GL261 or CT-2A glioma cells were orthotopically injected in the brain of ELTD1−/− and wildtype mice as described in the Supplementary Materials and Methods . For survival studies, mice were sacrificed when they lost more than 10% body weight or exhibited symptoms of a brain tumor. For the anti-PD-1 study, 200 µg of anti-mouse PD-1 (RMP1-14, Cat. # BE0146, InVivoMab, BioXcell) or rat-isotype IgG2a control (2A3, #BE089, InVivoMab, BioXcell) were administered intravenously every 3 days, starting on day 10 after tumor injection up to a total of 4 administrations.
Invivomab
Manufactured by BioXCell
Sourced in United States
InVivoMAb is a laboratory product that provides monoclonal antibodies for in vivo studies. The core function of InVivoMAb is to facilitate the investigation of biological processes and therapeutic interventions in living organisms.
Automatically generated - may contain errors
Lab products found in correlation
Countbright beads, by Thermo Fisher Scientific (2 mentions)
α il 22, by Genentech (2 mentions)
Cd19 beads, by Miltenyi Biotec (1 mentions)
Rat igg2b isotype control clone ltf 2, by BioXCell (1 mentions)
Rat igg2a isotype control clone 2a3, by BioXCell (1 mentions)
B95 8, by American Type Culture Collection (1 mentions)
Be0119, by BioXCell (1 mentions)
Mab466, by R&D Systems (1 mentions)
Be0117, by BioXCell (1 mentions)
Armenian hamster igg, by BioLegend (1 mentions)
Invivomab anti mouse ifnγ, by BioXCell (1 mentions)
Cxcr3 173, by BioLegend (1 mentions)
Human b cell isolation kit 2, by Miltenyi Biotec (1 mentions)
Cpg odn 2006, by InvivoGen (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Pen strep, by Merck Group (1 mentions)
Bp0089, by BioXCell (1 mentions)
Be0146, by BioXCell (1 mentions)
Anti mouse ly6g, by BioXCell (1 mentions)
Igg2a isotype control, by BioXCell (1 mentions)
33 protocols using invivomab
1
Orthotopic Glioma Model in ELTD1 Knockout Mice
2
Combinatorial Immunotherapy Enhances Tumor Regression
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C57Bl/6 female mice were injected subcutaneously (s.c.) with 5 × 106 EG.7-OVA or with 8 × 105 MCA205-Tlr3+/+ or with the same dose of MCA205-Tlr3-/- in Matrigel (0.1 mL/mouse) (BD Biosciences). When tumors reached a mean diameter of 9 ± 2 mm, they were treated intraperitoneally (i.p.) with 100 mg/kg CTX or 2.5 mg/kg CDDP followed by 3 injections of anti-PDL1 (clone 10F.9G2) and/or anti-PDL2 (clone TY25) Abs (InVivoMAb, BioXcell) in dilution buffer (InVivoPure pH 6.5, BioXcell). The first injection (150 μg/mouse) was given s.c. peritumorally 3 days after chemotherapy, the subsequent injections (250 μg/mouse) were given i.p. on days 7 and 10 after chemotherapy. In some experiments, mice received one s.c. peritumoral injection of 1000U IFN-I or mock instead of chemotherapy followed by three anti-PDL1/2 Ab administrations as detailed above. Control groups received the same volume of saline instead of the drugs and of control isotypes (IgG2b and IgG2a, InVivoMAb, BioXcell), instead of the specific Abs. Tumor growth was measured by a caliper twice a week. In some experiments, long-term survivors were challenged with 106 live EG.7-OVA cells s.c. into the right flank and the development of a new tumor mass was monitored twice a week and measured with a caliper.
3
Activation of B-1 cells by BLyS
4
Evaluation of Immunotherapies in Tumor-Bearing Mice
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All treatments began 10 days after tumor injection, and each treatment group consisted of 10 mice. Treatments included (i) anti-mouse Ly6G/Ly6C (anti-Gr-1 mAb) InVivoMab injected IP twice a week at 100 μg/mouse for 3 weeks; (ii) IgG2b isotype control, anti-keyhole limpet hemocyanin InVivoMab injected IP twice a week at 100 μg/kg body weight for 3 weeks; (iii) anti-mouse PD-1 InVivoMab injected intraperitoneally once a week at 100 μg/kg of their body weight for 3 weeks; (iv) gemcitabine injected IP twice a week at 100 mg/kg of body weight for 6 weeks and (v) vehicle PBS injected IP twice a week at 100 μl/mouse for 6 weeks. All InVivoMabs were purchased from BioX Cell (West Lebanon, NH). Gemcitabine was procured from Henry Ford Hospital Pharmacy, Detroit, MI.
5
Analyzing Memory B Cell Response
6
Depletion of Immune Cells in BRAF-Inhibitor Therapy
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Tumor-bearing animals were treated with BRAFi, oHSV and depletion antibodies. One hundred and fifty micrograms of in vivo depletion antibodies was injected via intraperitoneal injection every second day, starting 1 day prior to BRAF inhibition treatment, with a 2-day break between the second and third dose. A total of 11 injections were given. The following antibodies were used: anti-CD8 (clone 2.43), anti-CD4 (clone GK1.5), anti-NK1.1 (clone PK136) and isotype control (clone LTF-2) (InVivoMab, BioXCell). The depletion experiment was performed one time (n=6 mice per group).
7
Adenocarcinoma Tumor Model: Depletion and Checkpoint Blockade
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To establish the adenocarcinoma tumor models C57BL/6 WT female mice at 8–10 weeks of age, were intravenously injected with 7 × 104 KP cells or with 2 × 105 KP OVA variants (WT or KOCXCL5, KOCXCL5 (lenti-CXCL5) or KOCXCL5 (lenti-vec)). In one experiment (supplementary Figure 5) mice were challenged with 5 × 104 cells. Otherwise indicated, mice were sacrificed at initial (9 days) or at established stage of tumorigenesis (18 d). To deplete neutrophils in vivo, KP OVA bearing mice were intraperitoneally treated every 3 d, starting from the 6th to 12th d, with 200 µg of anti-Ly6G antibody (InVivo Plus, clone 1A8, Bio X Cell) or isotype control (Rat IgG2a isotype control, clone 2A3, Bio X Cell) and sacrificed at d 13.
In PD-L1 blockade experiments, mice were challenged with KP OVA WT or KOCXCL5 cells, intraperitoneally treated every 3 d with 200 µg of αPD-L1 (InVivoMab, clone 10 F.9G2, BioXcell) or isotype control (InVivoMab, rat IgG2b isotype control, clone LTF-2, BioXcell). Mice were sacrificed at d 18.
In PD-L1 blockade experiments, mice were challenged with KP OVA WT or KOCXCL5 cells, intraperitoneally treated every 3 d with 200 µg of αPD-L1 (InVivoMab, clone 10 F.9G2, BioXcell) or isotype control (InVivoMab, rat IgG2b isotype control, clone LTF-2, BioXcell). Mice were sacrificed at d 18.
8
Isolation and Cryopreservation of PBMCs
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Human B-lymphoma cell lines Farage (EBV+), DB (EBV-), EBV-producing marmoset B-cell line B95–8, and murine B-lymphoma cell line A20 were obtained from American Type Culture Collection (Manassas, VA, USA). Peripheral blood mononuclear cells (PBMCs) were isolated from peripheral blood with ficoll using density gradient centrifugation. Then ficoll medium interface was carefully removed, washed with salt-buffered solution, then centrifuged, leaving purified PBMCs. To freeze, freshly isolated PBMCs were resuspended to 5 × 106 cells/mL in freezing medium containing 10% DMSO and 40% fetal bovine serum (FBS) in RPMI-1640 medium, and placed inside a freezing container at − 80 °C overnight. The following day, samples were moved to a liquid nitrogen tank for long-term storage. Cells were cultured in humidified atmosphere of 95% air and 5% CO2 at 37 °C. Anti-human PD-L1 antibody and anti-human PD-1 antibody were from Innovent (Suzhou, China). Anti-mouse PD-L1 antibody Invivomab was from Bio X Cell (West Lebanon, NH, USA).
9
In Vivo T Cell Depletion and Cxcl10 Neutralization
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In the in vivo T cell subsets depletion study, InVivoMAb anti-mouse CD4 (YTS191, BE0119, BioXCell), CD8α (YTS169.4, BE0117 BioXcell) antibodies were intraperitoneally injected at a dose of 200 μg per mouse. To neutralize Cxcl10 in vivo, 50 µg of anti-mouse Cxcl10 (134013, MAB466, R&D systems) was intraperitoneally injected into mice every 3 days for 2 weeks.
10
Immune Modulation for Retinal Protection
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I) For CD4+ T cell depletion, intraperitoneal injection of Ultra-LEAF™ Purified anti-mouse CD4 Antibody (10 μg/g, clone GK1.5, BioLegend, San Diego, CA, USA) or corresponding isotype antibody (Ultra-LEAF Purified Rat IgG2b, κ Isotype Ctrl Antibody, clone RTK4530, 10 μg/g, BioLegend) was performed in mice on day 15, 25, and 40 PMI. ii) For CXCR3 blockade, intraperitoneal injection of Ultra-LEAF™ Purified anti-mouse CD183 (CXCR3) Antibody (10 μg/g, clone CXCR3-173, BioLegend) or corresponding isotype antibody (Armenian Hamster IgG, clone: HTK888, BioLegend) was performed in mice every 7 days since day 15 PMI. iii) For systemic IFN-g blocking, mice were injected i.p. with 200 mg of anti-IFN-g antibody (InVivoMAb anti-mouse IFNγ, BioXcell, clone XMG1.2) every 3 days since day 15 PMI. An IgG1 isotype antibody (HRPN, BioXcell) was used to serve as control. iv) To block VCAM-1, a InVivoMAb anti-mouse VCAM-1 (10 mg/kg, BioXcell; corresponding isotype antibody: InVivoMAb rat IgG1 isotype control, clone HRPN, BioXcell) was administrated intravenously in MB-injected mice every 3 days since day 15 PMI. For retinal evaluation in abovementioned in vivo antibody intervention experiments, one retina from each recipient mouse was randomly selected and used for RGC density, while the other was for Iba1 and GFAP staining.
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