Journal: Methods (San Diego, Calif.)

Article Title: Isolation and analysis of rare cells in the blood of cancer patients using a negative depletion methodology

doi: 10.1016/j.ymeth.2013.09.006

Figure Lengend Snippet: Flow chart of the CD45-based negative depletion and analysis pathway.

Article Snippet: The immunomagnetic labeling protocol involves two reagents: a CD45 tetrameric antibody complex (TAC) (Stem Cell Technologies) and dextran-coated magnetic nanoparticles (Stem Cell Technologies).

Techniques:

Journal: Methods (San Diego, Calif.)

Article Title: Isolation and analysis of rare cells in the blood of cancer patients using a negative depletion methodology

doi: 10.1016/j.ymeth.2013.09.006

Figure Lengend Snippet: Table of primary antibodies.

Article Snippet: The immunomagnetic labeling protocol involves two reagents: a CD45 tetrameric antibody complex (TAC) (Stem Cell Technologies) and dextran-coated magnetic nanoparticles (Stem Cell Technologies).

Techniques:

Journal: Methods (San Diego, Calif.)

Article Title: Isolation and analysis of rare cells in the blood of cancer patients using a negative depletion methodology

doi: 10.1016/j.ymeth.2013.09.006

Figure Lengend Snippet: List of CD45 antibody clones, fluoroprobes, and manufacturers tested.

Article Snippet: The immunomagnetic labeling protocol involves two reagents: a CD45 tetrameric antibody complex (TAC) (Stem Cell Technologies) and dextran-coated magnetic nanoparticles (Stem Cell Technologies).

Techniques: Clone Assay

Journal: Methods (San Diego, Calif.)

Article Title: Isolation and analysis of rare cells in the blood of cancer patients using a negative depletion methodology

doi: 10.1016/j.ymeth.2013.09.006

Figure Lengend Snippet: (A) Histograms of various anti-CD45 antibody clones, based on FCM analysis of peripheral blood leukocytes, both before (blue) and after (red) immunomagnetic labeling with anti-CD45-TAC/magnetic nanoparticles. Unstained controls are shown in orange. (B) Bar graph comparing the percentage of cells positive for each clone of anti-CD45 antibody, based on FCM analysis, both before and after immunomagnetic labeling.

Article Snippet: The immunomagnetic labeling protocol involves two reagents: a CD45 tetrameric antibody complex (TAC) (Stem Cell Technologies) and dextran-coated magnetic nanoparticles (Stem Cell Technologies).

Techniques: Clone Assay, Labeling

Journal: Methods (San Diego, Calif.)

Article Title: Isolation and analysis of rare cells in the blood of cancer patients using a negative depletion methodology

doi: 10.1016/j.ymeth.2013.09.006

Figure Lengend Snippet: Four-color images of a cytospin stained for DAPI, CK, CD45, and EpCAM of (a–e) leukocytes, (f–j) breast cancer cell line MCF7, and (k–o) an enriched peripheral blood sample from a metastatic breast cancer patient. The first column is a combined image of the four colors and the remaining columns (left to right) are DAPI, anti-CK-AF488, anti-CD45-AF594, and anti-EpCAM-AF647, respectively. For the patient sample, Cell #1 is CK+CD45+EpCAM− and Cell #2 is CK−CD45+EpCAM+.

Article Snippet: The immunomagnetic labeling protocol involves two reagents: a CD45 tetrameric antibody complex (TAC) (Stem Cell Technologies) and dextran-coated magnetic nanoparticles (Stem Cell Technologies).

Techniques: Staining

Journal: Methods (San Diego, Calif.)

Article Title: Isolation and analysis of rare cells in the blood of cancer patients using a negative depletion methodology

doi: 10.1016/j.ymeth.2013.09.006

Figure Lengend Snippet: (A) Four-color images of a cytospin stained for DAPI, CK, CD45, and VIM of (a–e) leukocytes, and (f–j) an enriched peripheral blood sample from a cancer patient. The columns (left to right) are DAPI, anti-CK-AF488, anti-CD45-AF594, anti-VIM-AF647, and a combined image of the four colors, respectively. (B) Four-color image of a cytospin stained for DAPI, CK, Vim, and N-CAD of an enriched peripheral blood sample from a cancer patient. (a) DAPI; (b) anti-CK-AF488; (c) anti-Vim-AF555; (d) anti-N-CAD-AF633 and (e) combined image of the four colors. The cells in the yellow boxes are CK+Vim+N-CAD+. Reproduced with permission from Balasubramanian et al. [15].

Article Snippet: The immunomagnetic labeling protocol involves two reagents: a CD45 tetrameric antibody complex (TAC) (Stem Cell Technologies) and dextran-coated magnetic nanoparticles (Stem Cell Technologies).

Techniques: Staining

Journal: Methods (San Diego, Calif.)

Article Title: Isolation and analysis of rare cells in the blood of cancer patients using a negative depletion methodology

doi: 10.1016/j.ymeth.2013.09.006

Figure Lengend Snippet: Four-color image of a cytospin stained for DAPI, CK, CD45, and HER2 of an enriched peripheral blood sample from a metastatic breast cancer patient. (a) DAPI; (b) anti-CK-AF488; (c) anti-CD45-AF594; (d) anti-HER2-AF647 and (e) combined image of the four colors. The cell in this figure is CK+CD45−HER2+, a HER2-expressing CTC.

Article Snippet: The immunomagnetic labeling protocol involves two reagents: a CD45 tetrameric antibody complex (TAC) (Stem Cell Technologies) and dextran-coated magnetic nanoparticles (Stem Cell Technologies).

Techniques: Staining, Expressing

Journal: Toxicology mechanisms and methods

Article Title: Activation of Protein Kinase C and Protein Kinase D in Human Natural Killer Cells: Effects of Tributyltin, Dibutyltin, and Tetrabromobisphenol A

doi: 10.3109/15376516.2015.1070226

Figure Lengend Snippet: Effects of exposures to TBBPA on the phosphorylation/activation state of PKC and PKD in NK cells: Representative experiment for 10 min exposures to TBBPA. Experiment was repeated using cells from seven different donors.

Article Snippet: NK cells were purified by mixing 30–40 mL of buffy coat with 0.6 mL of RosetteSep human NK cell enrichment antibody cocktail (Stem Cell Technologies, Vancouver, BC, Canada).

Techniques: Activation Assay

Journal: Toxicology mechanisms and methods

Article Title: Activation of Protein Kinase C and Protein Kinase D in Human Natural Killer Cells: Effects of Tributyltin, Dibutyltin, and Tetrabromobisphenol A

doi: 10.3109/15376516.2015.1070226

Figure Lengend Snippet: Effects of incubations with K562 target cells on phosphorylation/activation state of PKC in NK cells: A) levels of phospho-PKC normalized to the control. B) Representative Western blot. Experiments were repeated using cells from five different donors. Asterisks indicate a significant difference as compared with control, p<0.05.

Article Snippet: NK cells were purified by mixing 30–40 mL of buffy coat with 0.6 mL of RosetteSep human NK cell enrichment antibody cocktail (Stem Cell Technologies, Vancouver, BC, Canada).

Techniques: Activation Assay, Western Blot

Journal: Toxicology mechanisms and methods

Article Title: Activation of Protein Kinase C and Protein Kinase D in Human Natural Killer Cells: Effects of Tributyltin, Dibutyltin, and Tetrabromobisphenol A

doi: 10.3109/15376516.2015.1070226

Figure Lengend Snippet: Effects of incubations with K562 target cells on phosphorylation/activation state of PKD in NK cells: A) levels of phospho-PKD normalized to the control. B) Representative Western blot. Experiments were repeated using cells from five different donors. Asterisks indicate a significant difference as compared with control, p<0.05.

Article Snippet: NK cells were purified by mixing 30–40 mL of buffy coat with 0.6 mL of RosetteSep human NK cell enrichment antibody cocktail (Stem Cell Technologies, Vancouver, BC, Canada).

Techniques: Activation Assay, Western Blot

Journal: Toxicology mechanisms and methods

Article Title: Activation of Protein Kinase C and Protein Kinase D in Human Natural Killer Cells: Effects of Tributyltin, Dibutyltin, and Tetrabromobisphenol A

doi: 10.3109/15376516.2015.1070226

Figure Lengend Snippet: Effects of exposures to TBT on the phosphorylation/activation state of PKC in NK cells: A) levels of phospho-PKC normalized to the control for 10 min exposures to TBT. B) Representative experiment for 10 min exposures to TBT. C) Representative experiment for 1 h exposures to TBT. D) Representative experiment for 6 h exposures to TBT. Experiments were repeated using cells from five different donors. Asterisks indicate a significant difference as compared with control, p<0.05.

Article Snippet: NK cells were purified by mixing 30–40 mL of buffy coat with 0.6 mL of RosetteSep human NK cell enrichment antibody cocktail (Stem Cell Technologies, Vancouver, BC, Canada).

Techniques: Activation Assay

Journal: Toxicology mechanisms and methods

Article Title: Activation of Protein Kinase C and Protein Kinase D in Human Natural Killer Cells: Effects of Tributyltin, Dibutyltin, and Tetrabromobisphenol A

doi: 10.3109/15376516.2015.1070226

Figure Lengend Snippet: Effects of exposures to TBT on the phosphorylation/activation state of PKD in NK cells: A) levels of phospho-PKD normalized to the control for 10 min exposures to TBT. B) Representative experiment for 10 min exposures to TBT. C) Representative experiment for 1 h exposures to TBT. D) Representative experiment for 6 h exposures to TBT. Experiments were repeated using cells from five different donors. Asterisks indicate a significant difference as compared with control, p<0.05.

Article Snippet: NK cells were purified by mixing 30–40 mL of buffy coat with 0.6 mL of RosetteSep human NK cell enrichment antibody cocktail (Stem Cell Technologies, Vancouver, BC, Canada).

Techniques: Activation Assay

Journal: Toxicology mechanisms and methods

Article Title: Activation of Protein Kinase C and Protein Kinase D in Human Natural Killer Cells: Effects of Tributyltin, Dibutyltin, and Tetrabromobisphenol A

doi: 10.3109/15376516.2015.1070226

Figure Lengend Snippet: Effects of exposures to DBT on the phosphorylation/activation state of PKC in NK cells: A) levels of phospho-PKC normalized to the control for 10 min exposures to DBT B) Representative experiment for 10 min exposures to DBT. C) levels of phospho-PKC normalized to the control for the 1 h exposures to DBT. D) Representative experiment for 1 h exposures to DBT. E) Representative experiment for 6 h exposures to DBT. Experiments were repeated using cells from six different donors for 10 min and 1h and form three different donors for 6 h. Asterisks indicate a significant difference as compared with control, p<0.05.

Article Snippet: NK cells were purified by mixing 30–40 mL of buffy coat with 0.6 mL of RosetteSep human NK cell enrichment antibody cocktail (Stem Cell Technologies, Vancouver, BC, Canada).

Techniques: Activation Assay

Journal: Toxicology mechanisms and methods

Article Title: Activation of Protein Kinase C and Protein Kinase D in Human Natural Killer Cells: Effects of Tributyltin, Dibutyltin, and Tetrabromobisphenol A

doi: 10.3109/15376516.2015.1070226

Figure Lengend Snippet: Effects of exposures to DBT on the phosphorylation/activation state of PKD in NK cells: A) levels of phospho-PKD normalized to the control for 10 min exposures to DBT. B) Representative experiment for 10 min exposures to DBT. C) Representative experiment for 1 h exposures to DBT. D) Representative experiment for 6 h exposures to DBT. Experiments were repeated using cells from five different donors (10 min), six different donors (1 h) and three different donors (6 h). Asterisks indicate a significant difference as compared with control, p<0.05.

Article Snippet: NK cells were purified by mixing 30–40 mL of buffy coat with 0.6 mL of RosetteSep human NK cell enrichment antibody cocktail (Stem Cell Technologies, Vancouver, BC, Canada).

Techniques: Activation Assay