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

Article Title: Soluble Ig-like transcript 3 inhibits tumor allograft rejection in humanized SCID mice and T cell responses in cancer patients.

doi: 10.4049/jimmunol.178.11.7432

Figure Lengend Snippet: FIGURE 1. Effect of ILT3 on tumor allografts. sILT3 and mILT3 induce tolerance to allogeneic tumors in hu-SCID mice. A, Tumor growth was assessed in 5 groups, each consisting of 10 hu-SCID mice transplanted with KG1 and treated with rILT3-Fc (), human IgG (), not treated (E), and transplanted with empty vector-transfected KG1.MIG (‚) or ILT3-transfected KG1 (Œ). B, Engraftment of human T cells in blood, lymph nodes, and spleen of hu-SCID mice. HLA class I CD34 tumor metastasis in regional lymph nodes was seen in mice with KG1 and KG1.ILT3 tumors. C, Immunohistochemical staining of CD4 and CD8 human T cells infiltrating KG1 tumor growing in hu-SCID mice treated with rILT3-Fc. D, Growth of melanoma and pancreatic carcinoma cell lines in hu-SCID mice treated or not treated with rILT3-Fc.

Article Snippet: After 1 h of incubation at room temperature, the plate was washed and 100 l of anti-ILT3 biotinylated polyclonal Ab (R&D Systems) was added at a concentration of 33 ng/ml to each well, followed by 1-h incubation at room temperature.

Techniques: Plasmid Preparation, Transfection, Immunohistochemical staining, Staining

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

Article Title: Soluble Ig-like transcript 3 inhibits tumor allograft rejection in humanized SCID mice and T cell responses in cancer patients.

doi: 10.4049/jimmunol.178.11.7432

Figure Lengend Snippet: FIGURE 4. Detection of ILT3 in human sera. A, Sera from cancer patients and healthy blood donors were tested for sILT3 by ELISA. B, Healthy donor T cells were tested for MLC reactivity to irradiated KG1-stimulating cells in cultures containing pooled (ILT3-negative) sera from healthy controls or ILT3-positive sera from individual cancer patients (n 29) in the presence or absence of anti-ILT3 mAb or mouse IgG. C, The MLC-inhibitory activity of a cancer patient’s serum was tested before and after depletion of sILT3. D, CD8 T cells allostimulated in cultures containing ILT3-positive sera inhibited the alloreactivity of autologous, unprimed CD3 T cells in a dose-dependent manner. E, Immunoprecipitation and Western blot analysis of an ILT3-positive serum prior to (lane 4) and after depletion of ILT3 (lane 3) and of two ILT3-negative control sera (lanes 1 and 2).

Article Snippet: After 1 h of incubation at room temperature, the plate was washed and 100 l of anti-ILT3 biotinylated polyclonal Ab (R&D Systems) was added at a concentration of 33 ng/ml to each well, followed by 1-h incubation at room temperature.

Techniques: Enzyme-linked Immunosorbent Assay, Irradiation, Activity Assay, Immunoprecipitation, Western Blot, Negative Control

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

Article Title: Soluble Ig-like transcript 3 inhibits tumor allograft rejection in humanized SCID mice and T cell responses in cancer patients.

doi: 10.4049/jimmunol.178.11.7432

Figure Lengend Snippet: FIGURE 5. Immunohistochemical staining of macrophages with anti- CD68 and anti-ILT3 Abs in patients’ tumors and lymph nodes.

Article Snippet: After 1 h of incubation at room temperature, the plate was washed and 100 l of anti-ILT3 biotinylated polyclonal Ab (R&D Systems) was added at a concentration of 33 ng/ml to each well, followed by 1-h incubation at room temperature.

Techniques: Immunohistochemical staining, Staining

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

Article Title: Soluble Ig-like transcript 3 inhibits tumor allograft rejection in humanized SCID mice and T cell responses in cancer patients.

doi: 10.4049/jimmunol.178.11.7432

Figure Lengend Snippet: FIGURE 6. Identification of alter- natively spliced ILT3 in tumor biop- sies. Schematic representation of the structure of the ILT3 gene and of the mRNA deletion mutant. RT-PCR analysis of ILT3 mRNA in normal PBMC and tumor tissue from patients with colon carcinoma. All spliced variants were cloned and fully se- quenced. Arrows indicate the position of primers; second lane is the nega- tive control with no RNA added.

Article Snippet: After 1 h of incubation at room temperature, the plate was washed and 100 l of anti-ILT3 biotinylated polyclonal Ab (R&D Systems) was added at a concentration of 33 ng/ml to each well, followed by 1-h incubation at room temperature.

Techniques: Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Clone Assay, Control

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

Article Title: Soluble Ig-like transcript 3 inhibits tumor allograft rejection in humanized SCID mice and T cell responses in cancer patients.

doi: 10.4049/jimmunol.178.11.7432

Figure Lengend Snippet: FIGURE 7. Alternatively spliced mRNA encodes sILT3 protein. Lentivi- ral vectors were constructed and used to transfect cloned, alternatively spliced ILT3 cDNA into CHO-S and 293T cells. Empty vector and ILT3-Fc con- structs were used as controls. Western blot and ELISA analysis show that de- letion of exons 5–7 from ILT3 mes- sages results in a 45-kDa soluble form of the ILT3 protein.

Article Snippet: After 1 h of incubation at room temperature, the plate was washed and 100 l of anti-ILT3 biotinylated polyclonal Ab (R&D Systems) was added at a concentration of 33 ng/ml to each well, followed by 1-h incubation at room temperature.

Techniques: Construct, Clone Assay, Plasmid Preparation, Western Blot, Enzyme-linked Immunosorbent Assay