human annexin ii igg  (Bio-Rad)

Journal: Blood

Article Title: Phagocytosis by macrophages and endothelial cells inhibits procoagulant and fibrinolytic activity of acute promyelocytic leukemia cells.

doi: 10.1182/blood-2011-06-362186

Figure Lengend Snippet: Figure 1. PS exposure of NB4 and APL cells. Cells were costained with PI and either FITC–annexin V or FITC-lactadherin. (A) The cells were incubated with indicated concentrations of DNR for 24 hours. Percentage of annexin V/lactadherin- binding cells was analyzed by flow cytometry. After treatment with 1M DNR, 75% of NB4 cells and 80% of APL cells were positive for lactadherin. (B) The plasma membrane of APL cells displayed green fluorescence when stained by FITC– annexin V (left) or FITC-lactadherin (right). Cell nuclei displayed red fluorescence when labeled by PI. Co-stained areas appeared yellow. Scale bars represent 10 m.

Article Snippet: Goat anti–human annexin II IgG and goat anti–human IgG were from AbD Serotec.

Techniques: Incubation, Binding Assay, Cytometry, Clinical Proteomics, Membrane, Staining, Labeling

Journal: Blood

Article Title: Phagocytosis by macrophages and endothelial cells inhibits procoagulant and fibrinolytic activity of acute promyelocytic leukemia cells.

doi: 10.1182/blood-2011-06-362186

Figure Lengend Snippet: Figure 4. Effect of lactadherin on phagocytosis of target APL cells. A total of 1 106 CMFDA-labeled target APL cells were preincubated with 2nM lactadherin or annexin V for 10 minutes at room temperature, followed by removal of free protein- containing supernatant fluid. These cells were added to 5 105 CMTPX-stained THP-1–derived Ms or HUVECs that were seeded in 12-well culture plates before analyses by flow cytometry. Phagocytosis was quantified by measuring the percent- age of CMFDA (green)–positive red fluorescence (CMTPX) phagocytes. (A) Phago- cytic index was calculated in the absence or presence of lactadherin at indicated times before 2 hours. Lactadherin enhanced the extent of phagocytosis in a time-dependent manner. Asterisk and pound sign (*, #) indicate P .05 from phagocytosis by Ms and ECs without lactadherin, respectively. (B) Phagocytic percentage of 2 hour-incubation was assayed after pretreatment of target APL cells with lactadherin and annexin V separately. Lactadherin enhanced phagocytosis, whereas annexin V decreased engulfment. *P .05.

Article Snippet: Goat anti–human annexin II IgG and goat anti–human IgG were from AbD Serotec.

Techniques: Labeling, Staining, Derivative Assay, Cytometry, Incubation

Journal: Blood

Article Title: Phagocytosis by macrophages and endothelial cells inhibits procoagulant and fibrinolytic activity of acute promyelocytic leukemia cells.

doi: 10.1182/blood-2011-06-362186

Figure Lengend Snippet: Figure 6. Effect of lactadherin on PCA of coincu- bated target APL cells and phagocytes. Target APL cells were preincubated with 2nM lactadherin or an- nexin V for 10 minutes at room temperature. Clotting time (A), intrinsic FXa (B), extrinsic FXa (C), and throm- bin (D) of 1 106 target APL cells, or 1 106 target APL cells opsonized by annexin V or lactadherin, or incuba- tion of 1 106 target cells with phagocytes (THP-1– derived Ms or HUVECs) for 2 hours, or incubation of 1 106 annexin V–opsonized or lactadherin-opsonized target cells with phagocytes for 2 hours were deter- mined. Lactadherin and phagocytes cooperatively in- creased coagulation time and reduced enzyme com- plexes of target APL cells. *P .05 compared with single target APL cells. P .05; #P .01; and **P .001 compared with the mixture ofAPL targets and phagocytes (Ms and ECs separately).

Article Snippet: Goat anti–human annexin II IgG and goat anti–human IgG were from AbD Serotec.

Techniques: Coagulation, Derivative Assay, Incubation

Journal: Blood

Article Title: Phagocytosis by macrophages and endothelial cells inhibits procoagulant and fibrinolytic activity of acute promyelocytic leukemia cells.

doi: 10.1182/blood-2011-06-362186

Figure Lengend Snippet: Figure 7. Effect of phagocytosis on plasmin formation and annexin II expression. (A) Plasmin generation of 1 106 target APL cells, 5 105 phagocytes (THP-1–derived Ms or HUVECs), or incubation of 1 106 target APL cells with 5 105 phagocytes was evaluated at the given times. Plasmin production of the coincubated cells was time dependently reduced. *P .05 compared with 0 hour time point of each group. (B) Plasmin formation of 1 106 target APL cells, with or without 2nM annexin V or lactadherin, and with or without incubation with 5 105 phagocytes (THP-1–derived Ms or HUVECs) after 2 hours was measured. Plasmin formation of 1 106 viableAPL cells is also shown. *P .05 compared with the mixture of target APL cells and phagocytes (Ms and ECs separately). (C) NB4 cells were first labeled with goat anti–human annexin II IgG, and then with an Alexa Fluor 488–conjugated secondary Ab. Annexin II expression on permeabilized untreated NB4 cells (left) and 1M DNR-treated NB4 cells (right) was viewed with confocal microscopy. The cell nuclei were counterlabeled with PI (red), and scale bar represents 10 m. (D) Nonpermeabilized or permeabilized APL cells with and without 1M DNR treatment were stained as in panel C and analyzed by flow cytometry. Annexin II expression of DNR-treated cells decreased compared with untreated viable APL cells. (E) Flow cytometry was used to quantitate annexin II expression on cells that were treated as in panel C. Cells stained with goat anti–human IgG and Alexa Fluor 488–conjugated secondary Ab were used as control (black). The percentage of annexin II–positive viable APL cells (green) and target APL cells (pink) from one patient with permeabilization were 98.3% and 35.1%, respectively (left). Middle panel showed that, compared with controls, annexin II was expressed on the surface of nonpermeabilized THP-1–derived Ms (pink) and more so on permeabilized cells (green). Permeabilized HUVECs showed an increase in annexin II (green), but, compared with controls, nonpermeabilized HUVECs (pink) showed no increase in annexin II (right).

Article Snippet: Goat anti–human annexin II IgG and goat anti–human IgG were from AbD Serotec.

Techniques: Expressing, Derivative Assay, Incubation, Labeling, Confocal Microscopy, Staining, Cytometry, Flow Cytometry, Control