Synergy h1 hybrid multi mode plate reader

Manufactured by Agilent Technologies

Sourced in United States

The Synergy H1 Hybrid Multi-Mode Plate Reader is a versatile laboratory instrument designed for a wide range of applications. It is capable of performing absorbance, fluorescence, and luminescence measurements on microplates. The device features a monochromator-based optical system and a high-performance detection system to provide accurate and reliable results.

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Lab products found in correlation

Prism 9, by GraphPad (2 mentions) Elisa 96 well plates, by Corning (2 mentions) Rbd mouse fc tagged antigen, by Sino Biological (2 mentions) El406 washer dispenser bsl2 m, by Agilent Technologies (2 mentions) Nanodrop 2000 spectrophotometer, by Thermo Fisher Scientific (2 mentions) Bovine serum albumin (bsa), by HiMedia (2 mentions) One glo ex, by Promega (1 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions) Glycogen, by Thermo Fisher Scientific (1 mentions) Trypan blue, by Thermo Fisher Scientific (1 mentions) Stereo discovery v8 microscope, by Zeiss (1 mentions) Secondary goat anti mouse igg, by Southern Biotech (1 mentions) Nunclon delta surface 96 well plate, by Thermo Fisher Scientific (1 mentions) Celltiter glo 3d cell viability assay, by Promega (1 mentions) Prl tk dna, by Promega (1 mentions) Prl tk plasmid, by Promega (1 mentions) Dual luciferase reporter assay system, by Promega (1 mentions) Fugene 6, by Promega (1 mentions) Celltiter glo 3d reagent, by Promega (1 mentions) Pnpp substrate, by Merck Group (1 mentions)

Close spelling variants of this product

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20 protocols using synergy h1 hybrid multi mode plate reader

Bacterial Expression and Characterization of sFSTING

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SfSTING and mutant constructs as well as an sfGFP negative-control construct were cloned into pET vectors for IPTG-inducible expression. E. coli BL21 (DE3) (NEB) were transformed with these plasmids and then plated on LB medium plates supplemented with 100 μg ml⁻¹ ampicillin. After overnight incubation, three colonies from these plates were used to inoculate 5-ml MDG liquid cultures (0.5% glucose, 25 mM Na₂HPO₄, 25 mM KH₂PO₄, 50 mM NH₄Cl, 5 mM Na₂SO₄, 2 mM MgSO₄, 0.25% aspartic acid and trace metals) supplemented with 100 μg ml⁻¹ ampicillin and grown overnight at 37 °C with 230 r.p.m. shaking. Cultures were diluted 1:50 into fresh M9ZB medium (supplemented with 100 μg ml⁻¹ ampicillin) and grown for 3 h at 37 °C with 230 r.p.m. shaking. Cultures were then diluted to a uniform OD_600nm in M9ZB medium and further diluted 1:5 into fresh M9ZB medium supplemented with 5 μM IPTG to induce protein expression. A 200 μl volume of induced culture was added to a 96-well plate in technical triplicate and OD_600nm was recorded over 300 min in a Synergy H1 Hybrid Multi-Mode Plate Reader (BioTek) shaking at 37 °C. Plots were generated with GraphPad Prism 9.3.0.

Morehouse B.R., Yip M.C., Keszei A.F., McNamara-Bordewick N.K., Shao S, & Kranzusch P.J. (2022). Cryo-EM structure of an active bacterial TIR–STING filament complex. Nature, 608(7924), 803-807.

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Pseudovirus Entry Assay in ACE2-expressing 293T Cells

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HEK293T (293T) cells (ATCC CRL-11268) were cultured in 10% FBS, 1% PenStrep DMEM at 37°C in a humidified 8% CO₂ incubator. Transient transfection of mouse ACE2 in 293T cells was done 18–24 hours prior to infection using Lipofectamine 2000 (Life Technologies) and an HDM plasmid containing full length Mouse ACE2 (GenBank: Q8R010, synthesized by GenScript) in OPTIMEM. After 5 hr incubation at 37°C in a humidified 8% CO₂ incubator, DMEM with 10% FBS was added and cells were incubated at 37°C in a humidified 8% CO₂ incubator for 18–24 hr. Immediately prior to infection, 293T cells with transient expression of mouse ACE2 were washed with DMEM 1x, then plated with pseudovirus at a 1:75 dilution in DMEM. Infection in DMEM was done with cells between 60–80% confluence for 2.5 hr prior to adding FBS and PenStrep to final concentrations of 10% and 1%, respectively. Following 18–24 hr of infection, One-Glo-EX (Promega) was added to the cells and incubated in the dark for 5 min before reading on a Synergy H1 Hybrid Multi-Mode plate reader (Biotek). Cell entry levels of pseudovirus generated on different days (biological replicates) were plotted in GraphPad Prism as individual points, and average cell entry across biological replicates was calculated as the geometric mean.

Cameroni E., Bowen J.E., Rosen L.E., Saliba C., Zepeda S.K., Culap K., Pinto D., VanBlargan L.A., De Marco A., di Iulio J., Zatta F., Kaiser H., Noack J., Farhat N., Czudnochowski N., Havenar-Daughton C., Sprouse K.R., Dillen J.R., Powell A.E., Chen A., Maher C., Yin L., Sun D., Soriaga L., Bassi J., Silacci-Fregni C., Gustafsson C., Franko N.M., Logue J., Iqbal N.T., Mazzitelli I., Geffner J., Grifantini R., Chu H., Gori A., Riva A., Giannini O., Ceschi A., Ferrari P., Cippà P.E., Franzetti-Pellanda A., Garzoni C., Halfmann P.J., Kawaoka Y., Hebner C., Purcell L.A., Piccoli L., Pizzuto M.S., Walls A.C., Diamond M.S., Telenti A., Virgin H.W., Lanzavecchia A., Snell G., Veesler D, & Corti D. (2021). Broadly neutralizing antibodies overcome SARS-CoV-2 Omicron antigenic shift. Nature, 602(7898), 664-670.

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TRIzol RNA Extraction Optimization

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A standard TRIzol extraction protocol was followed using 10 μg of glycogen (Thermo‐Fisher Scientific, R0551 or AM9515) to facilitate precipitation of RNA. RNA was precipitated in isopropanol (0.5 mL per mL of TRIzol reagent), centrifuged at 13,500 rpm for 30 min at 4°C and washed with 75% nuclease‐free ethanol (1 ml per ml of TRIzol reagent). The RNA pellet was then resuspended in 20 μl of nuclease‐free water. Quantification and analysis was completed with Nanodrop 2000 spectrophotometer (Thermo Fisher Scientific, ND‐2000) or a Synergy H1 Hybrid Multi–Mode plate reader (BioTek).

McCann J., Sosa‐Miranda C.D., Guo H., Reshke R., Savard A., Zardini Buzatto A., Taylor J.A., Li L, & Gibbings D.J. (2022). Contaminating transfection complexes can masquerade as small extracellular vesicles and impair their delivery of RNA. Journal of Extracellular Vesicles, 11(10), e12220.

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Burst and Sustained IgG Release Assay

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Burst release assays were conducted by injecting 100 μL of PNP hydrogel loaded with fluorescently-labeled rat IgG into a microcentrifuge tube filled with 400 μL of PBS. After 5 minutes, 300 μL of the supernatant was removed and analyzed to determine the amount of FITC-labeled IgG using a Synergy H1 Hybrid Multi-Mode Plate Reader (BioTek). Release assays were conducted by adding 100 μL hydrogel to microcentrifuge tubes and then centrifuging until the hydrogel was situated in the bottom of the tube. Then PBS was added on top of the hydrogel. At regular intervals, aliquots of the supernatant were taken for measurement in the plate reader as described above, and the sampled volume was replaced with PBS.

Kasse C.M., Yu A.C., Powell A.E., Roth G.A., Liong C.S., Jons C.K., Buahin A., Maikawa C.L., Zhou X., Youssef S., Glanville J.E, & Appel E.A. (2023). Subcutaneous delivery of an antibody against SARS-CoV-2 from a supramolecular hydrogel depot. Biomaterials Science, 11(6), 2065-2079.

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Quantifying Cell Migration and Invasion

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After HeLa cells grew to confluence in 6-well culture plates, uniform wound scratch lines were made within the marked location by using a pipette tip. The cells were allowed to migrate and close the wound in culture media with and without 1 μg/mL of doxycycline. The pictures of the scratch were captured at 0, 6, 24, and 48 hours by using a Zeiss SteREO Discovery V8 microscope (Zeiss). The wound gaps were measured by using ImageJ software (National Institutes of Health). Boyden chamber cell invasion was performed by using CytoSelect Colorimetric Cell Invasion Assay (Cell Biolabs). HeLa and 32D cells were starved in serum-free medium for 14 hours. Cells were plated in the upper chamber and allowed to migrate through a membrane filter for 48 hours with serum added to the lower chamber as chemoattractant. For HeLa cells, the migrated cells were stained by using crystal violet and counted under light microscope. The migrated cells were then harvested and lysed by using the extraction reagent, and the optical densities were measured at 560 nm by using the Synergy H1 Hybrid Multi-mode plate reader (BioTek). For 32D cells, the migrated cells were stained with 0.4% trypan blue (Thermo Fisher Scientific) and counted manually. Pictures were taken by using the Nuance Multispectral Imaging System.

Mahat U., Garg B., Yang C.Y., Mehta H., Hanna R., Rogers H.J., Flagg A., Ivanov A.I, & Corey S.J. (2022). Lymphocyte cytosolic protein 1 (L-plastin) I232F mutation impairs granulocytic proliferation and causes neutropenia. Blood Advances, 6(8), 2581-2594.

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RBD-hACE2 Binding Inhibition Assay

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Blockade of WT RBD binding to hACE2 was performed, as previously described (Piccoli et al., 2020 (link)). Unlabeled mAbs were serially diluted, mixed with RBD mouse Fc-tagged antigen (Sino Biological, final concentration 20 ng/mL) and incubated for 30 min at 37°C. The mix was added for 30 min to ELISA 96-well plates (Corning) pre-coated overnight at 4°C with 2 μg/mL hACE2 in PBS. Plates were washed (EL406 washer/dispenser BSL2 M, Biotek) and RBD binding was revealed using a secondary goat anti-mouse IgG (Southern Biotech). After washing, pNPP substrate was added and plates were read at 405 nm (Synergy H1 Hybrid Multi-Mode plate reader, Biotek). The percentage of inhibition was calculated as follow: (1−(OD sample−OD neg ctr)/(OD pos ctr−OD neg ctr)) × 100.

Thomson E.C., Rosen L.E., Shepherd J.G., Spreafico R., da Silva Filipe A., Wojcechowskyj J.A., Davis C., Piccoli L., Pascall D.J., Dillen J., Lytras S., Czudnochowski N., Shah R., Meury M., Jesudason N., De Marco A., Li K., Bassi J., O’Toole A., Pinto D., Colquhoun R.M., Culap K., Jackson B., Zatta F., Rambaut A., Jaconi S., Sreenu V.B., Nix J., Zhang I., Jarrett R.F., Glass W.G., Beltramello M., Nomikou K., Pizzuto M., Tong L., Cameroni E., Croll T.I., Johnson N., Di Iulio J., Wickenhagen A., Ceschi A., Harbison A.M., Mair D., Ferrari P., Smollett K., Sallusto F., Carmichael S., Garzoni C., Nichols J., Galli M., Hughes J., Riva A., Ho A., Schiuma M., Semple M.G., Openshaw P.J., Fadda E., Baillie J.K., Chodera J.D., Rihn S.J., Lycett S.J., Virgin H.W., Telenti A., Corti D., Robertson D.L, & Snell G. (2021). Circulating SARS-CoV-2 spike N439K variants maintain fitness while evading antibody-mediated immunity. Cell, 184(5), 1171-1187.e20.

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Intracellular ATP Quantification for Cell Viability

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Intracellular ATP was used as a readout indicating the metabolically active stage of viable cells. The CellTiter-Glo^® 3D Cell Viability Assay (Promega, United States) was deployed for direct detection of the intracellular level of ATP. ATPs can change luciferin to a luminescence-emitting oxyluciferin. Briefly, the neurospheroids and the separated trophozoites were transferred to a low attachment round-bottom well of the 96-well plate (SPL Life Sciences, South Korea) and a Nunclon Delta Surface 96-well plate (Thermo Fisher Scientific, MA), respectively, followed by adding the CellTiter-Glo^® solution. The luminescence signal was measured by using a BioTek Synergy H1 Hybrid Multi-Mode plate reader. After removing the background luminescence of the culture medium, the data were displayed as relative light units (RLUs). Changes in the RLUs of each sample were related to the RLUs of control neurospheroids that were free from B. mandrillaris trophozoites.

Whangviboonkij N., Pengsart W., Chen Z., Han S., Park S, & Kulkeaw K. (2023). Phenotypic assay for cytotoxicity assessment of Balamuthia mandrillaris against human neurospheroids. Frontiers in Microbiology, 14, 1190530.

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FOXO Transcription Factor Activity Assay

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The activity of the FOXO transcription factor was assessed using the pGL-3 × DBE plasmid (Promega)^{43 (link)}, which incorporates three iterations of FOXO-responsive promoters and encodes firefly luciferase. Concurrently, the pRL-TK plasmid (Promega), expressing renilla luciferase constitutively, was transfected as an internal control. Astrocytes were seeded in 60 mm diameter plates (1 × 10⁶ cells/plate) and transfected after 48 h (approximately 60% confluence) using Fugene 6 (Promega) with 1 µg DBE DNA, 0.2 µg pRL-TK DNA, and a DNA: Fugene ratio of 1:6. Following 48 h of transfection, the cells were treated by washing twice with phosphate buffered saline (PBS) and incubating for 24 h in DMEM containing either 4.5 g/L or 1 g/L glucose concentration, with or without 1 nM Klotho. Post-treatment, the cells were lysed as per the Dual-Luciferase^® Reporter Assay system (Promega) guidelines. Equal sample volumes were used, and the luminescence was developed and read according to the manufacturer's instructions (Promega) on opaque white plates using a Synergy H1 Hybrid Multi-Mode plate reader (BioTek). The results are presented as the ratio of firefly luciferase fluorescence to renilla luciferase, normalized to the mean of the control group.

Orellana A.M., Mazucanti C.H., dos Anjos L.P., de Sá Lima L., Kawamoto E.M, & Scavone C. (2023). Klotho increases antioxidant defenses in astrocytes and ubiquitin–proteasome activity in neurons. Scientific Reports, 13, 15080.

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Cell Spheroid Metabolic Activity Assay

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A viability assay was performed at 48 and 96 h post-treatment to determine the metabolic activity of the cell spheroids. According to the manufacturers’ instructions, 150 μL of the medium was removed from each well of the 96-well clear F-bottom black plate (Greiner Bio-One GmbH (Frickenhausen, Germany)) and 50 μL of CellTiter-Glo^® 3D Reagent (Promega GmbH (Madison, WI, USA)) was added. Spheroids were then incubated at room temperature for 30 min in the dark to stabilise the bioluminescent signal, which was then measured using the Synergy H1™ Hybrid Multi-mode plate reader (BioTek Instruments^®, Inc. (Winooski, VE, USA)).

Landgraf L., Kozlowski A., Zhang X., Fournelle M., Becker F.J., Tretbar S, & Melzer A. (2022). Focused Ultrasound Treatment of a Spheroid In Vitro Tumour Model. Cells, 11(9), 1518.

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Measuring H2O2 generation in INS1 cells

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Stably transfected mCherry‐HyPer‐PTS1‐2 INS1 β cells (50 000/well) were seeded in 24‐well plates in regular medium and kept at 37°C, 5% CO₂ for 48 hours. Medium was removed and replaced with PBS with 11 mmol/L glucose with or without 2.5 or 5 nmol/L C‐peptide (Phoenix Pharmaceuticals) for 20 minutes at 37°C, 5% CO₂ before adding 100 or 200 μmol/L palmitic acid for 30 minutes at 37°C, 5% CO₂. Cells exposed to equimolar amounts of BSA were used as a control. Generation of H₂O₂ was assayed by immediately measuring fluorescence of HyPer (excitation 488; emission 510 nm) and of mCherry (excitation at 587; emission at 610) in each well using the BioTek Synergy H1 Hybrid Multi‐Mode plate reader. Each condition was tested in a total of 18 independent experiments. Hyper fluorescence was normalized to the mCherry fluorescence in each well and expressed as Hyper‐live‐cell fluorescence arbitrary units mean ± SEM

Luppi P., Drain N., To R., Stolz D., Wallace C., Watkins S, & Drain P. (2020). Autocrine C‐peptide protects INS1 β cells against palmitic acid‐induced oxidative stress in peroxisomes by inducing catalase. Endocrinology, Diabetes & Metabolism, 3(3), e00147.

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