178
3. PBS buffer (10 mM, pH 7.0): Both interactants should be dissolved in the same buffer.
4. Deuterated dimethyl sulfoxide 99.8% (DMSO-d 6 ) and deuterium oxide 99.9% (D 2 O).
5. Bruker 400 MHz Avance spectrometer equipped with a
z- gradient unit (Bruker BioSpin, Rheinstetten, Germany): The
NMR system is controlled by the software TopSpin 3.1.
1. PBS buffer (10 mM, pH 7.0), water (double distilled),
methanol.
2. 1 mL Cuvettes.
3. Water bath.
4. 37 °C Shaker and incubator.
5. UV-Vis spectrometer (slit = 1, speed 240 nm/min).
1. Theophylline (internal standard).
2. Regenerated cellulose membrane syringe filters 0.2 μm.
3. Cellulose nitrate filters, 0.2 μm pore size, for mobile-phase
filtering.
4. Acetonitrile, water, and formic acid (LC-MS grade).
5. Mobile phase A: Water containing 0.1% (v/v) formic acid.
6. Mobile phase B: Acetonitrile containing 0.1% (v/v) formic acid.
7. C18 column 100 mm × 2.1 mm, 2.6 um, with ProGuard column 2.1 mm.
8. Triple-quadrupole mass spectrometer coupled to ultrahighperformance liquid chromatography (Bruker’s EVOQ Elite
ER-UHPLC).
1. Patient-derived primary GBM cell cultures (GBM31, GBM59,
GBM77) were established from fresh tumor tissue obtained
from first surgical debulking or stereotactic biopsies at Charing
Cross Hospital as described previously.
2. Tissue samples were provided by the Imperial College
Healthcare NHS Trust Tissue Bank, which is supported by the
National Institute for Health Research (NIHR) Biomedical
Research Centre based at Imperial College Healthcare NHS
Trust and Imperial College London.
2.2 UV-Vis
Spectroscopy
2.3 Liquid
Chromatography
and Mass
Spectrometry
2.4 In Vitro Assays
of TMZ@PSC4 Activity
Antonis D. Tsiailanis et al.
3. PBS buffer (10 mM, pH 7.0): Both interactants should be dissolved in the same buffer.
4. Deuterated dimethyl sulfoxide 99.8% (DMSO-d 6 ) and deuterium oxide 99.9% (D 2 O).
5. Bruker 400 MHz Avance spectrometer equipped with a
z- gradient unit (Bruker BioSpin, Rheinstetten, Germany): The
NMR system is controlled by the software TopSpin 3.1.
1. PBS buffer (10 mM, pH 7.0), water (double distilled),
methanol.
2. 1 mL Cuvettes.
3. Water bath.
4. 37 °C Shaker and incubator.
5. UV-Vis spectrometer (slit = 1, speed 240 nm/min).
1. Theophylline (internal standard).
2. Regenerated cellulose membrane syringe filters 0.2 μm.
3. Cellulose nitrate filters, 0.2 μm pore size, for mobile-phase
filtering.
4. Acetonitrile, water, and formic acid (LC-MS grade).
5. Mobile phase A: Water containing 0.1% (v/v) formic acid.
6. Mobile phase B: Acetonitrile containing 0.1% (v/v) formic acid.
7. C18 column 100 mm × 2.1 mm, 2.6 um, with ProGuard column 2.1 mm.
8. Triple-quadrupole mass spectrometer coupled to ultrahighperformance liquid chromatography (Bruker’s EVOQ Elite
ER-UHPLC).
1. Patient-derived primary GBM cell cultures (GBM31, GBM59,
GBM77) were established from fresh tumor tissue obtained
from first surgical debulking or stereotactic biopsies at Charing
Cross Hospital as described previously.
2. Tissue samples were provided by the Imperial College
Healthcare NHS Trust Tissue Bank, which is supported by the
National Institute for Health Research (NIHR) Biomedical
Research Centre based at Imperial College Healthcare NHS
Trust and Imperial College London.
2.2 UV-Vis
Spectroscopy
2.3 Liquid
Chromatography
and Mass
Spectrometry
2.4 In Vitro Assays
of TMZ@PSC4 Activity
Antonis D. Tsiailanis et al.