Journal: Biochemical Journal

Article Title: The VEGFR/PDGFR tyrosine kinase inhibitor, ABT-869, blocks necroptosis by targeting RIPK1 kinase

doi: 10.1042/bcj20230035

Figure Lengend Snippet: Figure 2. ABT-869 inhibits necroptosis in mouse and human cells. (A,B) Wild-type mouse dermal fibroblast (MDF) cells were treated with increasing concentrations of ABT-869 or control compounds, RIPK3

Article Snippet: Necroptosis-Inducing Stimuli Stimulus Component Supplier Concentration TSQ TNF Produced in-house [61] 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM Q-VD-OPh MP Biologicals 10 mM TSZ TNF Produced in-house 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM z-VAD-fmk ApexBio 10 mM TSI TNF Produced in-house 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM IDN-6556 (Emricasan) Tetralogic 5 mM Western Blot Primary Antibodies Protein Antibody Supplier Catalogue # Dilution Mouse MLKL Rat anti-MLKL clone 3H1 Produced in-house [20], available from Merck Millipore cat #MABC604 1 : 1000 Mouse RIPK1 Rabbit anti-RIPK1 Cell Signalling Technology cat #3493 1 : 1000 Mouse RIPK3 (CETSA) Rabbit anti-RIPK3 ProSci cat #2283 1 : 1000 Mouse RIPK3 (phosphorylation time course) Rat anti-mouse RIPK3 clone 8G7 Produced in-house [9], available from Merck Millipore cat #MABC1595 1 : 1000 Human MLKL Rat anti-MLKL clone 3H1 Produced in-house [20], available from Merck Millipore cat #MABC604 1 : 1000 Continued Figure 4.

Techniques: Control

Journal: Biochemical Journal

Article Title: The VEGFR/PDGFR tyrosine kinase inhibitor, ABT-869, blocks necroptosis by targeting RIPK1 kinase

doi: 10.1042/bcj20230035

Figure Lengend Snippet: Figure 3. ABT-869 binds to RIPK1 in mouse and human cells. Part 1 of 2 (A) Binding affinities (KD) of ABT-869 and WEHI-615 for human full-length MLKL, RIPK1 kinase domain and RIPK3 kinase domain measured by

Article Snippet: Necroptosis-Inducing Stimuli Stimulus Component Supplier Concentration TSQ TNF Produced in-house [61] 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM Q-VD-OPh MP Biologicals 10 mM TSZ TNF Produced in-house 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM z-VAD-fmk ApexBio 10 mM TSI TNF Produced in-house 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM IDN-6556 (Emricasan) Tetralogic 5 mM Western Blot Primary Antibodies Protein Antibody Supplier Catalogue # Dilution Mouse MLKL Rat anti-MLKL clone 3H1 Produced in-house [20], available from Merck Millipore cat #MABC604 1 : 1000 Mouse RIPK1 Rabbit anti-RIPK1 Cell Signalling Technology cat #3493 1 : 1000 Mouse RIPK3 (CETSA) Rabbit anti-RIPK3 ProSci cat #2283 1 : 1000 Mouse RIPK3 (phosphorylation time course) Rat anti-mouse RIPK3 clone 8G7 Produced in-house [9], available from Merck Millipore cat #MABC1595 1 : 1000 Human MLKL Rat anti-MLKL clone 3H1 Produced in-house [20], available from Merck Millipore cat #MABC604 1 : 1000 Continued Figure 4.

Techniques: Binding Assay

Journal: Biochemical Journal

Article Title: The VEGFR/PDGFR tyrosine kinase inhibitor, ABT-869, blocks necroptosis by targeting RIPK1 kinase

doi: 10.1042/bcj20230035

Figure Lengend Snippet: Figure 4. ABT-869 binds to mouse and human RIPK1 in vitro. Part 1 of 2 Thermal Shift Assays (TSA) with mouse and human RIPK1 and RIPK3 kinase domains. Increasing concentrations of ABT-869

Article Snippet: Necroptosis-Inducing Stimuli Stimulus Component Supplier Concentration TSQ TNF Produced in-house [61] 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM Q-VD-OPh MP Biologicals 10 mM TSZ TNF Produced in-house 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM z-VAD-fmk ApexBio 10 mM TSI TNF Produced in-house 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM IDN-6556 (Emricasan) Tetralogic 5 mM Western Blot Primary Antibodies Protein Antibody Supplier Catalogue # Dilution Mouse MLKL Rat anti-MLKL clone 3H1 Produced in-house [20], available from Merck Millipore cat #MABC604 1 : 1000 Mouse RIPK1 Rabbit anti-RIPK1 Cell Signalling Technology cat #3493 1 : 1000 Mouse RIPK3 (CETSA) Rabbit anti-RIPK3 ProSci cat #2283 1 : 1000 Mouse RIPK3 (phosphorylation time course) Rat anti-mouse RIPK3 clone 8G7 Produced in-house [9], available from Merck Millipore cat #MABC1595 1 : 1000 Human MLKL Rat anti-MLKL clone 3H1 Produced in-house [20], available from Merck Millipore cat #MABC604 1 : 1000 Continued Figure 4.

Techniques: In Vitro

Journal: Biochemical Journal

Article Title: The VEGFR/PDGFR tyrosine kinase inhibitor, ABT-869, blocks necroptosis by targeting RIPK1 kinase

doi: 10.1042/bcj20230035

Figure Lengend Snippet: Figure 5. ABT-869 inhibits RIPK1 kinase activity in vitro and in cells. Part 1 of 2 (A–H) In vitro phosphorylation assays with mouse and human RIPK1 and RIPK3 kinase domains measured by ADP-Glo Kinase

Article Snippet: Necroptosis-Inducing Stimuli Stimulus Component Supplier Concentration TSQ TNF Produced in-house [61] 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM Q-VD-OPh MP Biologicals 10 mM TSZ TNF Produced in-house 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM z-VAD-fmk ApexBio 10 mM TSI TNF Produced in-house 100 ng/ml Smac mimetic (Compound A) Tetralogic 500 nM IDN-6556 (Emricasan) Tetralogic 5 mM Western Blot Primary Antibodies Protein Antibody Supplier Catalogue # Dilution Mouse MLKL Rat anti-MLKL clone 3H1 Produced in-house [20], available from Merck Millipore cat #MABC604 1 : 1000 Mouse RIPK1 Rabbit anti-RIPK1 Cell Signalling Technology cat #3493 1 : 1000 Mouse RIPK3 (CETSA) Rabbit anti-RIPK3 ProSci cat #2283 1 : 1000 Mouse RIPK3 (phosphorylation time course) Rat anti-mouse RIPK3 clone 8G7 Produced in-house [9], available from Merck Millipore cat #MABC1595 1 : 1000 Human MLKL Rat anti-MLKL clone 3H1 Produced in-house [20], available from Merck Millipore cat #MABC604 1 : 1000 Continued Figure 4.

Techniques: Activity Assay, In Vitro, Phospho-proteomics