in gut microbiota species at the species level. ( c ) Metagenomic KEGG functional enrichment analysis of Cd177 flox/flox vs. Cd177 flox/flox ; MRP8 Cre group in AP mice. WT, wild-type; Sham, sham (operation); KO, knock-out; AP, acute pancreatitis. * P < 0.05, and ** P < 0.01 as indicated

Article Snippet: In addition, Cd177 flox/flox ; MRP8 Cre (conditional knock-out, cKO) mice were obtained from Shanghai Model Organisms Company (Shanghai, China) using Cre-loxP system (Supplementary Fig. c-d).

Techniques: Functional Assay, Knock-Out

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: CD177 + neutrophils alleviate acute pancreatitis-associated lung injury by protecting colonic barrier and restoring gut microbiota

doi: 10.1007/s00018-026-06116-1

Figure Lengend Snippet: The “gut-lung axis” mechanisms underlying the deterioration of acute pancreatitis-associated lung injury in AP mice after the conventional or conditional ablation of Cd177 (Created in BioRender)

Article Snippet: In addition, Cd177 flox/flox ; MRP8 Cre (conditional knock-out, cKO) mice were obtained from Shanghai Model Organisms Company (Shanghai, China) using Cre-loxP system (Supplementary Fig. c-d).

Techniques: