135
10. Sterile 1× phosphate-buffered saline (PBS) solution.
11. Transfection reagents.
12. Filter paper discs and forceps for isolation of stable clones.
1. Black-walled or white-walled 96-well clear-bottom plates for
tissue culture.
2. Sterile low adherence pipette tips with barrier filter.
3. Firefly luciferase substrate: 4 mg/mL d-luciferin 40× stock
solution in PBS stored in dark colored vials at −80 °C.
Alternatively, GloSensor c-AMP reagent (Promega) can be
used to monitor firefly luciferase activity.
4. ATM inhibitor: 1000× stock solution 3 M caffeine.
5. ATM inhibitor: 1000× stock solution 20 mM KU-60019.
6. ATM inhibitor: 1000× stock solution 20 mM KU-55933.
7. ATM inhibitor: 1000× stock solution 20 mM CGK733.
8. Live cell bioluminescence imaging system with very high
sensitivity and required software package for data generation
and analysis (IVIS, Envision Xcite multi-label Plate Readers
from Perkin Elmer; or similar system).
9. Liquid handling instrument (Biomek Nx from Beckman or
similar).
10. Plate handling robot (Plate Handler II robot from Perkin
Elmer or similar).
11. Optional: Cell culture incubator compatible with highthroughput instruments (LiCONiC StoreX STX44 IC precision incubator from Liconic Instruments or similar) for
high-throughput assays.
1. Appropriate mouse strain for desired experimental system such
as immunocompromised mice (nude, SCID, or NSG) for
human tumor xenografts.
2. Optional: Small animal shaver such as Wahl trimmer.
3. 40 mg/mL luciferin stock in PBS, store in tightly sealed dark
tubes at −20 or −80 °C.
4. 28–30 gauge insulin syringe for intra-peritoneal (IP) luciferin
injection in mice.
5. Bioluminescence imaging instrument (IVIS or similar instrument)
with a heated platform and isoflurane anesthesia injection and
controller systems.
6. Additional accessories such as nose cones, animal partitions,
black paper sheets, ear tags, markers, 70% alcohol, and 10%
bleach or similar solution for disinfecting bench surfaces.
2.3 Cell Imaging
2.4 Animal Imaging
Molecular Imaging of ATM Kinase Activity
10. Sterile 1× phosphate-buffered saline (PBS) solution.
11. Transfection reagents.
12. Filter paper discs and forceps for isolation of stable clones.
1. Black-walled or white-walled 96-well clear-bottom plates for
tissue culture.
2. Sterile low adherence pipette tips with barrier filter.
3. Firefly luciferase substrate: 4 mg/mL d-luciferin 40× stock
solution in PBS stored in dark colored vials at −80 °C.
Alternatively, GloSensor c-AMP reagent (Promega) can be
used to monitor firefly luciferase activity.
4. ATM inhibitor: 1000× stock solution 3 M caffeine.
5. ATM inhibitor: 1000× stock solution 20 mM KU-60019.
6. ATM inhibitor: 1000× stock solution 20 mM KU-55933.
7. ATM inhibitor: 1000× stock solution 20 mM CGK733.
8. Live cell bioluminescence imaging system with very high
sensitivity and required software package for data generation
and analysis (IVIS, Envision Xcite multi-label Plate Readers
from Perkin Elmer; or similar system).
9. Liquid handling instrument (Biomek Nx from Beckman or
similar).
10. Plate handling robot (Plate Handler II robot from Perkin
Elmer or similar).
11. Optional: Cell culture incubator compatible with highthroughput instruments (LiCONiC StoreX STX44 IC precision incubator from Liconic Instruments or similar) for
high-throughput assays.
1. Appropriate mouse strain for desired experimental system such
as immunocompromised mice (nude, SCID, or NSG) for
human tumor xenografts.
2. Optional: Small animal shaver such as Wahl trimmer.
3. 40 mg/mL luciferin stock in PBS, store in tightly sealed dark
tubes at −20 or −80 °C.
4. 28–30 gauge insulin syringe for intra-peritoneal (IP) luciferin
injection in mice.
5. Bioluminescence imaging instrument (IVIS or similar instrument)
with a heated platform and isoflurane anesthesia injection and
controller systems.
6. Additional accessories such as nose cones, animal partitions,
black paper sheets, ear tags, markers, 70% alcohol, and 10%
bleach or similar solution for disinfecting bench surfaces.
2.3 Cell Imaging
2.4 Animal Imaging
Molecular Imaging of ATM Kinase Activity