Red Fluorescent SR-VAD-OPH in vitro Poly Caspase Apoptosis Detection Reagent

O-phenoxy (OPH) inhibitors form a stable covalent thioether adduct with the reactive -SH group of active caspase enzymes. SR-VAD-OPH poly caspase inhibitor can be used with a fluorescence microscope, fluorescent plate reader, or flow cytometry.

Figure 1. Jurkat cells were treated with staurosporine (Positive, Induced, right histogram). Negative control cells were spiked with a DMSO vehicle control (Negative, Non-induced, left histogram). Cells were stained with SR-VAD-OPH for 1 hour, washed twice, and read on a flow cytometer. Only 8.0% of control cells displayed caspase activity, whereas staurosporine induced caspase activity in 90.3% of the experimental cells. This is a ratio of 11:1. Data courtesy of Ms. Tracy Hanson, ICT 092713.
Figure 2. Jurkat cells were treated with staurosporine to induce apoptosis, then labeled with SR-VAD-OPH for 60 minutes. Cells were washed twice and wet-mount slides were prepared. A grey DIC image (left) was taken, and red fluorescence (right) was detected using a band pass filter (excitation at 565 nm, emission at 590-600 nm). SR-VAD-OPH reveals complete caspase activation in the form of red fluorescence in all cells. Data courtesy of Ms. Tracy Hanson, ICT (ICT111809; SR_OPH_111_17_18).
Figure 3. Jurkat cells were treated with DMSO (left) or staurosporine (right), then labeled with SR-VAD-OPH, followed by Green Live/Dead Stain (Cat. 6342). Four populations of cells were detected: unstained live cells (lower left); red early apoptotic cells stained with SR-VAD-OPH (lower right); late apoptotic cells (upper right) are dually stained with SR-VAD-OPH and Green Live/Dead Stain; and green necrotic cells (upper left). Data courtesy of Tracy Hanson, ICT (ICT-202:97; 091015).
Figure 1. Jurkat cells were treated with staurosporine (Positive, Induced, right histogram). Negative control cells were spiked with a DMSO vehicle control (Negative, Non-induced, left histogram). Cells were stained with SR-VAD-OPH for 1 hour, washed twice, and read on a flow cytometer. Only 8.0% of control cells displayed caspase activity, whereas staurosporine induced caspase activity in 90.3% of the experimental cells. This is a ratio of 11:1. Data courtesy of Ms. Tracy Hanson, ICT 092713.
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Figure 1. Jurkat cells were treated with staurosporine (Positive, Induced, right histogram). Negative control cells were spiked with a DMSO vehicle control (Negative, Non-induced, left histogram). Cells were stained with SR-VAD-OPH for 1 hour, washed twice, and read on a flow cytometer. Only 8.0% of control cells displayed caspase activity, whereas staurosporine induced caspase activity in 90.3% of the experimental cells. This is a ratio of 11:1. Data courtesy of Ms. Tracy Hanson, ICT 092713.
Figure 2. Jurkat cells were treated with staurosporine to induce apoptosis, then labeled with SR-VAD-OPH for 60 minutes. Cells were washed twice and wet-mount slides were prepared. A grey DIC image (left) was taken, and red fluorescence (right) was detected using a band pass filter (excitation at 565 nm, emission at 590-600 nm). SR-VAD-OPH reveals complete caspase activation in the form of red fluorescence in all cells. Data courtesy of Ms. Tracy Hanson, ICT (ICT111809; SR_OPH_111_17_18).
Figure 3. Jurkat cells were treated with DMSO (left) or staurosporine (right), then labeled with SR-VAD-OPH, followed by Green Live/Dead Stain (Cat. 6342). Four populations of cells were detected: unstained live cells (lower left); red early apoptotic cells stained with SR-VAD-OPH (lower right); late apoptotic cells (upper right) are dually stained with SR-VAD-OPH and Green Live/Dead Stain; and green necrotic cells (upper left). Data courtesy of Tracy Hanson, ICT (ICT-202:97; 091015).

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Product Manual

SKU: 6357

Size: 4 146 µg Vials
Price:
$572.00
Ships: 1-2 business days
  1. For in vitro apoptosis detection, culture your cells to a concentration of 2-5 x 105 cells/mL.
  2. Prepare experimental and control populations; concentrate cells to 5 x 105-106 cells/mL in 300 µL samples.
  3. Reconstitute the SR-OPH reagent with 50 µL DMSO to form the 150X stock concentrate (may be frozen for future use).
  4. Dilute the SR-OPH stock concentrate with 200 µL PBS to form the 30X working solution.
  5. Add SR-OPH working solution directly to samples and controls at a ratio of 1:30. For example, add 10 µL of working solution to a 300 µL aliquot of suspension cells.
  6. Incubate ~1 hour.
  7. Wash and spin cells two or three times.
  8. If desired, label cells with Hoechst 33342 nuclear stain, DAPI nuclear stain, or other compatible fluorescent markers.
  9. If desired, fix cells with formalin-based agents.
  10. Analyze data using a fluorescence microscope, plate reader, or flow cytometer.

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