Synergy ht

Manufactured by Tecan

The Synergy HT is a multi-mode microplate reader from Tecan. It is designed to provide accurate and efficient measurement of absorbance, fluorescence, and luminescence in a variety of microplate formats.

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Lab products found in correlation

Infinite m200 pro, by Tecan (3 mentions) Britelite plus, by PerkinElmer (2 mentions) Du800, by Beckman Coulter (1 mentions) Wst 1 assay, by Beyotime (1 mentions) Automated microplate spectrophotometer, by Agilent Technologies (1 mentions) Fitc dextran, by Targetmol (1 mentions)

6 protocols using synergy ht

Lentivirus Neutralization Assay

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Neutralization of three pseudotyped lentiviruses (EBOV, SUDV, or BDBV) was validated with monoclonal antibodies (mAbs) in HEK-293 T cells. Cells were seeded in white-walled, clear-bottom 96-well plates at a density of 20,000 cells per well 1 d before the assay (day 0). On day 1, mAbs (2 µM in HBS with 10% glycerol) were filtered with 0.22-μm sterile membranes and diluted with D10 media. Subsequently, mAbs were serially diluted (10-fold dilution) in D10 media and mixed with lentiviruses (diluted in D10 medium, supplemented with polybrene, 1:1,000, v/v) for 1 h before being transferred to HEK-293 T cells. On day 4, medium was removed, and 100 µL of luciferase substrates (BriteLite Plus, Perkin Elmer) were added to each well. Luminescent signals were recorded on a microplate reader (BioTek Synergy™ HT or Tecan M200). Percent infection was normalized to cells only (0% infection) and virus only (100% infection) on each plate. Neutralization assays were performed in four technique duplicates.

Xu D., Powell A.E., Utz A., Sanyal M., Do J., Patten J.J., Moliva J.I., Sullivan N.J., Davey R.A, & Kim P.S. (2024). Design of universal Ebola virus vaccine candidates via immunofocusing. Proceedings of the National Academy of Sciences of the United States of America, 121(7), e2316960121.

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Bacterial Growth and Fluorescence Measurement

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Bacterial growth and fluorescence measurements were conducted using a BioTek Synergy HT plate reader (units designated as RFU) or a Tecan Infinite M200 Pro plate reader (units designated as RLU_t). To quantify the fluorescence intensity, the BioTek plate reader was equipped with an excitation filter at 645 nm with a 40 nm bandwidth and an emission filter of 710 nm (20 nm bandwidth), while the Tecan plate reader was set to read excitation at 680 nm (9 nm bandwidth) and emission 710 nm (20 nm bandwidth). Cultures were started from single colonies incubated overnight in LB media with selective antibiotics. Prior to experiments, overnight cultures were subsequently subcultured at a 1:100 dilution into fresh LB media with antibiotic. After 2 h (approximately mid-log phase), bacteria were isolated and washed with M9 media to remove any residual LB. E. coli strains were inoculated into experimental conditions at a final OD₆₀₀ of 0.1 (1 cm pathlength, Beckman Coulter DU 800) in final conditions specified within each experiment. Culturing conditions, whether overnight, subcultures, or experiments, were consistently 37 °C while shaking. Growth and fluorescence experiments conducted with the BioTek plate reader were measured every 15 min for OD₆₀₀ and fluorescence at 710 nm.

Nobles C.L., Clark J.R., Green S.I, & Maresso A.W. (2015). A dual component heme biosensor that integrates heme transport and synthesis in bacteria. Journal of microbiological methods, 118, 7-17.

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Cytotoxicity and Cellular Uptake of NAP

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WST-1 assay (Beyotime) was used to determine the cytotoxicity of NAP. In brief, A549 cells (2 × 10³) were seeded in 96-well plate and treated with increasing concentration of NAP (up to 400 μM) for 48 h. WST-1 was added and the plate was read after 1.5 h on an automated microplate spectrophotometer (BioTek Instruments, Vermont, USA) at 450 nm according to the manufacturer’s instructions.
Colony-formation assay was performed to detect long-term survival of A549 cells in NAP treatment. 400 cells seeded in 12-well plates were treated with indicated concentration of NAP for 15 days. The plates were washed twice with PBS and fixed with methanol for 10 min at room temperature and then stained with 1% crystal violet for 5 min. The number of colonies was counted using ImageJ software (version 1.44I).
Cellular uptake assay was performed by using FITC-Dextran (TargetMol). A549 cells were seeded onto 12-well plate and incubated with FITC-Dextran (0.1 mg/mL) for 24 h, then the cells were treated with 50, 100, 150 and 200 μM of NAP for 24 h and 48 h, respectively. The fluorescence (excited at 488 nm and emitted at 530 nm) was measured with a multifunctional microplate reader Spark 10 M (Tecan, Synergy-HT).

Zhang J., Sun Y., Zhong L.Y., Yu N.N., Ouyang L., Fang R.D., Wang Y, & He Q.Y. (2020). Structure-based discovery of neoandrographolide as a novel inhibitor of Rab5 to suppress cancer growth. Computational and Structural Biotechnology Journal, 18, 3936-3946.

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Ebola Virus Neutralization Assay

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Antisera were heat-inactivated (56 °C, 30 min) before neutralization assays. Briefly, HEK-293 T cells were seeded in white-walled, clear-bottom 96-well plates (20,000 cells per well) 1 d before the assay (day 0). On day 1, antisera were serially diluted in D10 media and then mixed with EBOV, SUDV, or BDBV (diluted in D10 medium, supplemented with polybrene, 1:1,000, v/v) for 1 h before being transferred to HEK-293 T cells. On day 4, medium was removed, and 100 µL of luciferase substrates (BriteLite Plus, Perkin Elmer) were added to each well. Luminescent signals were recorded on a microplate reader (BioTek Synergy™ HT or Tecan M200). Percent infection was normalized to cells only (0% infection) and virus only (100% infection) on each plate. Neutralization titers (NT₅₀) were calculated as the serum dilution where a 50% inhibition of infection was observed. Neutralization assays were performed in technical duplicates.

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Antibiotic Minimum Inhibitory Concentration (MIC) Assay

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MIC was defined as the lowest concentration of antibiotic at which no visible growth was detected after 16 h at 37 °C. Overnight cultures were diluted 1:150 in LB broth and added to a 96-well plate. Antibiotics were serially diluted 1:2 and added to columns of the 96-well plate and grown at 37 °C with continuous shaking. Cell growth was measured using optical density at 600 nm (OD₆₀₀). MIC assays were performed in either BioTek Synergy HT or Tecan InfiniteM200 Pro microplate readers.
For MIC calculations performed by WuXi, MIC was calculated as the lowest concentration that inhibits visible growth after 18 h. Bacterial colonies (4–8) of strains of interest were vortexed in saline and adjusted to an OD₆₀₀ of 0.2. Strains were diluted 1:200 into CAMHB media in 96-well plates. Antibiotics were serially diluted 1:3 in DMSO, and 1 μl of each dilution was added to bacteria. The plates were incubated for 18–20 h at 37 °C before observation.

Chain C., Sheehan J.P., Xu X., Ghaffari S., Godbole A., Kim H., Freundlich J.S., Rabinowitz J.D, & Gitai Z. (2024). A folate inhibitor exploits metabolic differences in Pseudomonas aeruginosa for narrow-spectrum targeting. Nature Microbiology, 9(5), 1207-1219.

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Antibiotic Minimum Inhibitory Concentrations

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The minimum inhibitory concentration (MIC) of each antibiotic was defined as the lowest concentration of antibiotic that resulted in no visible growth. MICs were measured by diluting overnight cultures 1:150 and then adding 2-fold dilutions of each antibiotic in 96-well plates and grown with shaking at 37°C. Cell growth was monitored by measuring the OD₆₀₀. Assays performed by Pharmacology Discovery Services (New Taipei City, TW) and Pharmaron Inc. (Beijing, ROC) are indicated in Table S1. For MIC measurements done in-house, OD₆₀₀ readings were measured by either a BioTek Synergy HT (Winooski, VT) or Tecan InfiniteM200 Pro (Männedorf, CH) microplate reader. In locations where drug concentrations are listed, they are included both as a relative fold change compared to the MIC for that bacterial strain and drug combination (1X MIC, 2X MIC, etc) as well as an absolute concentration (1 μg/mL, 2 μg/mL, etc).

Martin JK I.I., Sheehan J.P., Bratton B.P., Moore G.M., Mateus A., Li S.H., Kim H., Rabinowitz J.D., Typas A., Savitski M.M., Wilson M.Z, & Gitai Z. (2020). A Dual-Mechanism Antibiotic Kills Gram-Negative Bacteria and Avoids Drug Resistance. Cell, 181(7), 1518-1532.e14.

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