mouse monoclonal antibody mab  (Developmental Studies Hybridoma Bank)

Journal:

Article Title: Optimization of immunoprecipitation-western blot analysis in detecting GW182-associated components of GW/P bodies

doi: 10.1038/nprot.2009.34

Figure Lengend Snippet: Summary of the steps involved and the approximate time required in detecting GW182 and associated proteins that are components of GWBs. Note that the steps shown in the boxes on the left are required only the first time if sufficient reagent is prepared for future experiments.

Article Snippet: Mouse GW182 (2D6) monoclonal antibody (Santa Cruz Biotechnology Inc., cat. no. sc-sc-56313) may also be used in IP and WB applications; however, this antibody tends to be weaker and more protein must be loaded onto a gel when probing extracts or a greater volume of antibody must be used when coupling antibody to sepharose beads.

Techniques:

Journal:

Article Title: Optimization of immunoprecipitation-western blot analysis in detecting GW182-associated components of GW/P bodies

doi: 10.1038/nprot.2009.34

Figure Lengend Snippet: Antibodies to GW182-immunoprecipitated Ago2, Dicer and XRN1 proteins from HeLa cell extracts as examined by WB analysis. GW182 and associated proteins were immunoprecipitated using human sera 18033 containing antibodies to GW182 coupled with Protein A sepharose beads and by mouse anti-GW182 monoclonal 4B6 coupled to Protein G sepharose beads. Normal human serum (NHS), IC6 human serum and mouse monoclonal antibody to golgin 97 were used as IP controls. Immunopreciptates were carried out with buffer that did not contain detergent. Proteins (40 μg) were resolved on a 10% SDS-PAGE gel and were detected with a 4B6 monoclonal antibody (a) or 18033 human anti-GWB serum (b). Immunopreciptates were carried out with buffer that contained 0.3% NP-40 detergent. Proteins (40 μg) were resolved on a 6.5% SDS-PAGE gel and were detected using a mouse anti-Ago2 monoclonal 4F9 antibody (c). Protein (40 μg) from whole cell extracts or 18033 immunoprecipitates carried out using 0.3% NP-40 buffer were separated on a 6.5% SDS-PAGE gel and probed with NHS or antibodies to golgin 97, GW182, Ago2, XRN1 or Dicer. (d) Mouse monoclonal antibody to Dicer and rabbit anti-XRN1 antibody were purchased from Abcam Inc. (cat. no. ab14601) and Bethyl Laboratories (cat. no. A300-443A), respectively. Mouse monoclonal 4F9 to recombinant hAgo2 was obtained as described previously25.

Article Snippet: Mouse GW182 (2D6) monoclonal antibody (Santa Cruz Biotechnology Inc., cat. no. sc-sc-56313) may also be used in IP and WB applications; however, this antibody tends to be weaker and more protein must be loaded onto a gel when probing extracts or a greater volume of antibody must be used when coupling antibody to sepharose beads.

Techniques: Immunoprecipitation, SDS Page, Recombinant

Journal: Gene Expression

Article Title: Overexpression of CYP2D6 Attenuates the Toxicity of MPP + in Actively Dividing and Differentiated PC12 Cells

doi:

Figure Lengend Snippet: Expression of CYP2D6 in clonal PC12 cells. (a) Immu-noblot analysis of CYP2D6 expression was performed with anti-CYP2D6 antibody. The results of representative cell lines of mock (lane 1) and CYP2D6 (lane 2), and also that of COS7 cells (monkey; kidney, epithelial) transiently transfected with CYP2D6 cDNA (lane 3) are shown. The bands for endogenous or transfec-ted CYP2D6 were detected at 65 kDa (arrow I) and 62 kDa (arrow II), respectively. (b–d) Clonal cells of CYP2D6 were immuno-stained with anti-CYP2D6 antibody. CYP2D6 (b) and ER-Track-erTM Blue-White DPX (c) are colocalized intracellularly (d). Scale bar: 50 μM.

Article Snippet: After a 20-day incubation with Geneticin ® , the resistant colonies were examined by immunoblotting with mouse monoclonal antibody against human CYP2D6, MAB-2D6 (BD GentestTM, Discovery Labware, Inc., Woburn, MA, USA, a subsidiary of Becton, Dickinson and Company), for selection as CYP2D6-expressing clones.

Techniques: Expressing, Transfection, Staining

Journal: Gene Expression

Article Title: Overexpression of CYP2D6 Attenuates the Toxicity of MPP + in Actively Dividing and Differentiated PC12 Cells

doi:

Figure Lengend Snippet: Enzyme activity of CYP2D6 in clonal PC12 cells. Enzyme activity of CYP2D6 was assayed by HPLC with [guanidine-14C]debrisoquine as substrate. Profiles of negative control (before incubation) (a), positive control (after a 3-h incubation with recombinant CYP2D6) (b), and those after a 3-h incubation with microsomal fraction of clonal cells of mock (c) and CYP2D6 (d) are shown.

Article Snippet: After a 20-day incubation with Geneticin ® , the resistant colonies were examined by immunoblotting with mouse monoclonal antibody against human CYP2D6, MAB-2D6 (BD GentestTM, Discovery Labware, Inc., Woburn, MA, USA, a subsidiary of Becton, Dickinson and Company), for selection as CYP2D6-expressing clones.

Techniques: Activity Assay, Negative Control, Incubation, Positive Control, Recombinant

Journal: Gene Expression

Article Title: Overexpression of CYP2D6 Attenuates the Toxicity of MPP + in Actively Dividing and Differentiated PC12 Cells

doi:

Figure Lengend Snippet: CYTOTOXICITY OF MPP + AND MPTP ON PC12 CELLS

Article Snippet: After a 20-day incubation with Geneticin ® , the resistant colonies were examined by immunoblotting with mouse monoclonal antibody against human CYP2D6, MAB-2D6 (BD GentestTM, Discovery Labware, Inc., Woburn, MA, USA, a subsidiary of Becton, Dickinson and Company), for selection as CYP2D6-expressing clones.

Techniques: Transfection

Journal: Gene Expression

Article Title: Overexpression of CYP2D6 Attenuates the Toxicity of MPP + in Actively Dividing and Differentiated PC12 Cells

doi:

Figure Lengend Snippet: Mitochondrial activity to reduce MTT after exposure to MPP+ or MPTP. Clonal cells of CYP2D6 and mock, undifferenti-ated or differentiated by NGF, were incubated with MPP+ (a) and MPTP (c) for 24 h. The activity is expressed as percentage of the mean value of control (vehicle alone). Each point represents the mean value ± SEM (n = 4) for mock and 10 for CYP2D6. *p < 0.05 by unpaired Student’s t -test for mock versus CYP2D6. (b) MTT-reducing activity after exposure to MPP+ and percentage of CYP2D6-positive cells are positively correlated. The regression line is y = 0.78x + 44.51 (r = 0.736).

Article Snippet: After a 20-day incubation with Geneticin ® , the resistant colonies were examined by immunoblotting with mouse monoclonal antibody against human CYP2D6, MAB-2D6 (BD GentestTM, Discovery Labware, Inc., Woburn, MA, USA, a subsidiary of Becton, Dickinson and Company), for selection as CYP2D6-expressing clones.

Techniques: Activity Assay, Incubation, Control

Journal: Gene Expression

Article Title: Overexpression of CYP2D6 Attenuates the Toxicity of MPP + in Actively Dividing and Differentiated PC12 Cells

doi:

Figure Lengend Snippet: Mitochondrial ROS generation and CYP2D6 expression. Mitochondrial ROS generation in CYP2D6-transfected PC12 cells was visualized by oxidized chloromethyltetramethylrosamine after a 24-h exposure to 20 μM MPP+ (a) or 500 μM MPTP (d). Overexpression of CYP2D6 was detected with anti-CYP2D6 antibody (b, e). The merged images of mitochondrial ROS generation and CYP2D6 expression are shown (c, f). Scale bar: 40 μm.

Article Snippet: After a 20-day incubation with Geneticin ® , the resistant colonies were examined by immunoblotting with mouse monoclonal antibody against human CYP2D6, MAB-2D6 (BD GentestTM, Discovery Labware, Inc., Woburn, MA, USA, a subsidiary of Becton, Dickinson and Company), for selection as CYP2D6-expressing clones.

Techniques: Expressing, Transfection, Over Expression