BD PharmingenTM APO-BRDUTM Kit

Description

Schematic Representation of APO-BRDU Labeling. The enzyme deoxynucleotidyl transferase TdT catalyzes a template dependent addition of bromolated deoxyuridine triphosphates Br-dUTP to the 3-hydroxyl ends of double- and single-stranded DNA. After Br-dUTP incorporation, DNA break sites are identified by a FITC-labeled anti-BrdU mAb. Flow Cytometry Data of APO-BRDU Negative and Positive Control cells top row . Negative and Positive control cells were incubated with Br-dUTP in the presence of TdT enzyme in order to incorporate Br-dUTP into exposed 3-OH DNA ends. Br-dUTP sites are detected with a FITC-labeled anti-BrdU mAb. Non-apoptotic cells do not incorporate significant amounts of Br-dUTP due to the lack of exposed 3-OH ends, and consequently have relatively little fluorescence compared to apoptotic cells which have an abundance of 3-OH ends. Flow Cytometric Analysis of HPB-ALL Human Leukemia Cells Using APO-BRDU bottom row . HPB-ALL human leukemia cells were left untreated left panel or treated with anti-human Fas mAb, clone DX2 Cat. No. 33450D and Protein G for 2 hr middle panel or 12 hr right panel . Cells were fixed and incubated with Br-dUTP in the presence of TdT enzyme in order to incorporate Br-dUTP into exposed 3-OH DNA ends. Br-dUTP was detected with a FITC-labeled anti-BrdU mAb. Non-apoptotic cells M1 gates do not incorporate significant amounts of Br-dUTP due to lack of exposed 3-OH ends, and consequently have relatively little fluorescence compared to apoptotic cells which have an abundance of 3-OH end M2 gates . DX2-induced, Fas-mediated apoptosis is shown by increases in the number of cells staining with anti-BrdU-FITC mAb M2 gates after 2 and 12 hr. The M1 and M2 gates demarcate non-apoptotic and apoptotic populations, respectively. Flow Cytometer Setup for Becton Dickinson Hardware. Positive control cells were labeled with both PI DNA and FITC-BrdU mAb. Display 1: Non-clumped cells are gated. Gated From Display 1: Separate boxes are drawn around cells that stain positive upper box and negative lower box with the FITC-BrdU mAb. An example of FACScan gain settings is shown. Schematic Representation of APO-BRDU Labeling. The enzyme deoxynucleotidyl transferase TdT catalyzes a template dependent addition of bromolated deoxyuridine triphosphates Br-dUTP to the 3-hydroxyl ends of double- and single-stranded DNA. After Br-dUTP incorporation, DNA break sites are identified by a FITC-labeled anti-BrdU mAb. Show More Flow Cytometry Data of APO-BRDU Negative and Positive Control cells top row . Negative and Positive control cells were incubated with Br-dUTP in the presence of TdT enzyme in order to incorporate Br-dUTP into exposed 3-OH DNA ends. Br-dUTP sites are detected with a FITC-labeled anti-BrdU mAb. Non-apoptotic cells do not incorporate significant amounts of Br-dUTP due to the lack of exposed 3-OH ends, and consequently have relatively little fluorescence compared to apoptotic cells which have an abundance of 3-OH ends. Flow Cytometric Analysis of HPB-ALL Human Leukemia Cells Using APO-BRDU bottom row . HPB-ALL human leukemia cells were left untreated left panel or treated with anti-human Fas mAb, clone DX2 Cat. No. 33450D and Protein G for 2 hr middle panel or 12 hr right panel . Cells were fixed and incubated with Br-dUTP in the presence of TdT enzyme in order to incorporate Br-dUTP into exposed 3-OH DNA ends. Br-dUTP was detected with a FITC-labeled anti-BrdU mAb. Non-apoptotic cells M1 gates do not incorporate significant amounts of Br-dUTP due to lack of exposed 3-OH ends, and consequently have relatively little fluorescence compared to apoptotic cells which have an abundance of 3-OH end M2 gates . DX2-induced, Fas-mediated apoptosis is shown by increases in the number of cells staining with anti-BrdU-FITC mAb M2 gates after 2 and 12 hr. The M1 and M2 gates demarcate non-apoptotic and apoptotic populations, respectively. Show More Flow Cytometer Setup for Becton Dickinson Hardware. Positive control cells were labeled with both PI DNA and FITC-BrdU mAb. Display 1: Non-clumped cells are gated. Gated From Display 1: Separate boxes are drawn around cells that stain positive upper box and negative lower box with the FITC-BrdU mAb. An example of FACScan gain settings is shown. Show More Schematic Representation of APO-BRDU Labeling. The enzyme deoxynucleotidyl transferase TdT catalyzes a template dependent addition of bromolated deoxyuridine triphosphates Br-dUTP to the 3-hydroxyl ends of double- and single-stranded DNA. After Br-dUTP incorporation, DNA break sites are identified by a FITC-labeled anti-BrdU mAb. Flow Cytometry Data of APO-BRDU Negative and Positive Control cells top row . Negative and Positive control cells were incubated with Br-dUTP in the presence of TdT enzyme in order to incorporate Br-dUTP into exposed 3-OH DNA ends. Br-dUTP sites are detected with a FITC-labeled anti-BrdU mAb. Non-apoptotic cells do not incorporate significant amounts of Br-dUTP due to the lack of exposed 3-OH ends, and consequently have relatively little fluorescence compared to apoptotic cells which have an abundance of 3-OH ends. Flow Cytometric Analysis of HPB-ALL Human Leukemia Cells Using APO-BRDU bottom row . HPB-ALL human leukemia cells were left untreated left panel or treated with anti-human Fas mAb, clone DX2 Cat. No. 33450D and Protein G for 2 hr middle panel or 12 hr right panel . Cells were fixed and incubated with Br-dUTP in the presence of TdT enzyme in order to incorporate Br-dUTP into exposed 3-OH DNA ends. Br-dUTP was detected with a FITC-labeled anti-BrdU mAb. Non-apoptotic cells M1 gates do not incorporate significant amounts of Br-dUTP due to lack of exposed 3-OH ends, and consequently have relatively little fluorescence compared to apoptotic cells which have an abundance of 3-OH end M2 gates . DX2-induced, Fas-mediated apoptosis is shown by increases in the number of cells staining with anti-BrdU-FITC mAb M2 gates after 2 and 12 hr. The M1 and M2 gates demarcate non-apoptotic and apoptotic populations, respectively. Flow Cytometer Setup for Becton Dickinson Hardware. Positive control cells were labeled with both PI DNA and FITC-BrdU mAb. Display 1: Non-clumped cells are gated. Gated From Display 1: Separate boxes are drawn around cells that stain positive upper box and negative lower box with the FITC-BrdU mAb. An example of FACScan gain settings is shown.