Scientific Citations

More than ### citations published since the beginning of 2017.

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Atypical KRASG12R Mutant Is Impaired in PI3K Signaling and Macropinocytosis in Pancreatic Cancer

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Structural complementarity facilitates E7820-mediated degradation of RBM39 by DCAF15

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Defining the landscape of ATP-competitive inhibitor resistance residues in protein kinases

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Inducing and exploiting vulnerabilities for the treatment of liver cancer

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An organoid platform for ovarian cancer captures intra- and interpatient heterogeneity

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Small-molecule targeting of brachyury transcription factor addiction in chordoma

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TAS-120 Overcomes Resistance to ATP-Competitive FGFR Inhibitors in Patients with FGFR2 Fusion-Positive Intrahepatic Cholangiocarcinoma

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Oral Mucosal Organoids as a Potential Platform for Personalized Cancer Therapy

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Unique and Shared Epigenetic Programs of the CREBBP and EP300 Acetyltransferases in Germinal Center B Cells Reveal Targetable Dependencies in Lymphoma

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Analysis of Liver Cancer Cell Lines Identifies Agents With Likely Efficacy Against Hepatocellular Carcinoma and Markers of Response

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Scientific citations published in peer reviewed high impact journals

Atypical KRASG12R Mutant Is Impaired in PI3K Signaling and Macropinocytosis in Pancreatic Cancer

Two days after seeding, organoids were 605 treated with ERKi SCH772984 (0 nM to 10 μM) or MEKi AZD6244 (selumetinib) (0 to 2.5 μM), randomized on a Tecan D300e drug dispenser. O

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Structural complementarity facilitates E7820-mediated degradation of RBM39 by DCAF15

Titrations of BodipyFL-E7820 (4) were carried out by mixing 200 nM biotinylated Strep-II-Avi-tagged DCAF15 variants or an equivalent volume of the assay buffer, 2 nM terbium-coupled streptavidin in the same assay buffer. After dispensing the assay mixture, an increasing concentration of BodipyFL-E7820 (4) was dispensed in the 384-well plate using a Titrations of BodipyFL-E7820 (4) were carried out by mixing 200 nM biotinylated Strep-II-Avi-tagged DCAF15 variants or an equivalent volume of the assay buffer, 2 nM terbium-coupled streptavidin in the same assay buffer. After dispensing the assay mixture, an increasing concentration of BodipyFL-E7820 (4) was dispensed in the 384-well plate using a Titrations of BodipyFL-E7820 (4) were carried out by mixing 200 nM biotinylated Strep-II-Avi-tagged DCAF15 variants or an equivalent volume of the assay buffer, 2 nM terbium-coupled streptavidin in the same assay buffer. After dispensing the assay mixture, an increasing concentration of BodipyFL-E7820 (4) was dispensed in the 384-well plate using a D300e digital dispenser normalized to 2% DMSO and then incubated for 15 min at room temperature. normalized to 2% DMSO and then incubated for 15 min at room temperature. normalized to 2% DMSO and then incubated for 15 min at room temperature

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Defining the landscape of ATP-competitive inhibitor resistance residues in protein kinases

Doxycycline (from a 0.3% Tween stock solution) was added to the remaining cells where applicable and drugs (from DMSO solutions) were added using an HP D300 Digital Dispenser

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Inducing and exploiting vulnerabilities for the treatment of liver cancer

About 24 h later, drugs were added at the indicated concentrations using the HP D300 Digital Dispenser (HP)

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An organoid platform for ovarian cancer captures intra- and interpatient heterogeneity

The drugs and their combinations were added 1 h after plating the organoids using the Tecan D300e Digital Dispenser. Drugs were dispensed in a randomized manner and DMSO end concentration was 1% in all wells. 120 h after adding the drugs, ATP levels were measured using the Cell-Titer Glo2.0 (Promega BV) according to the manufacturer’s instructions and luminescence was measured using a SpectraMax microplate reader (Molecular Devices). Results were normalized to vehicle (DMSO = 100%) and baseline control (navitoclax 20 µM)

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Small-molecule targeting of brachyury transcription factor addiction in chordoma

384-well BioCoat Collagen I (Corning) microtiter plates at a density of 1200, 1800, 2000, 1600, and 1200 cells per well, respectively. The following day, compound or DMSO was added to wells using an HP D300 digital dispenser instrument. Each compound was tested using 9 concentrations, in quadruplicate (four wells treated in parallel). Cell viability was assayed 6 d after compound addition with th;384-well BioCoat Collagen I (Corning) microtiter plates at a density of 1200, 1800, 2000, 1600, and 1200 cells per well, respectively. The following day, compound or DMSO was added to wells using an HP D300 digital dispenser instrument. Each compound was tested using 9 concentrations, in quadruplicate (four wells treated in parallel). Cell viability was assayed 6 d after compound addition with th

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TAS-120 Overcomes Resistance to ATP-Competitive FGFR Inhibitors in Patients with FGFR2 Fusion-Positive Intrahepatic Cholangiocarcinoma

For IC50 measurement using the FGFR inhibitors, cells were dissociated into single cells and seeded into a 384-well tissue culture plate, each well with 200 viable cells and 40 μL of growth medium. After 24 hours, compounds were added to each well over a 15-point concentration range, along with DMSO controls, using a Tecan D300e digital drug dispenser. Cells were cultured for 5 days in the presence of compound before assessing viability by adding 15 μL of CellTiterGlo (Promega) to each well, incubating for 20 minutes at room temperature on a shaker, and measuring luminescence using an Envision

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Oral Mucosal Organoids as a Potential Platform for Personalized Cancer Therapy

The drugs were added 1 hour after plating the organoids using the Tecan D300e Digital Dispenser (Tecan). Nutlin-3 (Cayman Chemical, catalog no. 10004372), Niraparib (Selleckchem, catalog no. S2741), AZD4547 (ApeXbio, catalog no. A8250), everolimus (LC Laboratories, catalog no. E4040), vemurafenib (Selleckchem, catalog no. S1267), and alpelisib (LC Laboratories, catalog no. A4477) were dissolved in DMSO. Cisplatin (Sigma, catalog no. C2210000), carboplatin (Sigma, catalog no. C2538), and cetuximab (obtained from hospital pharmacy) were dissolved in PBS containing 0.3% Tween-20, which was required to dispense these drugs using the HP printer. All wells were normalized for solvent used. DMSO percentage never exceeded 1%, PBS/Tween-20 percentage never exceeded 2%. Drug exposure was performed in triplicate for each concentration shown. For a layout of the drug screen, see Supplementary Fig. S12

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Unique and Shared Epigenetic Programs of the CREBBP and EP300 Acetyltransferases in Germinal Center B Cells Reveal Targetable Dependencies in Lymphoma

After 24 h, the CCS1477 and HATi329 compounds were added in 4 replicates using a HP D300 digital dispenser (HP), with 12 wells of DMSO-treated control cells. CellTiter-Glo readings were acquired after 48 h using a Tecan Infinite 200 plate reader. T

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Analysis of Liver Cancer Cell Lines Identifies Agents With Likely Efficacy Against Hepatocellular Carcinoma and Markers of Response

We analyzed 31 therapeutic compounds on the whole panel of 34 LCCLs (Supplementary Table 4). Drug screening was performed using the HP D300 digital dispenser (Tecan, Mannedorf, Switzerland) to create dose-response curves as described in the Supplementary

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Patient-derived organoids can predict response to chemotherapy in metastatic colorectal cancer patients

and oxaliplatin (oxaliplatin Cmax in patients = 3.8 to 10.1 mM; oxaliplatin range in vitro = 0.319 to 200 mM) in patients using a Tecan D300e digital dispenser (31–33)

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Suz12 inactivation cooperates with JAK3 mutant signaling in the development of T-cell acute lymphoblastic leukemia

HTS dispenser (TTP Labtech). For single-drug dose response studies, cells were seeded into 96-well plates at 300 000 cells/mL, and the compounds were added using a D300e digital dispenser (Tecan). Cell proliferation was

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Small-molecule allosteric inhibitors of BAX

BAX is a critical effector of the mitochondrial cell death pathway in response to a diverse range of stimuli in physiological and disease contexts. Upon binding by BH3-only proteins, cytosolic BAX undergoes conformational activation and translocation, resulting in mitochondrial outer ;ed with indicated concentrations of BTSA1 and BAI1 or DMSO in no FBS media for 2.5 hr, followed by 10% FBS replacement to a final volume of 25 μl. Dilutions of compounds were performed using a TECAN D300e Digital Dispenser from 10 mM stocks. Cell viability was assayed at 6 hr by addition of CellTiter-Glo according to the manufacturer’s protocol (Promega), and luminescence measured using a F200

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Crystal structure of misoprostol bound to the labor inducer prostaglandin E2 receptor

or 1h. Then, e-Coelenterazine Prolume Purple (Methoxy e-CTZ; Nanolight) was added to a final concentration of 2 μM followed immediately by the addition of increasing agonist concentrations using the HP D300 digital dispenser (Tecan). After 10 min of incubation at room temperature, the BRET signal was subsequently detected at 0.2 sec integration time using a Synergy NEO plate reader (BioTek) equip

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Plasticity of the Mycobacterium tuberculosis respiratory chain and its impact on tuberculosis drug development

Compounds were solubilized in DMSO and dispensed into 384-well plates using a D300e Digital Dispenser (HP)

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Exploring use of unsupervised clustering to associate signaling profiles of GPCR ligands to clinical response

cin (100 µg mL−1) and puromycin (10 mg mL−1). BRET assays Ligand preparation: Agonists were dissolved in DMSO and spotted on 96-well white bottom microplates (Greiner bio-one) using the HP D300 Digital Dispenser (Tecan Life Sciences). DMSO concentration was normalized for each point at 0.334%. Gαi and Gαo-activation assay: HEK 293 were co-transfected with DOR or MOR (human or rat)

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Small molecule degraders of the hepatitis C virus protease reduce susceptibility to resistance mutations

384-well plates with 50 μL FluoroBrite DMEM media (Thermo Fisher Scientific A18967) containing 10% FBS per well a day before compound treatment. Compounds and 100 nM dBET6 were dispensed using a D300e Digital Dispenser (HP), normalized to 0.5% DMSO, and incubated with cells for 5 h. The assay plate was imaged immediately using an Acumen High Content Imager (TTP Labtech) with 488 nm and 56;384-well plates with 50 μL FluoroBrite DMEM media (Thermo Fisher Scientific A18967) containing 10% FBS per well a day before compound treatment. Compounds and 100 nM dBET6 were dispensed using a D300e Digital Dispenser (HP), normalized to 0.5% DMSO, and incubated with cells for 5 h. The assay plate was imaged immediately using an Acumen High Content Imager (TTP Labtech) with 488 nm and 56

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Neuronal differentiation and cell-cycle programs mediate response to BET-bromodomain inhibition in MYC-driven medulloblastoma

rimental details: Cell lines were seeded into 384-well, white-walled, clear bottom plates at a density of 500 cells/well. Twenty-four hours after seeding, combination drugs were administered using an HP D300 Digital Dispenser in matrix format. Drug was administered such that the final volume of DMSO did not exceed 0.5%. The cells were then incubated for seven days and cell luminescence was measure

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Dissecting heterogeneity in malignant pleural mesothelioma through histo-molecular gradients for clinical applications

lls were seeded in triplicate on 96-well plates (Corning, Falcon) at 0.5–1 × 104 cells per well and treated with gradient concentrations of compounds diluted in DMSO (Dimethyl sulfoxide) using HP D300 Digital Dispenser (Tecan). Three compounds were used KIN001-102/Akt Inhibitor VIII (124018, Merck), 681640/Wee1 Inhibitor (681640, Merck) and a ROCK1 inhibitor, GSK269962A (S7687, Selleckchem)

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An oligomeric state-dependent switch in the ER enzyme FICD regulates AMPylation and deAMPylation of BiP

of HKTx dispensed into 384‐well non‐binding, low volume, HiBase, black microplates (greiner bio‐one). Note, all FICD variants were dispensed directly into the microplate well solutions using a D300e digital dispenser (Tecan). The plate was then sealed and the final 20 μl (Fig 6F) or 15 μl reactions (Fig 6G) were incubated for 45 min at 10°C, whilst shaking at 300 rpm on a ThermoMix

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Long-term expanding human airway organoids for disease modeling

04 pGEM‐T Easy vector syst. I Promega A3600 Nanodrop Lite Thermo Fisher TruSeq Nano kit Illumina 20015965 Illumina HiSeqX Illumina DNeasy blood & tissue kit Qiagen 69506 D300e Digital Dispenser Tecan Micromanipulator Narishige M‐152 Microinjector Narishige IM‐5B Stereomicroscope Leica MZ75 FlexMAP3D Bio‐Rad Laboratories Methods and Prot;ted by pipetting before being resuspended in 5% BME/AO media (15–20,000 organoids ml−1) and dispensed into 384‐well microplates (Greiner). Each compound was dispensed at 1–100 μM using a TECAN D300e Digital Dispenser, and after 5 days, cell viability was assayed as described above. Paclitaxel (T7402), methotrexate (A6770), crizotinib (PZ0240), and cisplatin (C2210000) were obtained;04 pGEM‐T Easy vector syst. I Promega A3600 Nanodrop Lite Thermo Fisher TruSeq Nano kit Illumina 20015965 Illumina HiSeqX Illumina DNeasy blood & tissue kit Qiagen 69506 D300e Digital Dispenser Tecan Micromanipulator Narishige M‐152 Microinjector Narishige IM‐5B Stereomicroscope Leica MZ75 FlexMAP3D Bio‐Rad Laboratories Methods and Prot

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The ETS Inhibitors YK-4-279 and TK-216 Are Novel Antilymphoma Agents

For TK-216, cell lines were seeded in 384-well plates at the density of 2,000 cells/well using the VIAFLO 96/384 hand-held electronic channel pipette (Integra Biosciences AG) and then treated with D300e Digital Dispenser (Tecan)

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Capmatinib (INC280) Is Active Against Models of Non-Small Cell Lung Cancer and Other Cancer Types with Defined Mechanisms of MET Activation

Capmatinib was then added from a 10 mM dimethyl sulfoxide (DMSO) stock solution using a HP D300 Digital Dispenser (Tecan)

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KRAS G12C NSCLC Models Are Sensitive to Direct Targeting of KRAS in Combination with PI3K Inhibition

CellTiterGlo (Promega) reagent was added 96 hours after beginning treatment, and luminescence was measured on a Spectramax M5 spectrophotometer (Molecular Devices). All conditions were tested as triplicate biological replicates. All drug treatments were administered using a Tecan D300e Digital Dispenser. GI50 were calculated using GraphPad Prism Software using the function log (inhibitor) versus response-variable slope (4 parameters). Values are capped at the maximum drug dose used

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USP7 Cooperates with NOTCH1 to Drive the Oncogenic Transcriptional Program in T-Cell Leukemia

3,000 cells per well were seeded using a microplate Dispenser (MultiFlo™, BioTek) in 384-well clear bottom, black wall plates (Corning). Drugs were added using the Tecan D300e digital dispenser (Tecan). After 72-h incubation, alamarBlue™ cell viability reagent (ThermoFisher) was added and viability was quantified by measuring fluorescence in a plate reader (Tecan Infinite m1000 pro, Ex: 530 nm; Eλ: 590). Synergy analysis was conducted using Compusyn software69

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A Multi-center Study on the Reproducibility of Drug-Response Assays in Mammalian Cell Lines

Skip to article ;Using an automated liquid-handling robot such as the HP D300e Digital Dispenser, it is possible to dispense compounds directly into microtiter plates in an arbitrary pattern, randomizing the locations of control and technical replicates and converting systematic error into ;Using an automated liquid-handling robot such as the HP D300e Digital Dispenser, it is possible to dispense compounds directly into microtiter plates in an arbitrary pattern, randomizing the locations of control and technical replicates and converting systematic error into

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Adaptation of Human iPSC-Derived Cardiomyocytes to Tyrosine Kinase Inhibitors Reduces Acute Cardiotoxicity via Metabolic Reprogramming

Master drug stocks were stored at 10 mM in DMSO for TKIs and 2 M in water for 2DG with 0.03% Tween 20. Tween was added to facilitate dispensing of drugs in aqueous solvent by the HP D300 dispenser

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Aurora A kinase inhibition destabilizes PAX3-FOXO1 and MYCN and synergizes with Navitoclax to induce Rhabdomyosarcoma cell death

Sigma- Aldrich). Drugs (Supplemental Table 1) were purchased from Selleckchem and added to the cells using the HP D300 Digital Dispenser (Tecan). 24h after seeding cells, medium was changed to 19 μl culture medium

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Dual Inhibition of GLUT1 and the ATR/CHK1 Kinase Axis Displays Synergistic Cytotoxicity in KRAS-Mutant Cancer Cells

ell) in 100 mL complete medium and treated using a TECAN D300e digital dispenser (HP). After 72 hours of treatment, relative cell viability was assessed by measuring the ATP content in each well. Therefore, CellTiter-Glo Reagent (Promega, catalog no. G7573) was added 1:1. Upon 10 minutes of incubation, luminescence intensity was measured with a plate reader (Tecan Infinite M1000 Pro) and normalized to intensities of control wells

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Intra-tumour diversification in colorectal cancer at the single-cell level

All drugs were dispensed by a HP-D300 automated liquid dispenser (TECAN). Samples were incubated for 6 days at 37 °C, and cell viability was measured by CellTiter-Glo 3D kit (Promega) on a SpectraMax M5e (Molecular Devices). Cell viability measurements were performed in duplicate wells for each clone. Survival ratios in drug-treated organoids were normalized to the average survival in a DMSO control

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A Living Biobank of Breast Cancer Organoids Captures Disease Heterogeneity

Then, 21 concentrations of afatinib, gefitinib, pictilisib, GDC-0068, AZD8055, everolimus (all Selleckchem), or tamoxifen (Sigma) as well as DMSO controls were added in duplicate using a Tecan D300e digital dispenser (Life Sciences). To measure ATP as proxy for viable cells, 40 μL of CellTiter-Glo 3D Reagent (Promega) per well were added after five days, before shaking the plate for 30 min at room temperature and reading luminescence on a SpectraMax microplate reader (Molecular Devices)

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SHP2 is required for growth of KRAS-mutant non-small-cell lung cancer in vivo

Indicated cells were cultured and seeded into 96-well plates at a density of 500–2,000 cells per well, depending on growth rate. Twenty-four hours later, drugs were added at the indicated concentrations using the HP D300 Digital Dispenser (HP). Cells were imaged every 4 h in IncuCyte ZOOM (Essen Bioscience). Phase-contrast images were analyzed to detect cell proliferation based on cell confluence

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Cooperative targeting of melanoma heterogeneity with an AXL antibody-drug conjugate and BRAF/MEK inhibitors

For dose-response curves and viability assays in melanoma, 1 × 103 cells were plated in 96-well plates and ADC was added 3 h after seeding with the HP D300 Digital Dispenser (Tecan)

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Cooperative Enhancer Activation by TLX1 and STAT5 Drives Development of NUP214-ABL1/TLX1-Positive T Cell Acute Lymphoblastic Leukemia

synergy between the different compounds, ALL-SIL, mouse leukemic cells or patient-derived xenograft cells were seeded into 96-well plates and the compounds were added in a randomized fashion using a D300e digital dispenser (Tecan). The DMSO concentration was normalized. Cell proliferation was measured after 48 hours using the ATPlite luminescence system (PerkinElmer) using a Victor multilabel pla;synergy between the different compounds, ALL-SIL, mouse leukemic cells or patient-derived xenograft cells were seeded into 96-well plates and the compounds were added in a randomized fashion using a D300e digital dispenser (Tecan). The DMSO concentration was normalized. Cell proliferation was measured after 48 hours using the ATPlite luminescence system (PerkinElmer) using a Victor multilabel pla

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MAP3K1 and MAP2K4 mutations are associated with sensitivity to MEK inhibitors in multiple cancer models

in PBS and stained with 0.1% crystal violet in water. Incucyte proliferation assays were carried out in 96-well plates at a density of 500–1000 cells per well. 24 h later, drugs were added using HP D300 Digital Dispenser (HP) at indicated concentrations. Cells were imaged every 4 h in IncuCyte ZOOM (Essen Bioscience). Phase-contrast images were collected and analyzed to detect cell prolifer

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Additivity of inhibitory effects in multidrug combinations

In each of these experiments, an inoculum of 104 cells was inoculated into 150 μl of media in a Nunc 96-well flat-bottom microplate. Antibiotics were added into the wells as indicated using a D300e digital dispenser (Tecan), which dispenses a nanolitre-scale volume of each antibiotic. Each concentration combination was performed in duplicate wells. Multiple untreated control wells (no antibiotics) were designated on each plate (2–6% of the wells in each experiment). To avoid systematic positional effects across the plates, the wells chosen for each drug combination on the plates were randomized. The plates were incubated at 30 °C with shaking (Liconic orbital shaker STX44), and OD600 nm was measured at least every 25 min using a Tecan robotic system and the Infinite M200 Pro reader. To enhance uniformity, the plate orientations in the shaker were rotated 180° following every measurement

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Plasticity in binding confers selectivity in ligand-induced protein degradation

e IKZF1 homology model was taken from17. Time-resolved fluorescence resonance energy transfer (TR-FRET) Compounds in dimerization assays were dispensed in a 384-well microplate (Corning, 4514) using D300e Digital Dispenser (HP) normalized to 2% DMSO into 200 nM biotinylated His6-avi-bromodomain (WT or mutant, see Figure legends) or 80 nM biotinylated StrepII-avi-IKZF1Δ (See Figure legends), 100;e IKZF1 homology model was taken from17. Time-resolved fluorescence resonance energy transfer (TR-FRET) Compounds in dimerization assays were dispensed in a 384-well microplate (Corning, 4514) using D300e Digital Dispenser (HP) normalized to 2% DMSO into 200 nM biotinylated His6-avi-bromodomain (WT or mutant, see Figure legends) or 80 nM biotinylated StrepII-avi-IKZF1Δ (See Figure legends), 100

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Accessing chemical diversity from the uncultivated symbionts of small marine animals

cytometry-based bioassays CEMTART tat/rev++ cells were maintained as described for cytoprotection assays. Drugs suspended in DMSO were added to media (100 µl) in 96-well flat-bottom plates using an HP D300 Digital Dispenser. All wells were normalized with DMSO to 0.5% total volume. Divamide stock concentrations of either 5 or 2 µg/µl were utilized for assays employing a concentration range sui

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TBX2 is a neuroblastoma core regulatory circuitry component enhancing MYCN/FOXM1 reactivation of DREAM targets

n 384-well plates and incubated overnight. Next, cell lines and organoids were co-treated with three-fold dilution series of JQ1 (5.1 nM–33.3 μM) and THZ1 (0.051 nM–0.333 μM), using the HP D300 Digital Dispenser (Tecan). Control cells were treated with DMSO. Cell viability was determined after 72 h (classical cell lines) or 120 h (organoids) using the 3-(4.5-dimethylthiazol-2-yl)

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Plasmepsin II-III copy number accounts for bimodal piperaquine resistance among Cambodian Plasmodium falciparum

Drug susceptibility was measured using the SYBR Green I method as previously described44 [/articles/s41467-018-04104-z#ref-CR44]. In brief, tightly synchronized 0–6 h rings were grown for 72 h in the presence of different concentrations of drugs in 384-clear-bottom well plates at 1% hematocrit, 1% starting parasitemia and 40 μl of 0.5% Albumax culture media. Growth at 72 h was measured by SYBR Green I (Lonza, Visp, Switzerland) staining of parasite DNA. A 24-point dilution of PPQ (Sigma-Aldrich, St. Louis, MO) and a 12-point dilution series of the rest of the drugs (DHA, ART, AS, MEF, and LUM; Sigma-Aldrich, St. Louis, MO) were carried out in triplicate and repeated with at least three biological replicates. Drug stocks were resuspended in dimethyl sulfoxide except for CQ prepared in 0.1% Triton X-100 in water and PPQ prepared in 0.1% Triton X-100 and 0.5% lactic acid in water to ensure complete dissolution, as lactic acid enhanced PPQ solubility45 [/articles/s41467-018-04104-z#ref-CR45]. Drugs dispensed by a HP D300 Digital Dispenser (Hewlett Packard Palo Alto, CA), with the Triton X-100 enhancing dispensing of aqueous drug stocks. Relative fluorescence units was measured at an excitation of 494 nm and emission of 530 nm on a SpectraMax M5 (Molecular Devices Sunnyvale, CA) and analyzed using GraphPad Prism version 7 (GraphPad Software La Jolla, CA). EC50 values were determined with the curve-fitting algorithm log(inhibitor) vs. response–Variable slope except for PPQ. PPQs bimodal dose–response would not allow for any curve-fitting hence susceptibility was measured by calculating the area under the second response curve (AUC). AUC was calculated by fitting a six-dimensional least-square polynomial equation to each sample’s dose–response curve and integrating the fitted equation over the region corresponding to the second response curve. These equations were fit to the data using the polyfit module in NumPy, a fundamental package for scientific computing using Python. We chose to use a six-dimensional least-square polynomial equation because it captures the dynamics of the second response curve better than equations with fewer dimensions. Using equations with dimensions > 6 do not result in major differences in calculated AUC values. In fact, AUC values calculated using the boundaries identified by our equation fitting method (0.10 μM – 30 μM) gave roughly the same result. The boundaries of the second response curve were determined by identifying the drug concentration corresponding to the average local minima before and after the second response curve across all drug-assayed samples

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Expression-based drug screening of neural progenitor cells from individuals with schizophrenia

ed in this study were provided by Eli Lilly as 10 mM stocks in DMSO, in heat-sealed 96-well plates or purchased from Tocris and dissolved in DMSO (Sigma Catalog Number D5879, Lot Number SHBF7682V). HP D300 T8 cassettes for pilot experiment were from Tecan. All other automated liquid handling consumables, experiments in stages 1 and 2, were from Perkin Elmer. Concentration of each drug was select;ed in this study were provided by Eli Lilly as 10 mM stocks in DMSO, in heat-sealed 96-well plates or purchased from Tocris and dissolved in DMSO (Sigma Catalog Number D5879, Lot Number SHBF7682V). HP D300 T8 cassettes for pilot experiment were from Tecan. All other automated liquid handling consumables, experiments in stages 1 and 2, were from Perkin Elmer. Concentration of each drug was select

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Ezh2 inhibition in Kras-driven lung cancer amplifies inflammation and associated vulnerabilities

, 3.8 µM TGF-α, 1.2 µM IL-6, 0.9 µM TNF-α, 0.9 µM IFN-γ, and 4.0 µM LPS) with and without the addition of 5 µM GSK126. After 24 h, BTZ was added in a dosage range of 0.01 to 300 nM using the TECAN d300e compound printer, including four replicates per dosage condition with randomized well distribution and DMSO normalization. GSK126 treatment was renewed 24 h after the application of BTZ us

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Integrative Modeling Identifies Key Determinants of Inhibitor Sensitivity in Breast Cancer Cell Lines

In the case of the validation experiments with HCC1806 and HCC1937 cell lines expressing 4E-BP1 or GFP constructs, cells were treated in a 9-point 1:3 dilution series of AZD8055, BEZ235, or GDC0941 using a HP D300 Digital Dispenser (HewlettPackard), while all other experimental conditions remained the same. Each assay was carried out in biological triplicate. Each replicate of a dose–response experiment was further analyzed by normalization to the negative and positive control (the normalized data are provided in Supplementary Table S3) and fitting to a four-parameter sigmoid function that allowed for the calculation of the IC50 (dose at which viability is 50% of the untreated control). The IC50 estimates are provided in Supplementary Table S4. For model inference, full dose–response curve data were used

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Elucidating Gene-by-Environment Interactions Associated with Differential Susceptibility to Chemical Exposure

were distributed into six 96-well plates, with 12 individuals per plate exposed to a concentration (0, 0.03, 0.1, 0.3, 1, 3, 5, or 10μM) using a 20-mM stock of Abamectin digitally dispensed using an HP D300e (Hewlitt Packard HP D300e). Second, a narrower range including 0, 0.5, 0.7, and 0.9μM was performed using 192 individuals (48 per concentration). From these data, the final critical concentr;First, 576 individual zebrafish embryos were distributed into six 96-well plates, with 12 individuals per plate exposed to a concentration (0, 0.03, 0.1, 0.3, 1, 3, 5, or 10 μ M ) using a 20 -mM stock of Abamectin digitally dispensed using an HP D300e (Hewlitt Packard HP D300e)

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Structural insight into allosteric modulation of protease-activated receptor 2

Compounds were dissolved in DMSO to 20 mM and a digital dispenser HP D300 (Tecan) was employed to set up the concentration response experiments. Prior to the analysis of compound binding, three running buffer blanks were injected to equilibrate the instrument

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Physiologic Medium Rewires Cellular Metabolism and Reveals Uric Acid as an Endogenous Inhibitor of UMP Synthase

SW620 cells were seeded in white 96-well plates (Greiner) at a density of 2000 cells/well and allowed to attach for 24 hr. 5-FU and doxorubicin were prepared in DMSO and dispensed using an HP D300 compound dispenser

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Direct Activation of BAX by BTSA1 Overcomes Apoptosis Resistance in Acute Myeloid Leukemia

Dilutions of compounds was performed using a TECAN D300e Digital Dispenser from 10 mM stocks

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Efficient measurement and factorization of high-order drug interactions in Mycobacterium tuberculosis

m inhibitory concentration for each drug is given in Table 1. Assays were performed in clear, flat-bottom 384-well microplates by dispensing nanoliter volumes of drugs using a digital drug dispenser (D300e Digital Dispenser, HP). Dispense locations were randomized within each plate using the software to minimize the plate position effect. Mid-log phase cultures were diluted to an optical density (

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Cannabinoid CB2 receptor ligand profiling reveals biased signalling and off-target activity

The cells were incubated for 90 min in a humidified atmosphere at 37 °C and 5% CO2. Compounds in DMSO stocksolutions were added using a HP D300 Digital Dispenser (Tecan, Männedorf, Switzerland)

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Adaptive resistance of melanoma cells to RAF inhibition via reversible induction of a slowly dividing de-differentiated state

tion plates and a Seiko pin transfer robot system. Immediately after, A375 and WM115 cells were then treated with 1 μM vemurafenib and COLO858 cells were treated with 0.32 μM vemurafenib using an HP D300 Digital Dispenser. Forty‐eight hours after treatment, cells were fixed and analyzed for NGFR expression using immunofluorescence microscopy, as described earlier. To identify candidate compo;tion plates and a Seiko pin transfer robot system. Immediately after, A375 and WM115 cells were then treated with 1 μM vemurafenib and COLO858 cells were treated with 0.32 μM vemurafenib using an HP D300 Digital Dispenser. Forty‐eight hours after treatment, cells were fixed and analyzed for NGFR expression using immunofluorescence microscopy, as described earlier. To identify candidate compo

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Proliferation Markers Are Associated with MET Expression in Hepatocellular Carcinoma and Predict Tivantinib Sensitivity In Vitro

cells were treated with 5 concentrations of each compound (10-fold dilution from 0.001 to 10 mmol/L in duplicates) using the HP D300 digital dispenser (Tecan). Cell viability was measured 48 hours after drug treatment by colorimetric MTS assay following the supplier's recommendations (Promega). Each experiment was repeated at least twice for each cell line, and results were normalized on untreated cells. Curve fitting of dose–response data was performed using GraphPad Prism 6 Software, and the two following classical parameters representative of drug sensitivity were derived: (i) the GI50 corresponding to the concentration of drug that inhibits 50% of cell viability and (ii) the AUC corresponding to the area under the dose–response curve that provides an overall measure of cumulative response. When the GI50 was not reached, the values were set to the highest concentration tested (10 mmol/L)

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Genomic Activation of PPARG Reveals a Candidate Therapeutic Axis in Bladder Cancer

Assays were performed by seeding 384-well plates with 10,000 cells per well in MEMalpha containing 10% FBS and dosing compounds at indicated concentration using the HP D300 digital dispenser (HP/Tecan)

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Therapeutic efficacy of the small molecule GS-5734 against Ebola virus in rhesus monkeys

ted in BSL-4 at USAMRIID. HeLa or HFF-1 cells were seeded at 2,000 cells per well in 384-well plates. Ten serial dilutions of compound in triplicate were added directly to the cell cultures using the HP D300 digital dispenser (Hewlett Packard, Palo Alto, CA) in 2-fold dilution increments starting at 10 μM at 2 h prior to infection. The DMSO concentration in each well was normalized to 1% using an;ted in BSL-4 at USAMRIID. HeLa or HFF-1 cells were seeded at 2,000 cells per well in 384-well plates. Ten serial dilutions of compound in triplicate were added directly to the cell cultures using the HP D300 digital dispenser (Hewlett Packard, Palo Alto, CA) in 2-fold dilution increments starting at 10 μM at 2 h prior to infection. The DMSO concentration in each well was normalized to 1% using an

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Targeting the PI3K/mTOR Pathway in Lymphoma with PQR309 and PQR620: Single Agent Activity and Synergism with the BCL2 Inhibitor Venetoclax

IC50) was obtained in lymphoma cell lines [diffuse large B cell lymphoma (DLBCL), no.=26; mantle cell lymphoma (MCL), no.=8; anaplastic large T-cell lymphoma, no.=5; others, no=5] exposed to increasing doses of PQR620 for 72h using a Tecan D300e Digital Dispenser on ;obtained in lymphoma cell lines [diffuse large B cell lymphoma (DLBCL), no.=26; mantle cell lymphoma (MCL), no.=8; anaplastic large T-cell lymphoma, no.=5; others, no=5] exposed to increasing doses of PQR620 for 72h using a Tecan D300e Digital Dispenser on 384well

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Quantitative CRISPR interference screens in yeast identify chemical-genetic interactions and new rules for guide RNA design

Growth rates were determined by measuring the OD 600 approximately every 15 min for at least 80 cycles at 30 °C in TECAN sunrise or GENios plate readers. Drugs were dissolved in DMSO and dispensed to plates using an HP D300 Digital Dispenser (Tecan)

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Light-controlled modulation of gene expression by chemical optoepigenetic probes

vine serum albumin (BSA), pH 7.4) and 100 μL/well dispensed (Multidrop, Thermofisher) into white 96-well plates (Corning) followed by dispensing of test compounds from 10 mM stock solutions using an HP D300 Digital Dispenser (Hewlett-Packard). The LED-array was then placed on top of the 96-well plate and enzyme inhibitor solutions were exposed to light for 1 h (470 nm, 17 mW/cm2). Light exposure;vine serum albumin (BSA), pH 7.4) and 100 μL/well dispensed (Multidrop, Thermofisher) into white 96-well plates (Corning) followed by dispensing of test compounds from 10 mM stock solutions using an HP D300 Digital Dispenser (Hewlett-Packard). The LED-array was then placed on top of the 96-well plate and enzyme inhibitor solutions were exposed to light for 1 h (470 nm, 17 mW/cm2). Light exposure

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A PHGDH inhibitor reveals coordination of serine synthesis and one-carbon unit fate

th a ViewLux uHTS Microplate Imager (PerkinElmer). Follow-up assays were performed in black 384-well plates (Greiner) in 20 µL of enzyme buffer to which compounds were added in dose-response with an HP D300 digital dispenser (Hewlett-Packard), followed by addition of 20 µL of substrate buffer. Plates were incubated at room temperature (25 °C) and read at 0 and 20 minutes with a Spectramax M5 pl;Page 1. © 201 6 Nature America, Inc. All rights reserved. 452 nature chemical biology | vol 12 | June 2016 | www.nature.com/naturechemicalbiology ARTICLE published online: 25 april 2016 | doi: 10.1038/nchembio.2070 One

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N-methylation of a bactericidal compound as a resistance mechanism in Mycobacterium tuberculosis

Compounds were dispensed in a black 384-well low-volume microplate (Greiner Bio-One) using a Hewlett Packard HP D300 digital dispenser (Tecan Group Ltd.)

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Suramin inhibits cullin-RING E3 ubiquitin ligases

Compound stock solutions in DMSO were titrated into 384-well plates using an HP D300 digital dispenser

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High-Order Drug Combinations Are Required to Effectively Kill Colorectal Cancer Cells

For pairwise and triple combinations, compound doses were combined at a fixed ratio of 1:1 and 1:1:1, respectively (17). DMSO concentrations were normalized per well. In addition, vehicle (DMSO) and positive controls (staurosporine) were transferred. All treatments were performed in triplicates. One of the replicates received 50 nL of 2 mmol/L CellEvent Caspase-3/7 Green Detection Reagent (Thermo) using a HP D300 dispenser (Tecan). Cells were treated for 72 to 96 hours. Caspase-3/7 activity was imaged every 24 hours. At the end of the treatment, cells were fixed, and their DNA was stained and imaged (18, 19). High-order experiments combining more than 3 drugs were also setup with fixed ratios but only testing 4 dose points. To validate synergies of MDM2 combinations, matrices were tested to assess effects at different ratios of the single agents

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Molecular Landscape of Acquired Resistance to Targeted Therapy Combinations in BRAF-Mutant Colorectal Cancer

At 24 hours post-seeding, 100 µl of serum-free medium with or without cetuximab (5 μg/ml) was manually added to the cells. All other drugs were added directly on the plate by TECAN D300e digital dispenser (HP). After 72 hours’ treatment cell viability was assessed by ATP content using CellTiter-Glo Luminescent Assay (Promega). Viability was normalized as a percentage of control untreated cells. Data from growth-inhibition assays were plotted using the nonlinear regression curve fit modelling from GraphPad Prism-5 (GraphPad Software)

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Tecan D300e Digital Dispenser is a product of HP Inc. CA, USA.