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D300e digital dispenser

Manufactured by Tecan
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Sourced in Switzerland, Germany, France, United States

The D300e Digital Dispenser is a precision liquid handling instrument designed for accurate and reproducible dispensing of small volumes of liquids. The device utilizes digital technology to provide reliable and consistent liquid delivery within a specified volume range.

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Lab products found in correlation

Resazurin, by Merck Group (8 mentions) F200 pro microplate reader, by Tecan (8 mentions) Clariostar, by BMG Labtech (7 mentions) Celltiter glo luminescent cell viability assay, by Promega (6 mentions) Multidrop combi reagent dispenser, by Thermo Fisher Scientific (5 mentions) Alamarblue cell viability reagent, by Thermo Fisher Scientific (4 mentions) Celltiter glo 3d cell viability assay, by Promega (4 mentions) Celltiter glo luminescent assay, by Promega (4 mentions) Hyclone trypan blue, by Cytiva (4 mentions) Infinite m1000 pro plate reader, by Tecan (4 mentions) Victor multilabel plate reader, by PerkinElmer (4 mentions) Envision multilabel plate reader, by PerkinElmer (3 mentions) Celltiter glo assay, by Promega (3 mentions) Tc20 automated cell counter, by Bio-Rad (3 mentions) Celltiter glo assay reagent, by Promega (3 mentions) Prism software, by GraphPad (3 mentions) Multiflo, by Agilent Technologies (2 mentions) Infinite m1000 pro, by Tecan (2 mentions) Celltiter glo reagent, by Promega (2 mentions) Tryple express, by Thermo Fisher Scientific (2 mentions)

114 protocols using d300e digital dispenser

1

Evaluating Efflux Pump Inhibitors on Antibiotic Efficacy

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MIC experiments were carried out according to the CLSI guidelines (CLSI 2023 (link)). Overnight cultures were diluted to an OD600 = 0.001 in CAMHB followed by addition of efflux pump inhibitors (EPIs) at a final concentration of 100 µM. Afterwards, 50 µL of bacterial suspension were transferred to 384-well flat-bottomed microtitre plates (Greiner) where a dose-response of the antibiotics of interest were added by serial dilutions (range of 0–128 mg/L) using a Tecan D300e Digital Dispenser (Tecan, France). Plates were incubated at 37 °C for up to 20 h. Bacterial viability was evaluated using the resazurin microtitre assay (REMA) and measured by fluorescence using a Tecan Spaert multimode plate reader (Ex: 530 nm Em: 590 nm). MICs were defined as the concentration that prevented 90% of resazurin turnover compared to the non-treated bacteria.
To evaluate the efficacy of BDM88855.HCl (1’), BDM91288 (5) and PAβN; their concentration-dependent boosting of levofloxacin activity was determined using a checkerboard assay. 384-well flat-bottomed microtitre plates containing a dose-response of each EPI (concentration range = 0–300 µM) and levofloxacin (concentration range = 0–1 mg/L) were prepared with a Tecan D300e Digital Dispenser (Tecan, France). Bacterial suspensions, incubation and plate readouts were carried out as described above.
Vieira Da Cruz A., Jiménez-Castellanos J.C., Börnsen C., Van Maele L., Compagne N., Pradel E., Müller R.T., Meurillon V., Soulard D., Piveteau C., Biela A., Dumont J., Leroux F., Deprez B., Willand N., Pos K.M., Frangakis A.S., Hartkoorn R.C, & Flipo M. (2023). Pyridylpiperazine efflux pump inhibitor boosts in vivo antibiotic efficacy against K. pneumoniae. EMBO Molecular Medicine, 16(1), 93-111.
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2

Investigating Protein Degradation Pathways in HEK293T Cells

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HEK293TCas9 cells were resuspended at 0.15 × 106 per mL and plated on a 384 well plate with 50 μl per well and MLN4924, MLN7243 or MG132 with or without CR8 serially diluted with D300e Digital Dispenser (Tecan Inc.).
HEK293TCas9 cells (0.625 × 106 cells/6 well plate format) were seeded the day before transfection. The following day, 2.5 ug of pRSF91-GFP or pRSF91-CRBN9 (link) plasmid DNA was mixed with 250 μl OptiMem and 7.5 μl TransIT-LT1 (Mirus Bio) according to manufacture protocol. 48 hours post transfection cells were resuspended at 0.15 × 106 cells /mL and plated on a 384 well plate with 50 μl per well.
HEK293TCas9 cells were transduced with sgRNAs targeting either DDB1 or Luciferase in pXPR003 backbone (GPP) (Supplementary Oligo Table 1). After nine days of puromycin selection, cells were re-plated into a 96-well format with 2 × 104 cells per well and CR8 and Roscovitine were serially diluted with D300e Digital Dispenser (Tecan Inc.).
After 3 days of drug exposure, cell viability was assessed using the CellTiter-Glo luminescent assay (Promega, #G7572) on an EnVision Multilabel Plate Reader (Perkin Elmer) or CLARIOstar Plus, MARS 3.4 (BMG LabTech). Cell viabilities were calculated relative to DMSO controls.
Słabicki M., Kozicka Z., Petzold G., Li Y.D., Manojkumar M., Bunker R., Donovan K.A., Sievers Q.L., Koeppel J., Suchyta D., Sperling A.S., Fink E.C., Gasser J.A., Wang L.R., Corsello S.M., Sellar R.S., Jan M., Gillingham D., Scholl C., Fröhling S., Golub T.R., Fischer E.S., Thomä N.H, & Ebert B.L. (2020). The CDK inhibitor CR8 acts as a molecular glue degrader that depletes cyclin K. Nature, 585(7824), 293-297.
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3

Investigating Protein Degradation Pathways in HEK293T Cells

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HEK293TCas9 cells were resuspended at 0.15 × 106 per mL and plated on a 384 well plate with 50 μl per well and MLN4924, MLN7243 or MG132 with or without CR8 serially diluted with D300e Digital Dispenser (Tecan Inc.).
HEK293TCas9 cells (0.625 × 106 cells/6 well plate format) were seeded the day before transfection. The following day, 2.5 ug of pRSF91-GFP or pRSF91-CRBN9 (link) plasmid DNA was mixed with 250 μl OptiMem and 7.5 μl TransIT-LT1 (Mirus Bio) according to manufacture protocol. 48 hours post transfection cells were resuspended at 0.15 × 106 cells /mL and plated on a 384 well plate with 50 μl per well.
HEK293TCas9 cells were transduced with sgRNAs targeting either DDB1 or Luciferase in pXPR003 backbone (GPP) (Supplementary Oligo Table 1). After nine days of puromycin selection, cells were re-plated into a 96-well format with 2 × 104 cells per well and CR8 and Roscovitine were serially diluted with D300e Digital Dispenser (Tecan Inc.).
After 3 days of drug exposure, cell viability was assessed using the CellTiter-Glo luminescent assay (Promega, #G7572) on an EnVision Multilabel Plate Reader (Perkin Elmer) or CLARIOstar Plus, MARS 3.4 (BMG LabTech). Cell viabilities were calculated relative to DMSO controls.
Słabicki M., Kozicka Z., Petzold G., Li Y.D., Manojkumar M., Bunker R., Donovan K.A., Sievers Q.L., Koeppel J., Suchyta D., Sperling A.S., Fink E.C., Gasser J.A., Wang L.R., Corsello S.M., Sellar R.S., Jan M., Gillingham D., Scholl C., Fröhling S., Golub T.R., Fischer E.S., Thomä N.H, & Ebert B.L. (2020). The CDK inhibitor CR8 acts as a molecular glue degrader that depletes cyclin K. Nature, 585(7824), 293-297.
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4

Measuring Anchorage-Dependent and -Independent Cell Growth

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To measure anchorage-dependent growth, cells were plated at low density in 96 well plates and allowed to attach overnight. Inhibitors were then added with a D300e Digital Dispenser (Tecan). After 72 hours, cells were labeled with calcein AM (Thermo Fisher) according to the manufacturer’s protocol, and counted on a SpectraMax MiniMax 300 Imaging Cytometer (Molecular Devices).
To measure anchorage-independent growth, soft agar colony formation assays were performed. In a 96-well plate, wells were coated with 50 μl 2% SeaPrep Agarose (Lonza) mixed with cell culture medium. Next, 100 μl of 5000 cells mixed with 1% SeaPrep Agarose were added to each well and allowed to solidify. Fifty μl of medium were added to each well, and drugs were dispensed with a D300e Digital Dispenser (Tecan). After 72 hours, the alamarBlue cell viability reagent (Thermo Fisher) was added and incubated for 3 hours. Fluorescence was quantified on a SpectraMax MiniMax 300 Imaging Cytometer (Molecular Devices).
Blake D.R., Vaseva A.V., Hodge R.G., Kline M.P., Gilbert T.S., Tyagi V., Huang D., Whiten G.C., Larson J.E., Wang X., Pearce K.H., Herring L.E., Graves L.M., Frye S.V., Emanuele M.J., Cox A.D, & Der C.J. (2019). Application of a MYC degradation screen identifies sensitivity to CDK9 inhibitors in KRAS-mutant pancreatic cancer. Science signaling, 12(590), eaav7259.
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5

High-throughput drug synergy screening

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A total of 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,  λex: 530 nm; λem: 590 nm). Synergy analysis was conducted using SynergyFinder software and the Bliss Independence model 92 (link),115 (link).
Zhou Y., Han C., Wang E., Lorch A.H., Serafin V., Cho B.K., Diaz B.T., Calvo J., Fang C., Khodadadi-Jamayran A., Tabaglio T., Marier C., Kuchmiy A., Sun L., Yacu G., Filip S.K., Jin Q., Takahashi Y.H., Amici D.R., Rendleman E.J., Rawat R., Bresolin S., Paganin M., Zhang C., Li H., Kandela I., Politanska Y., Abdala-Valencia H., Mendillo M.L., Zhu P., Palhais B., Van Vlierberghe P., Taghon T., Aifantis I., Goo Y.A., Guccione E., Heguy A., Tsirigos A., Wee K.B., Mishra R.K., Pflumio F., Accordi B., Basso G, & Ntziachristos P. (2020). Posttranslational regulation of the exon skipping machinery controls aberrant splicing in leukemia. Cancer discovery, 10(9), 1388-1409.
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6

Cell-free Ribosome Translation Inhibition Assay

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Cell-free in-vitro translation inhibition assays were performed using luciferase mRNA and bacterial S30 extracts containing either wild-type bacterial or human hybrid ribosomes.55 (link) In brief, firefly luciferase mRNA was transcribed in vitro using T7 RNA polymerase (Thermo) using a plasmid as template in which the mammalian promoter in pGL4.14 (Promega) has been replaced by theT7 bacteriophage promoter. Test articles in aqueous solution containing 0.3% Tween20 were dispensed into white 96-well plates (Eppendorf) using the TECAN D300e digital dispenser. The test article dispension volume was balanced to a total of 1.5 μL by 0.3% Tween20 in water. The reaction volume was brought to 15 μL by addition of 13.5 μL Translation Master Mix comprised of bacterial S30 extract, 0.2 mM amino acid mix, 6 μg tRNA (Sigma), 0.4 μg hFluc mRNA, 0.3 μL protease inhibitor (cOmplete, EDTA-free, Roche), 12 U RNAse inhibitor (Ribolock, Thermo Scientific), and 6 μL S30 premix without amino acids (Promega). Dispension and mixing of reagents was performed on ice prior to incubating the sealed plates at 37 °C. After 30 minutes of incubation, the reaction was stopped on ice and 75 μL of luciferase assay reagent (Promega) was added to each well. Luminescence was recorded with a plate reader (BIO-TEK FLx800, Witec AG, Littau, Switzerland).
Lubriks D., Zogota R., Sarpe V.A., Matsushita T., Sati G.C., Haldimann K., Gysin M., Böttger E.C., Vasella A., Suna E., Hobbie S.N, & Crich D. (2021). Synthesis and Antibacterial Activity of Propylamycin Derivatives Functionalized at the 5”- and Other Positions with a View to Overcoming Resistance due to Aminoglycoside Modifying Enzymes.. ACS infectious diseases, 7(8), 2413-2424.
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7

Trametinib and Refametinib Dose-Response Assay

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Trametinib (Selleck Chemicals, Houston, TX, USA, S2673) and Refametinib (Selleck Chemicals, S1089) were dissolved to 10 mM in DMSO and printed at nine concentrations from 5 nM to 25 µM in triplicates in white-bottom sterile polystyrene 384-well plates (Corning, Wiesbaden, Germany, 3570) using a Tecan D300e Digital Dispenser (Tecan, Männedorf, Switzerland/HP Inc., Palo Alto, CA, USA) and the corresponding software D300e Control (v3.1.3, Tecan). DMSO was printed as a negative control. Pre-printed plates were sealed and stored at −80 °C until used. Prepared assay plates were equilibrated at room temperature and 500 cells per well were seeded in 30 µL culture medium with a Multidrop Combi dispenser (Thermo Fisher Scientific). Assay plates were incubated at 37 °C and 5% CO2 for 5 days (120 h).
Mergener S., Siveke J.T, & Peña-Llopis S. (2021). Monosomy 3 Is Linked to Resistance to MEK Inhibitors in Uveal Melanoma. International Journal of Molecular Sciences, 22(13), 6727.
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8

High-Throughput Cytotoxicity Screening

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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 (link).
Jin Q., Martinez C.A., Arcipowski K.M., Zhu Y., Diaz B.T., Wang K.K., Johnson M.R., Volk A.G., Wang F., Wu J., Grove C., Wang H., Sokirniy I., Thomas P.M., Goo Y.A., Abshiru N.A., Hijiya N., Peirs S., Vandamme N., Berx G., Goosens S., Marshall S.A., Rendleman E.J., Takahashi Y.H., Wang L., Rawat R., Bartom E.T., Collings C.K., Van Vlierberghe P., Strikoudis A., Kelly S., Ueberheide B., Mantis C., Kandela I., Bourquin J.P., Bornhauser B., Serafin V., Bresolin S., Paganin M., Accordi B., Basso G., Kelleher N.L., Weinstock J., Kumar S., Crispino J.D., Shilatifard A, & Ntziachristos P. (2018). USP7 cooperates with NOTCH1 to drive the oncogenic transcriptional program in T cell leukemia. Clinical cancer research : an official journal of the American Association for Cancer Research, 25(1), 222-239.
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9

Cytotoxicity Assay of Small Molecules

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Rhabdomyosarcoma cell lines Rh30 and Rh6 were seeded in 384-well plates at 2500 cells per well, in 30 µL of media, using a BioTek MultiFlo dispenser. Cultures were grown overnight and then treated with drugs FGF401 (23029), PP1 (14244), Ruxolitinib (11609), and Stattic (14590) that were all purchased from Cayman Chemical. All of the pharmaceuticals were administered at the following concentrations using a D300e digital dispenser (Tecan Trading AG): 0.01, 0.0316, 0.1, 0.316, 1, 3.16, and 10 µM. After 72 h of drug treatment, cultures were treated with an equal volume of CellTiter-Glo reagent (Promega 2020-01-29). Luminescence was measured in a Biotek Instruments Inc. plate reader. The data was then analyzed using Prism GraphPad software.
Crawford K.A., Berlow N.E., Tsay J., Lazich M., Mancini M., Noakes C., Huang T, & Keller C. (2020). Case report for an adolescent with germline RET mutation and alveolar rhabdomyosarcoma. Cold Spring Harbor Molecular Case Studies, 6(3), a004853.
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10

Screening PDAC Cell Lines for RMC-7977 Sensitivity

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19 PDAC cell lines were tested for sensitivity to RMC-7977 as part of a panel of human cancer cell lines of various histotypes screened at Crown Bioscience. These PDAC cell lines harbored KRASG12D, KRASG12V, KRASG12C, KRASQ61H, and BRAFV487_P492delinsA mutations. To measure inhibition of cell proliferation, cells were cultured in methylcellulose and treated in triplicates with serial dilutions of RMC-7977 (top concentration of 1 µM) or DMSO dispensed by a Tecan D300e digital dispenser (Tecan Trading AG). Cells were incubated for 120 hours prior to measurement of ATP levels using CellTiter-Glo. Technical triplicates were run for each biological replicate and a total of three biological replicates was done for each cell line. CTG assay readouts were plotted as a function of log molar [inhibitor] and a 4-parameter sigmoidal concentration response model was fitted to the data. Mean ± s.d. was plotted for each tested dilution.
Wasko U.N., Jiang J., Curiel-Garcia A., Wang Y., Lee B., Orlen M., Drizyte-Miller K., Menard M., Dilly J., Sastra S.A., Palermo C.F., Dalton T., Hasselluhn M.C., Decker-Farrell A.R., Chang S., Jiang L., Wei X., Yang Y.C., Helland C., Courtney H., Gindin Y., Zhao R., Kemp S.B., Clendenin C., Sor R., Vostrejs W., Amparo A.A., Hibshman P.S., Rees M.G., Ronan M.M., Roth J.A., Bakir B., Badgley M.A., Chabot J.A., Kluger M.D., Manji G.A., Quintana E., Wang Z., Smith J.A., Holderfield M., Wildes D., Aguirre A.J., Der C.J., Vonderheide R.H., Stanger B.Z., Singh M, & Olive K.P. (2023). Tumor-selective effects of active RAS inhibition in pancreatic ductal adenocarcinoma. bioRxiv.
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