M200 pro reader

Manufactured by Tecan

Sourced in Switzerland

The M200 Pro reader is a microplate reader designed for absorbance and fluorescence measurements. It is capable of reading microplates with up to 384 wells.

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Lab products found in correlation

Luciferin, by GoldBio (2 mentions) Dexamethasone (dex), by Thermo Fisher Scientific (2 mentions) White 96 well plate, by Greiner (2 mentions) A dmem, by Thermo Fisher Scientific (2 mentions) Multicycle, by Actimetrics (2 mentions) Revertaid h minus first strand cdna synthesis kit, by Thermo Fisher Scientific (2 mentions) Agilent cary 60 uv vis, by Agilent Technologies (1 mentions) Celltiter 96 aqueous one solution cell proliferation assay, by Promega (1 mentions) Adp glo kinase assay, by Promega (1 mentions) Taqman gene expression master mix, by Thermo Fisher Scientific (1 mentions) Rt pcr grade water, by Thermo Fisher Scientific (1 mentions) Rneasy mini kit, by Qiagen (1 mentions) Lightcycler 480, by Roche (1 mentions) Infinite m200 pro plate reader, by Tecan (1 mentions)

10 protocols using m200 pro reader

Continuous OD monitoring of yeast growth

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For continuous OD monitoring (e.g. Figure 3D), cultures were set up in 96-well plates sealed with parafilm in a Tecan M200Pro reader with orbital shaking. OD at wavelength 595 nm was taken every 15 min and analyzed by using the Magellan 7 software (Tecan). For cultures whose OD reading was taken intermittently every several hours (e.g. Figure 5F), the culture plates were placed in non-shaking incubator within a humid chamber before each reading after agitation. The OD at wavelength 595 nm was recorded by using a Tecan M200Pro reader and the data files were processed in R. The growth assays lasted until the fastest growing culture reached saturation, at which time the last OD readings of all strains were recorded.
The drug concentrations of hygromycin B and radicicol were adjusted for different media (e.g., YPD vs SC-ura) and/or the strain background, so that the wild-type control show the same growth delay as was observed for the strain RLY2628 in YPD media containing the stated drug concentration. The growth of genetic variants was normalized to the corresponding wild-type controls.
A comprehensive description of all methods used can be found in Supplemental Information.

Chen G., Mulla W.A., Kucharavy A., Tsai H.J., Rubinstein B., Conkright J., McCroskey S., Bradford W.D., Weems L., Haug J.S., Seidel C.W., Berman J, & Li R. (2015). Targeting the Adaptability of Heterogeneous Aneuploids. Cell, 160(4), 771-784.

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Circadian Rhythm Assay in Fibroblasts

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Fibroblasts were cultured to confluence (~30,000 cells/well) in 96-well white plates (Greiner Bio-One, Gloucestershire, UK) using ADMEM (ThermoScientific) containing 10% FBS and 1% penicillin–streptomycin. Cells were synchronized with 200 nM dexamethasone, diluted in serum-free medium for an hour and washed twice with the same solution (ThermoScientific) before reconstituting with serum-free medium containing 1× B27 (ThermoScientific) and 400 μM luciferin (Gold Biotechnology, St. Louis, Missouri, USA). Bioluminescence was recorded for 4 days in Tecan M200 Pro readers. Actimetrics MultiCycle was used to determine the period and amplitude (baseline subtracted 24 h running average of the raw luminescence data smoothed over 8 h). For luminometric experiments involving the 62 fibroblast cell lines, three technical replicates per dose, per drug, were conducted once and averaged.

Sanghani H.R., Jagannath A., Humberstone T., Ebrahimjee F., Thomas J.M., Churchill G.C., Cipriani A., Attenburrow M.J., Perestenko O.V., Cowley S.A., Cader M.Z., Peirson S.N., Harrison P.J., Foster R.G., Goodwin G.M, & Vasudevan S.R. (2020). Patient fibroblast circadian rhythms predict lithium sensitivity in bipolar disorder. Molecular Psychiatry, 26(9), 5252-5265.

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Circadian Rhythm Profiling in Fibroblasts

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Fibroblasts were cultured to confluence (~30 000 cells/well) in 96-well white plates (Greiner Bio-One, Gloucestershire, UK) using ADMEM (ThermoScientific) containing 10% FBS and 1% penicillin-streptomycin. Cells were synchronized with 200nM dexamethasone, diluted in serum-free medium for an hour and washed twice with the same solution (ThermoScientific) before reconstituting with serum-free medium containing 1x B27 (ThermoScientific) and 400μM luciferin (Gold Biotechnology, St. Louis, Missouri, USA). Bioluminescence was recorded for 4 days in Tecan M200 Pro readers. Actimetrics MultiCycle was used to determine the period and amplitude (baseline subtracted 24h running average of the raw luminescence data smoothed over 8h). For luminometric experiments involving the 62 fibroblast cell lines, three technical replicates per dose, per drug, were conducted once and averaged.

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Quantifying Mycobacterial Redox State

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A total of 2 × 10⁵ adipocytes and preadipocytes were seeded in a 96-well black plate and were infected at an MOI of 1 for 24 h with a plasmid expressing pMrx1-roGFP2. Ten days postinfection, the infected cells were washed once in PBS, and a fluorescence reading for emission at 520 nm was taken by excitation at 390 nm and 490 nm. For oxidizing or reducing, the cells were treated with 10 mM cumene hydroperoxide (CHP) or 40 mM DTT for 15 min, and then a fluorescence reading was taken using a Tecan M200 Pro reader. A log-phase culture grown in 7H9 complete medium in vitro was taken as the control; it was washed once in PBS, and after resuspension of the pellet in PBS, 200 μl was transferred to the black plate in triplicate for fluorescence reading. Bacterial samples were either left untreated or treated with 10 mM CHP or 40 mM DTT for full oxidization or reduction in order to calculate the E_MSH value for this in vitro-grown culture or for Mtb^A and Mtb^P as described in reference 32 (link).

Nandy A., Mondal A.K., Pandey R., Arumugam P., Dawa S., Jaisinghani N., Rao V., Dash D, & Gandotra S. (2018). Adipocyte Model of Mycobacterium tuberculosis Infection Reveals Differential Availability of Iron to Bacilli in the Lipid-Rich Caseous Environment. Infection and Immunity, 86(6), e00041-18.

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UV-Vis and Fluorescence Spectroscopy

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UV–Vis spectra were recorded on a spectrophotometer Agilent Cary 60 UV–Vis (Santa-Clara, CA, USA) in MeOH in the wavelength range from 200 to 800 nm. Fluorescence spectra were obtained from 50 μL of 500 μM stocks of compounds in anhydrous MeOH in black 384-well plates in Tecan M200Pro reader. Excitation wavelength was 350 nm (9 nm bandpass), and emission range was from 400 to 850 nm (20 nm bandpass) with the 2 nm step.

Klimenko A., Huber R., Marcourt L., Tabakaev D., Koval A., Dautov S.S., Dautova T.N., Wolfender J.L., Thew R., Khotimchenko Y., Queiroz E.F, & Katanaev V.L. (2023). Shallow- and Deep-Water Ophiura Species Produce a Panel of Chlorin Compounds with Potent Photodynamic Anticancer Activities. Antioxidants, 12(2), 386.

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IFN-γ ELISA Quantification Protocol

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An anti-IFN-γ monoclonal antibody has been used as primary antibody (Clone 2G1, Invitrogen, M700A) which was coated on the MaxiSorp 96-well plate (Nunc, 442404) overnight at 4 °C. Antibody was removed, and plates were blocked by PBS plus 1 % BSA for 2 h. Cells were co-cultured as previously mentioned, and supernatants were transferred into pre-coated plates for incubation. A biotin labeled anti-IFN-γ antibody has been used as a secondary antibody (Clone B133.5, M701B, Invitrogen), and HRP conjugated streptavidin (Thermo Scientific, N200) was added after secondary antibody incubation. 100 μL/well TMB (Sigma Aldrich, T5525) has been added as substrate of HRP. Plates were read by infinite M200 PRO reader (TECAN Austria GmbH, Grödlg, Austria). OD₄₅₀ was used for the readout and OD₆₃₀ was used as reference readout for background control. Technical replicates have been performed for ELISA assay.

Ma Y., Liu F., Li B., Peng K., Zhou H., Xu Y., Qiao D., Deng L., Tian G., Nielsen M, & Wang M. (2022). Identification and assessment of TCR-T cells targeting an epitope conserved in SARS-CoV-2 variants for the treatment of COVID-19. International Immunopharmacology, 112, 109283.

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Measuring Cell Metabolic Activity

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Metabolic activity was measured using the CellTiter 96 AQueous One Solution Cell Proliferation Assay (Promega, Madison, WI, USA) according to the manufacturer’s instructions. Briefly, 77 µL of cell-specific media was added to 33 µL of cell suspension. After transferring the mixture into a 96-well-plate, 20 µL of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) stain was added before incubating the plate for 4 h. Afterwards, the absorption of the plate at 490 nm was read in a M200 PRO reader (Tecan, Männedorf, Switzerland) and measured relative to a medium control. Afterwards, results were analyzed and displayed relative to a 37 °C control for each treatment condition. A medium control without cells was subtracted from each absorbance value measured before analysis.

Hahn O., Heining F.M., Janzen J., Becker J.C., Bertlich M., Thelen P., Mansour J.J., Duensing S., Pahernik S., Trojan L, & Popeneciu I.V. (2020). Modulating the Heat Sensitivity of Prostate Cancer Cell Lines In Vitro: A New Impact for Focal Therapies. Biomedicines, 8(12), 585.

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Kinase Assay for Aurora-A Activity

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AURKA kinase assays were performed using the ADP-Glo Kinase assay (Promega) according to the manufacturer protocol. Measurement of Aurora-A activity was carried out using reactions containing (per well):10 ng Aurora-A catalytic domain, 50 uM ATP, and 0.5 myelin basic protein (MBP), in 5% dimethyl sulfoxide (DMSO). Reactions were incubated at room temperature for 60 min, and luminescence data (integration time: 0.5–1 s) was recorded on a Tecan M200 Pro reader (Männedorf, Switzerland).

Ton A.T., Singh K., Morin H., Ban F., Leblanc E., Lee J., Lallous N, & Cherkasov A. (2020). Dual-Inhibitors of N-Myc and AURKA as Potential Therapy for Neuroendocrine Prostate Cancer. International Journal of Molecular Sciences, 21(21), 8277.

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Quantifying Osteogenic Gene Expression

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RNA was isolated from four samples per group in day 0, 7, 14 and 21 using the RNeasy Mini Kit (Qiagen, Hilden, Germany) followed the manufacturer's instructor. RNA content (ng per sample) and ratio of absorbance at 260 nm to 280 nm, indicating RNA purity, was measured using Tecan Infinite® M200 Pro reader. Presence of the RNA in the samples was checked using standard agarose gel electrophoresis. The isolated RNA was processed followed manufacturers instruction. Briefly, osteogenic markers osteocalcin (OCN), collagen type I (COL1A2), integrin binding sialoprotein (ISBP), and EE1 (eukaryotic elongation factor, was used as housekeeping gene) genes were analysed. Detail description of the analysed genes in ESI Table S1.† For the reactions RevertAid H Minus First Strand cDNA Synthesis Kit (Thermo Scientific, Waltham, MA), TaqMan probes with a fluorescent label at the 5′ end and the quencher on the 3′ end (reviewed in ESI†), TaqMan Gene Expression Master Mix (Thermo Scientific, Waltham, MA, USA) and RT-PCR Grade Water (Thermo Scientific, Waltham, MA) was used. Results were generated from the values obtained by calculation according to the formula 2^−ΔC_p. Light Cycler 480 (Roche, Basel, Switzerland) was used to measure the fluorescence intensity.

Jarolimova P., Voltrova B., Blahnova V., Sovkova V., Pruchova E., Hybasek V., Fojt J, & Filova E. (2020). Mesenchymal stem cell interaction with Ti6Al4V alloy pre-exposed to simulated body fluid. RSC Advances, 10(12), 6858-6872.

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Monitoring Cellular ROS Levels

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The impact of caffeine and 4-HNE on ROS generation was measured using dihydroethidium probe (DHE, Molecular Probes). The fluorescence was continuously measured every 15 s for 40 min, and then as a single point after 24 h using Tecan M200 Pro reader (Ex 530 nm/Em 580 nm).
Additionally, the impact of 4-HNE and caffeine on intracellular ROS production was measured using 2′,7′-dichlorfluorescein-diacetate (DCFH-DA, Fluka) assay as previously described [11, 31] . Cells were loaded with 10 μM DCFH-DA for 30 min at 37 °C. The excess probe has been removed, treatment added, and fluorescence intensity continuously measured throughout 8 h at 37 °C and 5% CO 2 using TECAN Infinite M200 PRO plate reader equipped with gas control mode, excitation wavelength at 500 nm and emission at 529 nm. The arbitrary units, relative fluorescence units (RFU), were based directly on fluorescence intensity.

Al-Menhali A.S., Banu S., Angelova P.R., Barcaru A., Horvatovich P., Abramov A.Y, & Jaganjac M. (2020). Lipid peroxidation is involved in calcium dependent upregulation of mitochondrial metabolism in skeletal muscle. Biochimica et biophysica acta. General subjects, 1864(3).

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