Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Activation of the intracellular pattern-recognition receptor NOD2 promotes acute myeloid leukemia cell apoptosis and provides a survival advantage in an animal model of AML

doi: 10.4049/jimmunol.1900885

Figure Lengend Snippet: A) Peripheral blood monocytes from healthy donors were stimulated with 10 μg/mL MTP-PE and/or 10 ng/mL IFN-γ for 24 hours; TNFα levels were measured by ELISA (n=3). B) MOLM-13 cells were stimulated with 10 μg/mL MTP-PE and/or 10 ng/mL IFN-γ for 48 hours; subsequently, TNFα levels measured by ELISA (n=3). C) MTP-PE internalization and NOD2 signaling pathway. D) Expression of NOD2 pathway members were extracted from gene arrays from healthy-donor peripheral blood monocytes treated with/without 25 ng/mL IFN-γ, as described earlier (17). E) Primary blasts from AML patients (n=7) were treated with/without 10 ng/mL IFN-γ for 24 hours; transcript levels were measured by qPCR. F) MOLM-13 cells were treated with/without 10 ng/mL IFN-γ for 24 hours. Whole cell lysates were separated by western blots and probed for the molecules indicated; membranes were reprobed with a calreticulin antibody as a loading control (n=3). * denotes p≤0.05; ** p≤0.01.

Article Snippet: Enzyme-linked Immunosorbent Assay (ELISA) Supernatants from cell cultures treated with/without 10 μg/mL MTP-PE and/or 10 ng/mL IFN-γ were analyzed at 24 or 48 hours using Human TNF-alpha DuoSet ELISA (R&D Systems) according to manufacturer’s instruction.

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Western Blot, Control

Journal:

Article Title: Comparative analysis of MVA-CD40L and MVA-TRICOM vectors for enhancing the immunogenicity of chronic lymphocytic leukemia (CLL) cells

doi: 10.1016/j.leukres.2009.12.013

Figure Lengend Snippet: Expression of CD40L and TRICOM molecules following infection of CLL cells with MVA-TRICOM or MVA-CD40L

Article Snippet: For antibody blocking experiments, CLL cells were pre-treated with purified antibody to human CD40L (Clone 24-31; eBioscience, San Diego, CA) or control mouse IgG1 (Serotec, Inc., Raleigh, NC) at a concentration of 10 μg/mL for 1 h at 37°C, then infected with MVA as above in the presence of the same concentration of antibody.

Techniques: Expressing, Infection

Journal:

Article Title: Comparative analysis of MVA-CD40L and MVA-TRICOM vectors for enhancing the immunogenicity of chronic lymphocytic leukemia (CLL) cells

doi: 10.1016/j.leukres.2009.12.013

Figure Lengend Snippet: Comparison of number of CLL patient samples exhibiting upregulation of costimulatory molecules following infection with MVA-TRICOM or MVA-CD40L

Article Snippet: For antibody blocking experiments, CLL cells were pre-treated with purified antibody to human CD40L (Clone 24-31; eBioscience, San Diego, CA) or control mouse IgG1 (Serotec, Inc., Raleigh, NC) at a concentration of 10 μg/mL for 1 h at 37°C, then infected with MVA as above in the presence of the same concentration of antibody.

Techniques: Infection

Journal:

Article Title: Comparative analysis of MVA-CD40L and MVA-TRICOM vectors for enhancing the immunogenicity of chronic lymphocytic leukemia (CLL) cells

doi: 10.1016/j.leukres.2009.12.013

Figure Lengend Snippet: CLL cells from 4 patients were infected with MVA-WT, MVA-TRICOM, or MVA-CD40L. Following 24 hours of infection, CLL cells were washed, irradiated, and cocultured with allogeneic CD3+ T cells from a healthy donor at a ratio of effector to stimulator cells of 10:1. Error bars indicate the standard deviation of replicate measurements. Note that, due to variability in responses, different scales were used on the graphs.

Article Snippet: For antibody blocking experiments, CLL cells were pre-treated with purified antibody to human CD40L (Clone 24-31; eBioscience, San Diego, CA) or control mouse IgG1 (Serotec, Inc., Raleigh, NC) at a concentration of 10 μg/mL for 1 h at 37°C, then infected with MVA as above in the presence of the same concentration of antibody.

Techniques: Infection, Irradiation, Standard Deviation

Journal:

Article Title: Comparative analysis of MVA-CD40L and MVA-TRICOM vectors for enhancing the immunogenicity of chronic lymphocytic leukemia (CLL) cells

doi: 10.1016/j.leukres.2009.12.013

Figure Lengend Snippet: (A) CLL cells were infected with MVA-WT or MVA-CD40L. Following 24 hours of infection, CLL cells were analyzed by flow cytometry for expression of B7-1, ICAM-1, and LFA-3 by CD40L-expressing and non-expressing populations. The plots shown were gated on CD19+ B cells. After infection with MVA-CD40L, 3 populations of CLL cells that expressed negative, low, or high levels of CD40L were observed; these are designated as “negative” (N), “low MFI” (L), and “high MFI” (H) populations on the plots shown. (B) CLL cells were treated with blocking antibody (anti-CD40L or isotype-control mouse IgG1) before and during infection with MVA-CD40L.

Article Snippet: For antibody blocking experiments, CLL cells were pre-treated with purified antibody to human CD40L (Clone 24-31; eBioscience, San Diego, CA) or control mouse IgG1 (Serotec, Inc., Raleigh, NC) at a concentration of 10 μg/mL for 1 h at 37°C, then infected with MVA as above in the presence of the same concentration of antibody.

Techniques: Infection, Flow Cytometry, Expressing, Blocking Assay

Journal:

Article Title: Comparative analysis of MVA-CD40L and MVA-TRICOM vectors for enhancing the immunogenicity of chronic lymphocytic leukemia (CLL) cells

doi: 10.1016/j.leukres.2009.12.013

Figure Lengend Snippet: CLL cells from 6 patients were infected with MVA-WT, MVA-TRICOM, or MVA-CD40L. Following 24 hours of infection, CLL cells were washed, irradiated, and cocultured with autologous CD3+ T cells at a ratio of effector to stimulator cells of 2.5:1. Error bars indicate the standard deviation of replicate measurements. Note that, due to variability in responses, different scales were used on the graphs. Costimulatory molecule expression by the CLL cells following infection is shown in Table 1.

Article Snippet: For antibody blocking experiments, CLL cells were pre-treated with purified antibody to human CD40L (Clone 24-31; eBioscience, San Diego, CA) or control mouse IgG1 (Serotec, Inc., Raleigh, NC) at a concentration of 10 μg/mL for 1 h at 37°C, then infected with MVA as above in the presence of the same concentration of antibody.

Techniques: Infection, Irradiation, Standard Deviation, Expressing

Journal:

Article Title: Comparative analysis of MVA-CD40L and MVA-TRICOM vectors for enhancing the immunogenicity of chronic lymphocytic leukemia (CLL) cells

doi: 10.1016/j.leukres.2009.12.013

Figure Lengend Snippet: CLL cells from 4 patients were infected with MVA-WT, MVA-TRICOM, or MVA-CD40L. Following 24 hours of infection, CLL cells were washed and, without irradiation, cocultured with autologous CD3+ T cells at a ratio of effector to stimulator cells of 2.5:1. Proliferation by the CLL cells alone is indicated by white bars; proliferation by T cells and CLL cells together is indicated by black bars. Error bars indicate the standard deviation of replicate measurements. Note that, due to variability in responses, different scales were used on the graphs.

Article Snippet: For antibody blocking experiments, CLL cells were pre-treated with purified antibody to human CD40L (Clone 24-31; eBioscience, San Diego, CA) or control mouse IgG1 (Serotec, Inc., Raleigh, NC) at a concentration of 10 μg/mL for 1 h at 37°C, then infected with MVA as above in the presence of the same concentration of antibody.

Techniques: Infection, Irradiation, Standard Deviation