Journal: Scientific reports

Article Title: Prognostic impact of PD-L1 and TIGIT expression in non-small cell lung cancer following concurrent chemo-radiotherapy.

doi: 10.1038/s41598-023-29724-4

Figure Lengend Snippet: Figure 3. Expression rate of the TIL markers on CD8 + cells after induction therapy. The results are shown according to (A) PD-1 and (B) TIGIT expression.

Article Snippet: For CD8 and PD-1 multistaining, we used citrate buffer (pH 6.0) at 98 °C as antigen retrieval buffer and rabbit anti-CD8 antibody (clone 1779R, Novus Biologicals, Centennial CO, US) diluted at 1:300 and mouse anti-PD-1 antibody (clone NAT105, Abcam, Cambridge, UK) diluted at 1:50 as the primary antibody and Alexa Flour 488-conjugated goat anti-rabbit IgG (Thermo Fisher Scientific, Massachusetts, US) diluted at 1:500 and Alexa Fluor 594-conjugated goat anti-mouse IgG (Thermo Fisher Scientific, Waltham MA, US) diluted to 1:500 as the secondary antibodies.

Techniques: Expressing

Journal: Journal of Clinical Investigation

Article Title: Distinct histopathological phenotypes of severe alcoholic hepatitis suggest different mechanisms driving liver injury and failure

doi: 10.1172/jci157780

Figure Lengend Snippet: Figure 1. IHC staining analyses of inflammatory cells in the liver of SAH patients. (A and B) Liver tissues from SAH patients (n = 10) were subjected to sequential multiplex immunofluorescence staining of α-SMA, HepPar1, MPO, CD8, CD4, CD20, and IBA1. (A) Typical starry image of immunofluorescent staining from 1 SAH patient with a large number of inflammatory cells surrounding hepatocytes. (B) Two different patterns of representative images from SAH55 and SAH1 patients. (C) Liver tissues from healthy controls (HC) (n = 5) and SAH patients (n = 40) were subjected to IHC staining for MPO, CD8, and CD4. The number of MPO+, CD8+, CD4+ cells in the parenchymal area (SAH-P) and fibrotic area (SAH-F) were quantified. From 5 to 7 parenchymal or fibrotic areas were randomly selected for quantitation, and average numbers of cells are shown. For the graph, each dot represents one patient. (D and E) Analyses of SAH data from C show negative correlation between P-Neu and F-CD8+ T cells or F-CD4+ T cells and positive correlation between F-CD8+ T cells and F-CD4+ T cells. P values are indicated. Data are represented as mean ± SEM. ***P < 0.001. Statistical significance was assessed using 1-way ANOVA followed by Tukey’s post hoc test for multiple groups (C) and Pearson’s correlation analysis (E).

Article Snippet: The following antibodies were used: HepPar1 (catalog M7158, Dako), CD8a (catalog NBP2-34318-0.1 mg, Novus), CD4 (catalog ab133616, Abcam), MPO (catalog ab208670, Abcam), CD20 (catalog PA5-16701, Thermo Fisher), IBA1 (catalog MABN92, EMD Millipore), and α-SMA (catalog M085129-2, Agilent).

Techniques: Immunohistochemistry, Multiplex Assay, Immunofluorescence, Staining, Quantitation Assay

Journal: Journal of Clinical Investigation

Article Title: Distinct histopathological phenotypes of severe alcoholic hepatitis suggest different mechanisms driving liver injury and failure

doi: 10.1172/jci157780

Figure Lengend Snippet: Figure 2. Hepatic neutrophil and CD8+ T cell infiltration are associated with liver injury in patients with SAH. (A) Upper panel: correlation analysis between the numbers of P-Neu per field and MELD score, serum ALT, or AST level; lower panel: correlation analysis between the numbers of F-CD8+ T cells per field and MELD score, serum ALT, or ALT level. (B–E) SAH patients were dichotomized into Neulo and Neuhi groups (using the average quantity at 50 cells/field of intrahepatic neutrophils in our study cohort as the cutoff) or into CD8lo and CD8hi groups. MELD score, serum AST, and ALT levels were compared between groups. For panels B and D, all SAH patients were used for the comparison. For panels C and E, only SAH patients with abnormal ALT (>40 U/L) levels were used for the comparison. Data are represented as mean ± SEM. *P < 0.05. Statistical significance was assessed using Pearson’s correlation analysis (A) and 2-tailed Student’s t test for comparing 2 groups (B–E).

Article Snippet: The following antibodies were used: HepPar1 (catalog M7158, Dako), CD8a (catalog NBP2-34318-0.1 mg, Novus), CD4 (catalog ab133616, Abcam), MPO (catalog ab208670, Abcam), CD20 (catalog PA5-16701, Thermo Fisher), IBA1 (catalog MABN92, EMD Millipore), and α-SMA (catalog M085129-2, Agilent).

Techniques: Comparison

Journal: Journal of Clinical Investigation

Article Title: Distinct histopathological phenotypes of severe alcoholic hepatitis suggest different mechanisms driving liver injury and failure

doi: 10.1172/jci157780

Figure Lengend Snippet: Figure 4. Hepatic neutrophil infiltration negatively correlates with fibrosis progression in patients with SAH, cirrhosis, and alcoholic steatohepatitis. (A and B) Liver tissues from SAH patients (n = 33) were subjected to Sirius red staining. Representative images from NeuhiCD8lo and NeuloCD8hi groups are shown (A). Scale bars: 500 μm. (B) Fibrosis scores were evaluated by the Ishak system, and SAH patients were divided into 2 groups based on fibrosis scores and hepatic neutrophils. CD8+ T cell numbers were compared between the 2 groups. (C) Liver tissues from healthy control (n = 6) and alcoholic cirrhosis (ALC) patients (n = 32) were subjected to IHC staining of MPO, CD8, and MPO+; CD8+ cells in the parenchymal area (ALC-P) and fibrotic area (ALC-F) were quantified. (D) Liver tissues from healthy controls (n = 6) and alcoholic steatohepatitis patients (n = 12) were subjected to IHC staining of MPO+ and CD8+ cells. Representative images (top) and quantification are shown (bottom). Red arrows indi- cate MPO+ staining, and blue arrows indicate CD8+ staining. Scale bars: 100 μm. Data are represented as mean ± SEM. *P < 0.05; ***P < 0.001. Statistical significance was assessed using 2-tailed Student’s t test for comparing 2 groups (B and D) and 1-way ANOVA followed by Tukey’s post hoc test for multiple groups (C).

Article Snippet: The following antibodies were used: HepPar1 (catalog M7158, Dako), CD8a (catalog NBP2-34318-0.1 mg, Novus), CD4 (catalog ab133616, Abcam), MPO (catalog ab208670, Abcam), CD20 (catalog PA5-16701, Thermo Fisher), IBA1 (catalog MABN92, EMD Millipore), and α-SMA (catalog M085129-2, Agilent).

Techniques: Staining, Control, Immunohistochemistry

Journal: Journal of Clinical Investigation

Article Title: Distinct histopathological phenotypes of severe alcoholic hepatitis suggest different mechanisms driving liver injury and failure

doi: 10.1172/jci157780

Figure Lengend Snippet: Figure 6. Hepatic NCF1 expression is upregulated and positively correlated with neutrophil-related genes in patients and mice with alcoholic steatohep- atitis. (A) Liver tissues from alcoholic steatohepatitis patients were subjected to immunofluorescence staining of MPO and NCF1. Representative images are shown. Scale bar: 23 μm. (B) Correlation analyses were performed between NCF1 and neutrophil-related genes based on RNA-Seq data (n = 12). Values in x and y axes represent relative transcripts per million (TPM). (C and D) Liver tissues from ethanol- (n = 3) and pair-fed (n = 3) mice were subjected to microarray analyses. Heatmap analyses of neutrophil-related genes (C) and CD8+ T cell–related genes (D). Microarray data were pulled from our previous paper (50); GEO database number is GSE67546. Statistical significance was assessed using Pearson’s correlation analysis (B).

Article Snippet: The following antibodies were used: HepPar1 (catalog M7158, Dako), CD8a (catalog NBP2-34318-0.1 mg, Novus), CD4 (catalog ab133616, Abcam), MPO (catalog ab208670, Abcam), CD20 (catalog PA5-16701, Thermo Fisher), IBA1 (catalog MABN92, EMD Millipore), and α-SMA (catalog M085129-2, Agilent).

Techniques: Expressing, Immunofluorescence, Staining, RNA Sequencing, Microarray

Journal: Journal of Clinical Investigation

Article Title: Distinct histopathological phenotypes of severe alcoholic hepatitis suggest different mechanisms driving liver injury and failure

doi: 10.1172/jci157780

Figure Lengend Snippet: Figure 11. Model depicting distinct histopathological phenotypes in SAH patients based on liver immune phenotyping and the role of neutrophilic NCF1-dependent ROS in AH pathogenesis. SAH exists as 2 distinct histopathological pheno- types, NeuhiCD8lo and NeuloCD8hi, suggesting different pathogenesis leading to liver injury and/or failure among these patients. Patients with high hepatic neutrophils but low CD8+ T cells are younger and have higher MELD scores and ALT levels, but have less fibrosis compared with those with NeuloCD8hi. Among those with Neuhi, neutrophilic NCF1-depen- dent ROS promotes AH progression through the NCF1/SIRT1/AMPK axis on lipid metabolism and NCF1/p38 MAPK/miR-223 on alcohol-induced inflammation and fibrosis.

Article Snippet: The following antibodies were used: HepPar1 (catalog M7158, Dako), CD8a (catalog NBP2-34318-0.1 mg, Novus), CD4 (catalog ab133616, Abcam), MPO (catalog ab208670, Abcam), CD20 (catalog PA5-16701, Thermo Fisher), IBA1 (catalog MABN92, EMD Millipore), and α-SMA (catalog M085129-2, Agilent).

Techniques:

Journal: BioMed Research International

Article Title: B7-H4 Expression in Precancerous Lesions of the Uterine Cervix

doi: 10.1155/2021/5857092

Figure Lengend Snippet: Correlation between B7-H4 expression and the number of infiltrating T lymphocytes and cytokines in patients with cervical disease.

Article Snippet: After blocking with 5% goat serum, slides were incubated with the following primary antibodies overnight at 4°C: rabbit anti-human Foxp3 (dilution 1 : 300, NB100-39002SS, Novus Biologicals, Littleton, CO, USA), mouse anti-human CD4 (dilution 1 : 200, NBP2-27216, Novus Biologicals), mouse anti-human CD8 alpha (dilution 1 : 200, NBP2-32836, Novus Biologicals), and rabbit anti-human IFN- γ (dilution 1 : 100, 8455P, Cell Signaling Technology).

Techniques: Expressing

Journal: BioMed Research International

Article Title: B7-H4 Expression in Precancerous Lesions of the Uterine Cervix

doi: 10.1155/2021/5857092

Figure Lengend Snippet: Double immunofluorescence-based staining of (a) CD4 + Foxp3 + and (b) interferon (IFN)- γ + CD8 + T cells. Magnification: ×200.

Article Snippet: After blocking with 5% goat serum, slides were incubated with the following primary antibodies overnight at 4°C: rabbit anti-human Foxp3 (dilution 1 : 300, NB100-39002SS, Novus Biologicals, Littleton, CO, USA), mouse anti-human CD4 (dilution 1 : 200, NBP2-27216, Novus Biologicals), mouse anti-human CD8 alpha (dilution 1 : 200, NBP2-32836, Novus Biologicals), and rabbit anti-human IFN- γ (dilution 1 : 100, 8455P, Cell Signaling Technology).

Techniques: Double Immunofluorescence Staining

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Reproducible, high-dimensional imaging in archival human tissue by Multiplexed Ion Beam Imaging by Time-of-Flight (MIBI-TOF)

doi: 10.1038/s41374-022-00778-8

Figure Lengend Snippet: MIBI-TOF staining panel.

Article Snippet: Individual metal labeled antibodies for IHC were also obtained from Ionpath: CD3 (D7A6E), CD8 (C8/144B), CD68 (D4B9C), Pax5 (D7H5X), PanCK (AE1/AE3).

Techniques: Staining