mab3241  (R&D Systems)

Journal: Oncotarget

Article Title: Chemokines CCL2, 3, 14 stimulate macrophage bone marrow homing, proliferation, and polarization in multiple myeloma

doi:

Figure Lengend Snippet: A. qPCR analysis of expression of different MO chemokines in MM patient BM cells. The value indicates the relative expression to GAPDH. One representative sample of 4 patient samples analyzed is shown. B. Levels of CCL3, CCL14, and CCL2, measured by ELISA, in BM plasma of healthy donors (CTR) and patients with MM or MGUS. The numbers of CTR, MGUS and MM patients used for measuring CCL2 are 4, 10, and 23, respectively; for CCL14 are 7, 10, and 11, respectively; and for CCL3 are 7, 10, and 13, respectively. C. Immunohistochemistry analysis of CCL3, CCL14, and CCL2 expression in BM biopsies of 2 healthy donors (CTR1 and CTR2) and 2 representatives (MM1 and MM2) out of five MM patients. D. Linear regression analysis of the relationship between the percentage of BM MΦs and concentration of chemokines CCL14 ( n = 20) and CCL3 ( n = 18) in BM plasma in MM patients. * p < 0.05.

Article Snippet: Neutralizing antibodies to human CCL3 (MAB270), CCL14 (MAB3241) and CCL2 (MAB679), and mouse CCL3 (AB-450-NA), CCL2 (AB-479-NA) were purchased from R&D Systems.

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Clinical Proteomics, Immunohistochemistry, Concentration Assay

Journal: Oncotarget

Article Title: Chemokines CCL2, 3, 14 stimulate macrophage bone marrow homing, proliferation, and polarization in multiple myeloma

doi:

Figure Lengend Snippet: A. qPCR analysis of CCL3, CCL14, and CCL2 expression in CD138 + primary MM cells and CD138 − non-malignant cells in BM aspirates from 3 different MM patients (MM1 to MM3). The y-axis indicates the fold change relative to GAPDH value. B. Concentration of chemokines secreted by 4 human MM cell lines. MM cells (2 ×10 5 cell per well) were cultured in vitro for 24 hours; then the chemokine expression in culture supernatant was analyzed by ELISA. C. BMSCs were cocultured with ARP-1 or MM.1S MM cells for 48 hours. CCL3, CCL14, and CCL2 expression in BMSCs was analyzed by qPCR. D. BMSCs were cultured in transwells with MM cells ARP-1 for 48 hours, followed by removal of MM cells and cultured alone in fresh medium for 24 hours. BMSCs from the same donors were cultured alone in parallel as controls. Chemokine concentrations were determined by ELISA using culture supernatants collected at the end of culture. * p < 0.05, ** p < 0.01.

Article Snippet: Neutralizing antibodies to human CCL3 (MAB270), CCL14 (MAB3241) and CCL2 (MAB679), and mouse CCL3 (AB-450-NA), CCL2 (AB-479-NA) were purchased from R&D Systems.

Techniques: Expressing, Concentration Assay, Cell Culture, In Vitro, Enzyme-linked Immunosorbent Assay

Journal: Oncotarget

Article Title: Chemokines CCL2, 3, 14 stimulate macrophage bone marrow homing, proliferation, and polarization in multiple myeloma

doi:

Figure Lengend Snippet: A. nMΦ or mMΦ were generated as described in the Methods. The cells were then cultured in vitro in the presence of each of the chemokine neutralizing antibodies (αCCL3, αCCL14 and αCCL2, at a final concentration of 10 μg/ml each), and their combination (MIX). An equal amount of IgG was used as control. Cell proliferation at different time points was examined by MTS. B. nMΦs were cultured for 3 days in the presence of each of the recombinant chemokines (final concentration of 1 μg/ml) or their combination (MIX). mMΦs cultured in medium alone were used as a positive control. Cell proliferation at different time points was determined by MTS assay. C. Western blot showing the levels of p27Kip1 and cyclin D1 in nMΦs and mMΦs from 2 healthy donors (Do1 and Do2). D. Western blot showing the levels of p27Kip1 and cyclin D1 in nMΦs and mMΦs. mMΦs were generated in coculture with MM cells in transwells in the absence (CTR) or presence of control IgG or a combination of the chemokine neutralizing antibodies (MIX). E. Western blot showing the levels of different kinases, IL-6, c-myc, cyclin D1, and p27Kip1 in nMΦs in overnight cultures in medium or with addition of each of the chemokines CCL2, CCL14, or CCL3 individually or all three combined (MIX). mMΦs were used as a positive control. F. Proliferative response of mMΦs in medium alone (CTR) or in the presence of the PI3K-Akt inhibitor LY294002 (LY; 50 μM) or Erk1/2 inhibitor U0126 (UO; 2 μM). * p < 0.05, ** p < 0.01.

Article Snippet: Neutralizing antibodies to human CCL3 (MAB270), CCL14 (MAB3241) and CCL2 (MAB679), and mouse CCL3 (AB-450-NA), CCL2 (AB-479-NA) were purchased from R&D Systems.

Techniques: Generated, Cell Culture, In Vitro, Concentration Assay, Control, Recombinant, Positive Control, MTS Assay, Western Blot