Journal: Frontiers in Immunology

Article Title: Colorectal Cancer-Associated Immune Exhaustion Involves T and B Lymphocytes and Conventional NK Cells and Correlates With a Shorter Overall Survival

doi: 10.3389/fimmu.2021.778329

Figure Lengend Snippet: Antibodies used in immunostaining.

Article Snippet: IDO1 , GT273 , Mouse , GTX634652 , Genetex, Irvine, CA, USA.

Techniques: Immunostaining

Journal: Acta Pharmacologica Sinica

Article Title: IDO-1 inhibitor INCB24360 elicits distant metastasis of basal extruded cancer cells in pancreatic ductal adenocarcinoma

doi: 10.1038/s41401-022-01035-w

Figure Lengend Snippet: a Immunostaining IDO-1 and E-cadherin antibodies in human PDACs. IDO-1 expression patterns in normal pancreatic ducts and precursor lesions (panINs) and early invasive lesions (white arrowheads, ducts; the lower panel in the left panel, E-cadherin staining, and nuclear morphology in the boxed region). b IDO-1 expression in the neoplastic duct, invasive lesions, and metastatic lymph lesions of PDACs (the upper panel, the split channels of the boxed region; the lower left in the right panel, the magnification of the boxed region). c Comparing the IDO-1 staining intensity in the neoplastic cells in PDAC or invasive lesions (sample size, n > 15) with lymph metastatic neoplastic cells and normal/precursors ducts (sample size, n = 4). Data, mean ± SD. One-way ANOVA. ** P < 0.01. d Immunostaining IDO-1 and E-cadherin antibodies in human PDACs revealed IDO-1 is also exclusively upregulated in the cells with apical or basal extrusion (the lower panels, single channel of the boxed region). The 3D-rendered image revealed the morphology of basal extruding cells. e , f Significant fraction of IDO-1 + neoplastic cells in PDAC have discontinuous or uneven E-cadherin membrane staining. The circled area in the right and left panels (yellow arrowheads, IDO-1 high , and E-cadherin low neoplastic cells). The graph showed the counts of IDO-1 high and E-cadherin low cells and IDO-1 low and E-cadherin high cells in PDAC tissues.

Article Snippet: Following antibodies were used for immunostaining or immunoblotting: IDO-1 (Cell Signaling Technology, D5J4E TM , rabbit, mAb #86630, 1:1000), CD45RA (BD Pharmingen, Clone: L48, 1:200), Foxp3 (Abcam, ab20034, 1:200), human VEGFR2/KDR/Flk-1 antibody (R&D, AF357, 1:200), E-Cadherin antibody (R&D, AF748, 1:100), CD11b antibody (Abcam, ab34216, 1:10000), CD8 antibody (Abcam, ab4505, 1:1000), CD11b antibody (Abcam, ab133357, 1:10000), purified mouse anti-human CD16 antibody (BD Pharmingen, Clone GO22, 1:200), anti-glucose transporter GLUT1 antibody (Abcam, ab115730, 1:1000), purified mouse anti-human CD3 antibody (BD Pharmingen, Clone SP34-2, 556648, 1:500), recombinant anti-Cytokeratin 19 antibody (Abcam, EP1580Y, 1:1000), 6-diamidino-2-phenylindole (DAPI, Sigma, D9542, 1:1000), IDO monoclonal antibody (Sigma, Clone mIDO-48, 1:500), IDO-1 monoclonal antibody (Enzo Life Sciences, Clone 2E2, 1:200), Vimentin antibody (Thermo Fisher Scientific, MA5-11883, 1:1000), Ki67 (EPR3610, Abcam, 1:1000), E-cadherin (Abcam, Ab1416, 1:500).

Techniques: Immunostaining, Expressing, Staining

Journal: Acta Pharmacologica Sinica

Article Title: IDO-1 inhibitor INCB24360 elicits distant metastasis of basal extruded cancer cells in pancreatic ductal adenocarcinoma

doi: 10.1038/s41401-022-01035-w

Figure Lengend Snippet: a CK-19, IDO-1, and vimentin antibodies immunostaining showed that IDO-1 + and vimentin + cells in the neoplastic cells of PDAC that localized on the basal domain of ducts expressed a lower level of CK-19 (white arrows, basal extruding cells with EMT). Scale bar, 20 µm. Student t test. ** P < 0.001. Data, mean ± SEM. b E-cadherin, IDO-1, and vimentin antibodies immunostaining showed that vimentin antibodies stained a few IDO-1 + and E-cadherin + cells in PDAC tissues. Student t test. * P < 0.05, ** P < 0.01. Data, mean ± SEM. c 1 ng/mL Abraxane and 5 ng/mL gemcitabine-treated KPIC cells for 12 h were stained by CK-19, E-cadherin, and vimentin antibodies. The results CK-19 immunostaining intensity in cells with higher vimentin levels and lower vimentin levels showed that KPIC cells with higher levels of vimentin also expressed higher levels of CK-19 and vice versa. Student’s t test. ** P < 0.01. Data, mean ± SEM. d 5 ng/mL and 10 ng/mL gemcitabine-treated KPIC cells with over 80% confluence in dishes were performed CK-19, vimentin, and TUNEL staining. The measurement of staining intensity showed that TUNEL + cells with intact cellular structure expressed higher levels of CK-19 compared to TUNEL - KPIC cells. Data, mean ± SD. t test, ** P < 0.01.

Article Snippet: Following antibodies were used for immunostaining or immunoblotting: IDO-1 (Cell Signaling Technology, D5J4E TM , rabbit, mAb #86630, 1:1000), CD45RA (BD Pharmingen, Clone: L48, 1:200), Foxp3 (Abcam, ab20034, 1:200), human VEGFR2/KDR/Flk-1 antibody (R&D, AF357, 1:200), E-Cadherin antibody (R&D, AF748, 1:100), CD11b antibody (Abcam, ab34216, 1:10000), CD8 antibody (Abcam, ab4505, 1:1000), CD11b antibody (Abcam, ab133357, 1:10000), purified mouse anti-human CD16 antibody (BD Pharmingen, Clone GO22, 1:200), anti-glucose transporter GLUT1 antibody (Abcam, ab115730, 1:1000), purified mouse anti-human CD3 antibody (BD Pharmingen, Clone SP34-2, 556648, 1:500), recombinant anti-Cytokeratin 19 antibody (Abcam, EP1580Y, 1:1000), 6-diamidino-2-phenylindole (DAPI, Sigma, D9542, 1:1000), IDO monoclonal antibody (Sigma, Clone mIDO-48, 1:500), IDO-1 monoclonal antibody (Enzo Life Sciences, Clone 2E2, 1:200), Vimentin antibody (Thermo Fisher Scientific, MA5-11883, 1:1000), Ki67 (EPR3610, Abcam, 1:1000), E-cadherin (Abcam, Ab1416, 1:500).

Techniques: Immunostaining, Staining, TUNEL Assay

Journal: Acta Pharmacologica Sinica

Article Title: IDO-1 inhibitor INCB24360 elicits distant metastasis of basal extruded cancer cells in pancreatic ductal adenocarcinoma

doi: 10.1038/s41401-022-01035-w

Figure Lengend Snippet: a The typical morphology of KPIC PDAC organoids taken by DIC microscopy and E-cadherin, CK-19, and vimentin antibodies co-immunostaining or Glut-1 and IDO-1 antibodies co-immunostaining at day 7. The spliced images in the middle panel from the boxed region showed that the PDAC KPIC organoids did not express vimentin. b–d Treating KPIC PDAC organoids with 0, 10, and 20 ng/mL IFN-γ for 24 h increases apical extrusion of IDO-1 + KPIC cells ( b ). IDO-1 expression levels in KPIC 2D culture after treatment with 0, 10, and 20 ng/mL IFN-γ and 20 µM 1-MT for 24 h ( c , d ). The intensity of the IDO-1 signal in each cell was measured by Imaris 9.7. One-way ANOVA, ** P < 0.01. e Treating KPIC PDAC organoids with 0, 1, 10, and 20 ng/mL IFN-γ for 24 h and 48 h increased the distorting of ductal structure. Counting the ratio of ductal shape changes at 24 h and 48 h (replicates, n = 5; black arrowheads, the protruding basal cells).

Article Snippet: Following antibodies were used for immunostaining or immunoblotting: IDO-1 (Cell Signaling Technology, D5J4E TM , rabbit, mAb #86630, 1:1000), CD45RA (BD Pharmingen, Clone: L48, 1:200), Foxp3 (Abcam, ab20034, 1:200), human VEGFR2/KDR/Flk-1 antibody (R&D, AF357, 1:200), E-Cadherin antibody (R&D, AF748, 1:100), CD11b antibody (Abcam, ab34216, 1:10000), CD8 antibody (Abcam, ab4505, 1:1000), CD11b antibody (Abcam, ab133357, 1:10000), purified mouse anti-human CD16 antibody (BD Pharmingen, Clone GO22, 1:200), anti-glucose transporter GLUT1 antibody (Abcam, ab115730, 1:1000), purified mouse anti-human CD3 antibody (BD Pharmingen, Clone SP34-2, 556648, 1:500), recombinant anti-Cytokeratin 19 antibody (Abcam, EP1580Y, 1:1000), 6-diamidino-2-phenylindole (DAPI, Sigma, D9542, 1:1000), IDO monoclonal antibody (Sigma, Clone mIDO-48, 1:500), IDO-1 monoclonal antibody (Enzo Life Sciences, Clone 2E2, 1:200), Vimentin antibody (Thermo Fisher Scientific, MA5-11883, 1:1000), Ki67 (EPR3610, Abcam, 1:1000), E-cadherin (Abcam, Ab1416, 1:500).

Techniques: Microscopy, Immunostaining, Expressing

Journal: Acta Pharmacologica Sinica

Article Title: IDO-1 inhibitor INCB24360 elicits distant metastasis of basal extruded cancer cells in pancreatic ductal adenocarcinoma

doi: 10.1038/s41401-022-01035-w

Figure Lengend Snippet: a Testing IDO-1 expression in KPIC and KPIC Ido-1 KO cells. b Representative images of Ki67 antibody immunostaining and count of Ki67 + cells in KPIC and KPIC Ido-1 KO cells. Data, mean ± SD. t test, ** P < 0.01. c Representative images of dextran uptake in KPIC and KPIC Ido-1 KO cells after starvation and count of dextran vesicles in SIM Z-stacked images. Data, mean ± SD. t test, ** P < 0.01. d , e Comparing the growth patterns and speeds of KPIC Ido-1 KO PDAC organoids with KPIC organoids. Data, CI 95%. Two-way ANOVA. f KPIC Ido-1 KO organoids did not form a subcutaneous tumor in C57/B6 mice after transplantation for 4 weeks ( n = 7; white circled region, subcutaneous tumor of KPIC organoids). LS-MS data of KPIC and KPIC Ido-1 KO cells showed that CD86 ( g ), CD276 ( h ), H2-K1 ( i ), CD74 ( j ), and H2-D1 ( k ) significantly changed after Ido-1 knockout. Data, mean ± SD. t test; ** P < 0.01. Repeats, 3 times.

Article Snippet: Following antibodies were used for immunostaining or immunoblotting: IDO-1 (Cell Signaling Technology, D5J4E TM , rabbit, mAb #86630, 1:1000), CD45RA (BD Pharmingen, Clone: L48, 1:200), Foxp3 (Abcam, ab20034, 1:200), human VEGFR2/KDR/Flk-1 antibody (R&D, AF357, 1:200), E-Cadherin antibody (R&D, AF748, 1:100), CD11b antibody (Abcam, ab34216, 1:10000), CD8 antibody (Abcam, ab4505, 1:1000), CD11b antibody (Abcam, ab133357, 1:10000), purified mouse anti-human CD16 antibody (BD Pharmingen, Clone GO22, 1:200), anti-glucose transporter GLUT1 antibody (Abcam, ab115730, 1:1000), purified mouse anti-human CD3 antibody (BD Pharmingen, Clone SP34-2, 556648, 1:500), recombinant anti-Cytokeratin 19 antibody (Abcam, EP1580Y, 1:1000), 6-diamidino-2-phenylindole (DAPI, Sigma, D9542, 1:1000), IDO monoclonal antibody (Sigma, Clone mIDO-48, 1:500), IDO-1 monoclonal antibody (Enzo Life Sciences, Clone 2E2, 1:200), Vimentin antibody (Thermo Fisher Scientific, MA5-11883, 1:1000), Ki67 (EPR3610, Abcam, 1:1000), E-cadherin (Abcam, Ab1416, 1:500).

Techniques: Expressing, Immunostaining, Transplantation Assay, Knock-Out

Journal: Oncotarget

Article Title: P53 prevent tumor invasion and metastasis by down-regulating IDO in lung cancer

doi: 10.18632/oncotarget.17408

Figure Lengend Snippet: The relationship between IDO1 expression and clinicopathological characteristic in Lung cancer patients

Article Snippet: For incubation with primary mAb, Tissue slides were incubated at 4°C overnight with rabbit anti-IDO1 mAb (1:50; Abnova).

Techniques: Expressing

Journal: Oncotarget

Article Title: P53 prevent tumor invasion and metastasis by down-regulating IDO in lung cancer

doi: 10.18632/oncotarget.17408

Figure Lengend Snippet: (A) IDO1 expression of was determined by way of qRT-PCR in 64 paired human lung cancer and their corresponding nontumorous samples (NT) and normalized against an endogenous U6 RNA control. (B) IDO1expression in different clinical stages of lung cancer. Patients were staged in accordance with the 8th Edition of the TNM staging for lung cancer of UICC. (C) up-regulation of IDO1 in lung cancer was associated with lymph node-metastasis; patients were classified into lymph node-metastasis negative group (LN-negative) and positive group (LN-positive).

Article Snippet: For incubation with primary mAb, Tissue slides were incubated at 4°C overnight with rabbit anti-IDO1 mAb (1:50; Abnova).

Techniques: Expressing, Quantitative RT-PCR, Control

Journal: Oncotarget

Article Title: P53 prevent tumor invasion and metastasis by down-regulating IDO in lung cancer

doi: 10.18632/oncotarget.17408

Figure Lengend Snippet: (A) The expression levels of IDO1 were determined by qRT-PCR in lung cancer cell lines. U6 RNA served as an internal control. (B) And (C) Successful over-expression of IDO1 was confirmed by qRT-PCR after infection with IDO1-expressing lentivirus or vector control lentivirus. The values of IDO1 expression were calculated as fold change relative to the vector control.

Article Snippet: For incubation with primary mAb, Tissue slides were incubated at 4°C overnight with rabbit anti-IDO1 mAb (1:50; Abnova).

Techniques: Expressing, Quantitative RT-PCR, Control, Over Expression, Infection, Plasmid Preparation

Journal: Oncotarget

Article Title: P53 prevent tumor invasion and metastasis by down-regulating IDO in lung cancer

doi: 10.18632/oncotarget.17408

Figure Lengend Snippet: Overexpression of IDO1 significantly enhanced abilities of cell migration (A) and invasion (B) in NCI-H292 and HCC827 Lung cancer cell after infection with IDO1-expressing or vector lentivirus.

Article Snippet: For incubation with primary mAb, Tissue slides were incubated at 4°C overnight with rabbit anti-IDO1 mAb (1:50; Abnova).

Techniques: Over Expression, Migration, Infection, Expressing, Plasmid Preparation

Journal: Oncotarget

Article Title: P53 prevent tumor invasion and metastasis by down-regulating IDO in lung cancer

doi: 10.18632/oncotarget.17408

Figure Lengend Snippet: Downregulation of IDO1 significantly suppressed cell migration and invasion in NCI-H1299 Lung cancer cell

Article Snippet: For incubation with primary mAb, Tissue slides were incubated at 4°C overnight with rabbit anti-IDO1 mAb (1:50; Abnova).

Techniques: Migration

Journal: Oncotarget

Article Title: P53 prevent tumor invasion and metastasis by down-regulating IDO in lung cancer

doi: 10.18632/oncotarget.17408

Figure Lengend Snippet: (A) The distribution of p53 protein expression in lung cancer and corresponding nontumorous tissue (B) Correlation between p53 expression and IDO1 levels in the 64 lung cancer tissue samples. The expression levels of p53 were classified into low (scores of 0 and 1) and high groups (scores of 2 and 3) according to the scores of p53 immunohistochemical staining.

Article Snippet: For incubation with primary mAb, Tissue slides were incubated at 4°C overnight with rabbit anti-IDO1 mAb (1:50; Abnova).

Techniques: Expressing, Immunohistochemical staining, Staining

Journal: Oncotarget

Article Title: P53 prevent tumor invasion and metastasis by down-regulating IDO in lung cancer

doi: 10.18632/oncotarget.17408

Figure Lengend Snippet: (A) and (B) silencing of IDO 1 was confirmed by Western blot in NCI-H292 and HCC827 cells after transfection with specific si-IDO1. β-Actin served as an internal control. (C) and (D) migration and invasion assays were carried out in NCI-H292 and HCC827 cells after transfection with negative control (NC) or si-IDO1.

Article Snippet: For incubation with primary mAb, Tissue slides were incubated at 4°C overnight with rabbit anti-IDO1 mAb (1:50; Abnova).

Techniques: Western Blot, Transfection, Control, Migration, Negative Control

Journal: Oncotarget

Article Title: P53 prevent tumor invasion and metastasis by down-regulating IDO in lung cancer

doi: 10.18632/oncotarget.17408

Figure Lengend Snippet: (A) Western blot analysis was used to detect the IDO1 expression in NCI-H1299 cells after transfection with si-p53, si-IDO1 or NC. β-Actin served as an internal control. (B) NCI-H1299 cells after transfection with si-p53, si-IDO1 or NC were subjected to migration and invasion assays.

Article Snippet: For incubation with primary mAb, Tissue slides were incubated at 4°C overnight with rabbit anti-IDO1 mAb (1:50; Abnova).

Techniques: Western Blot, Expressing, Transfection, Control, Migration