# | Aladdin ID | Assay Type | Description | Organism | Compounds | Reference | BAO Format | Source | |
---|---|---|---|---|---|---|---|---|---|
1. | ALA3427823 | Binding | Inhibition of PIM2 (unknown origin) assessed as remaining activity at 100 uM by radioactive filter binding assay in presence of [33P]-ATP relative to control | Homo sapiens | 16 | ALA3425453 | single protein format | Scientific Literature | |
2. | ALA3430828 | Binding | Delta TM value showing the stabilisation of PIM2 produced by compound binding | Homo sapiens | 156 | ALA1145498 | single protein format | Scientific Literature | |
3. | ALA3606455 | Binding | Inhibition of human PIM2 | Homo sapiens | 1 | ALA3603755 | single protein format | Scientific Literature | |
4. | ALA3630921 | Binding | Inhibition of PIM2 (unknown origin) at 10 uM after 120 mins P33 radiolabeled kinase activity assay | Homo sapiens | 3 | ALA3627608 | single protein format | Scientific Literature | |
5. | ALA3631439 | Binding | Inhibition of PIM2 (unknown origin) assessed as remaining activity at 10 uM by high-throughput assay relative to control | Homo sapiens | 1 | ALA3627674 | single protein format | Scientific Literature | |
6. | ALA3636741 | Binding | Inhibition of Pim2 (unknown origin) using 5FAM-ARKRRRHPSGPPTA as substrate after 90 mins | Homo sapiens | 39 | ALA3632547 | single protein format | Scientific Literature | |
7. | ALA3705221 | Binding | ATP Depletion Assay: The activity of PIM2 is measured using a luciferase-luciferin based ATP detection reagent to quantify ATP depletion resulting from kinase-catalyzed phosphoryl transfer to a peptide substrate. Compounds to be tested are dissolved in 100% DMSO and directly distributed into white 384-well plates at 0.5 ul per well. To start the reaction, 10 ul of 10 nM Pim2 kinase and 20 uM BAD peptide (RSRHSSYPAGT-OH) in assay buffer (50 mM HEPES pH 7.5, 5 mM MgCl2, 1 mM DTT, 0.05% BSA) is added into each well. After 15 minutes, 10 ul of 8 uM ATP in assay buffer is added. Final assay concentrations are 5 nM PIM2, 4 uM ATP, 10 uM BAD peptide and 2.5% DMSO. The reaction is performed until approximately 50% of the ATP is depleted, then stopped with the addition of 20 ul KinaseGlo Plus (Promega Corporation) solution. The stopped reaction is incubated for 10 minutes and the remaining ATP detected via luminescence on the Victor2 (Perkin Elmer). | Homo sapiens | 19 | ALA3638844 | single protein format | BindingDB Database | |
8. | ALA3705115 | Binding | PIM-2 Enzyme Assay: The assay for the determination of PIM activity is based on the incorporation of [33P] ATP into PIM2tide substrate and capture of the radiolabeled peptide onto a Whatman P81 (phosphocellulose) filter plate. Assay mixtures contained 4 uM [gamma-33P] ATP (20 uCi/mL), 1.0 uM PIM2tide and 1.5 nM PIM-2 in place of PIM-1. | Homo sapiens | 114 | ALA3639321 | single protein format | BindingDB Database | |
9. | ALA3706072 | Binding | Depletion Assay: The activity of PIM2 is measured using a luciferase-luciferin based ATP detection reagent to quantify ATP depletion resulting from kinase-catalyzed phosphoryl transfer to a peptide substrate. Compounds to be tested are dissolved in 100% DMSO and directly distributed into white 384-well plates at 0.5 piper well. To start the reaction, 10 μl of 10 nM Pim2 kinase and 20 μM BAD peptide (RSRHSSYPAGT-OH) in assay buffer (50 mM HEPES pH 7.5, 5 mM MgCl2, and 1 mM DTT, 0.05% BSA) is added into each well. After 15 minutes, 10 μl of 8 μM ATP in assay buffer is added. Final assay concentrations are 5 nM PIM2, 4 μM ATP, 10 μM BAD peptide and 2.5% DMSO. The reaction is performed until approximately 50% of the ATP is depleted, then stopped with the addition of 20 μl KinaseGlo Plus (Promega Corporation) solution. The stopped reaction is incubated for 10 minutes and the remaining ATP detected via luminescence on the Victor2 (Perkin Elmer). | Homo sapiens | 54 | ALA3638819 | single protein format | BindingDB Database | |
10. | ALA3707966 | Binding | Kinase Assay: The inhibitory activity of putative kinase inhibitors and the potency of selected compounds were determined using a trans-phosphorylation assay.Specific peptide or protein substrates are trans-phosphorylated by their specific ser-thr or tyr kinase in the presence of ATP traced with 33P-gamma-ATP, and in the presence of their own optimal buffer and cofactors.At the end of the phosphorylation reaction, more than 98% unlabeled ATP and radioactive ATP is captured by an excess of the ion exchange dowex resin; the resin then settles down to the bottom of the reaction plate by gravity.Supernatant is subsequently withdrawn and transferred into a counting plate, then evaluated by (3-counting.Reagents/Assay ConditionsDowex Resin Preparation500 g of wet resin (SIGMA, custom prepared resin DOWEX 1x8 200-400 mesh, 2.5 Kg) are weighed out and diluted to 2 L in 150 mM sodium formate, pH 3.00.The resin is allowed to settle down (some hours) and then the supernatant is discarded. | Homo sapiens | 3 | ALA3638988 | single protein format | BindingDB Database | |
11. | ALA3707802 | Binding | AlphaScreen Assay: Pim 1, Pim 2 & Pim 3 AlphaScreen assays using high ATP (11-125.times. ATP Km) were used to determine the biochemical activity of the inhibitors. The activity of Pim 1, Pim 2, & Pim 3 is measured using a homogeneous bead based system quantifying the amount of phosphorylated peptide substrate resulting from kinase-catalyzed phosphoryl transfer to a peptide substrate. Compounds to be tested are dissolved in 100% DMSO and directly distributed to a white 384-well plate at 0.25 .mu.l per well. To start the reaction, 5 .mu.l of 100 nM Bad peptide (Biotin-AGAGRSRHSSYPAGT-OH (SEQ ID NO:1)) and ATP (concentrations described below) in assay buffer (50 mM Hepes, pH=7.5, 5 mM MgCl.sub.2, 0.05% BSA, 0.01% Tween-20, 1 mM DTT) is added to each well. This is followed by the addition of 5 .mu.l/well of Pim 1, Pim 2 or Pim 3 kinase in assay buffer (concentrations described below). Final assay concentrations (described below) are in 2.5% DMSO. The reactions are performed for .about.2 hours. | Homo sapiens | 201 | ALA3639088 | single protein format | BindingDB Database | |
12. | ALA3707781 | Binding | ATP Depletion Assay: The activity of PIM1, PIM2, and PIM3 is measured using a luciferase-luciferin based ATP detection reagent to quantify ATP depletion resulting from kinase-catalyzed phosphoryl transfer to a peptide substrate. | Homo sapiens | 137 | ALA3638421 | single protein format | BindingDB Database | |
13. | ALA3707982 | Binding | Binding Assay: PIM-1, -2, and -3 enzymes were generated as fusion proteins expressed in bacteria and purified by IMAC column chromatography (Sun, X., Chiu, J. F., and He, Q. Y. (2005) Expert Rev. Proteomics, 2:649-657). A fluorescent-labeled Pim-specific peptide substrate, was custom synthesized by American Peptide Company (Sunnyvale, Calif.). Reaction Buffer contained 10 mM HEPES, pH 7.2, 10 mM MgCl2, 0.01% Tween 20, 2 mM DTT. Termination Buffer contained 190 mM HEPES, pH 7.2, 0.015% Brij-35, 0.2% Coating Reagent 3 (Caliper Life Sciences, Hopkinton, Mass.), 20 mM EDTA. Separation Buffer contained 100 mM HEPES, pH 7.2, 0.015% Brij-35, 0.1% Coating Reagent 3, 1:200 Coating Reagent 8 (Caliper Life Sciences, Hopkinton, Mass.), 10 mM EDTA and 5% DMSO. | Homo sapiens | 29 | ALA3638875 | assay format | BindingDB Database | |
14. | ALA3707582 | Binding | Kinase Inhibition Assay: The buffer for PIM-2 assay was composed of HEPES 50 mM, at pH 7.5, with 1 mM MgCl2, 1 mM DTT, 3 microM Na3VO4, and 0.2 mg/mL BSAFull-length human PIM-2 was expressed and purified as described in Fedorov O, et al., PNAS 2007 104, 51, 20523-28.Assay Conditions (Final Concentrations)Enzyme concentration=1.5 nMAktide substrate (Chemical Abstract Service Registry Number 324029-01-8)=5 microMATP=4 microM33P-γ-ATP=1 nM. | Homo sapiens | 17 | ALA3639053 | single protein format | BindingDB Database | |
15. | ALA3707524 | Binding | Inhibition Assay: Inhibition assay using PIM-1, PIM-2 and PIM-3. | Homo sapiens | 1 | ALA3639232 | single protein format | BindingDB Database | |
16. | ALA3734496 | Binding | Inhibition of recombinant human PIM2 using fluorescein-labeled peptide as substrate by fluorescence-electrophoretic mobility shift assay | Homo sapiens | 1 | ALA3727354 | single protein format | Patent Bioactivity Data | |
17. | ALA3738379 | Binding | Inhibition of PIM2 (unknown origin) by electrophoretic mobility shift assay | Homo sapiens | 1 | ALA3734671 | single protein format | Scientific Literature | |
18. | ALA3739158 | Binding | Inhibition of PIM2 (unknown origin) at 200 nM by electrophoretic mobility shift assay | Homo sapiens | 2 | ALA3734671 | single protein format | Scientific Literature | |
19. | ALA3742689 | Binding | Inhibition of PIM2 (unknown origin) | Homo sapiens | 1 | ALA3739276 | single protein format | Scientific Literature | |
20. | ALA3750240 | Binding | Inhibition of PIM2 kinase (unknown origin) using NH2-AGAGRSRHSSYPAGT-OH as substrate by kinase-Glo assay | Homo sapiens | 14 | ALA3745712 | single protein format | Scientific Literature |
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