Clariostar plus microplate reader

Manufactured by BMG Labtech

Sourced in Germany, United States, Australia

The CLARIOstar Plus is a high-performance microplate reader engineered by BMG LABTECH. It is a versatile instrument capable of measuring absorbance, fluorescence, and luminescence in microplates.

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Lab products found in correlation

Pierce bca protein assay kit, by Thermo Fisher Scientific (4 mentions) Nano glo luciferase assay reagent, by Promega (3 mentions) Prism 9, by GraphPad (3 mentions) Fitc dextran, by Merck Group (2 mentions) Fitc dextran, by BMG Labtech (2 mentions) Celltiter glo luminescent cell viability assay, by Promega (2 mentions) Lumitrac 96 well plate, by Greiner (2 mentions) Anti human igg hrp, by Jackson ImmunoResearch (2 mentions) Tmb substrate solution, by Thermo Fisher Scientific (2 mentions) Celltiter 96 aqueous one solution cell proliferation assay, by Promega (2 mentions) Fastap alkaline phosphatase, by Thermo Fisher Scientific (2 mentions) Nad p h quinone oxidoreductase 1 nqo1, by R&D Systems (2 mentions) Kp372 1, by Thermo Fisher Scientific (2 mentions) Fastap reaction buffer, by Thermo Fisher Scientific (2 mentions) Methanol, by Thermo Fisher Scientific (2 mentions) Dulbecco phosphate buffered saline (dpbs), by Thermo Fisher Scientific (2 mentions) Matrisperse cell recovery solution, by Corning (1 mentions) Axiovert inverted microscope, by Zeiss (1 mentions) Methyl thiazolyl tetrazolium (mtt), by Merck Group (1 mentions) Propidium iodide, by Merck Group (1 mentions)

103 protocols using clariostar plus microplate reader

Quantifying Promoter Activities in Microtiter Plates

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Promoter activities were assessed by measuring mVenus fluorescence intensity in 96-well microtiter plates (Nunc™). Each well contained 0.1 ml AMM inoculated with 10⁴ spores. Plates were incubated for 18 h at 37 °C. Subsequently, absorbance and fluorescence signals were quantified using a CLARIOstar Plus® microplate reader (BMG LABTECH). Absorbance was measured at 280 nm, spiral well scan. For detection of mVenus fluorescence, 497-20 nm excitation and 540-20 nm emission was used. Each reporter strain was analyzed in biological triplicates followed by subtraction of background fluorescence recorded from untransformed recipient strain (wt).
For measuring the bioluminescence of luciferase reporter strains, LUMITRAC 96-well plates (Greiner Bio-ONE) were used. Spores were inoculated in AMM to obtain a final concentration of 1.5 × 10⁴ spores in 0.1 ml. After incubation for 18 h at 37 °C, 20 μL of 0.6 mM D-luciferin (Synchem UG & Co.KG, Felsberg/Altenburg, Germany) in PBS was added. The bioluminescence was recorded at 580-80 nm using spiral well scan employing a CLARIOstar Plus® microplate reader (BMG LABTECH). Each reporter strain was analyzed in three biological triplicates followed by subtraction of background luminescence recorded from untransformed wt strain.

Yap A., Glarcher I., Misslinger M, & Haas H. (2022). Characterization and engineering of the xylose-inducible xylP promoter for use in mold fungal species. Metabolic Engineering Communications, 15, e00214.

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Amyloid Protein Aggregation Kinetics

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50 μM monomer solutions were aggregated in the presence of 20 μM Thioflavin T (ThT) and 0.02% NaN₃. 170 μl of each sample (5 replicates) was loaded into a 96 well full-area plate (non-binding, clear bottomed, #655906, Greiner Bio-One, Frickenhausen, Austria) and incubated at 37°C for ∼140 h in a CLARIOstar Plus microplate reader (BMG Labtech, Ortenberg, Germany). Aggregation was promoted through linear shaking (300 rpm, 300 s before each cycle) with the addition of a single borosilicate bead (3 mm diameter) to each well. Fluorescent intensity measurements were taken using spiral averaging (5 mm diameter) using excitation 440 nm, dichroic 460 nm, and emission 480 nm filters, 3 gains and 50 flashes per well. The data were plotted using GraphPad Prism version 9.3.1 for Windows (GraphPad Software, San Diego, CA, United States). The reported t_1/2 and t_lag values are averages of the independent replicates. The sigmoidal model provided by the software was used to estimate the t_1/2 of aggregation. In particular, the t_1/2 was estimated as the x-value at 50% of maximum fluorescence. The t_lag was estimated by extrapolation from the maximum aggregation rate.

Thrush R.J., Vadukul D.M, & Aprile F.A. (2022). A Facile Method to Produce N-Terminally Truncated α-Synuclein. Frontiers in Neuroscience, 16, 881480.

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Measurement of Maximum LDH Activity

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The maximum LDH activity was measured with the Pierce LDH Cytotoxicity Assay Kit (Cat. No. 88953; Thermo Fisher Scientific) according to manufacturer instructions. Briefly, 20,000 MEF were cultured overnight in a flat bottom, 96-well plate at 37 °C, 5% CO₂. Cells were lysed with lysis buffer provided in the kit and 50 μl of supernatant was transferred to a fresh 96-well flat bottom plate. The kit’s proprietary reaction solution was added to the supernatant and incubated at room temperature for 30 m. After stopping the reaction, absorbance was measured at 490 nm and 680 nm in a CLARIOstar Plus microplate reader (BMG LABTECH). LDH activity was determined by subtracting the 680 nm from the 490 nm absorbance values.

Solano Fonseca R., Metang P., Egge N., Liu Y., Zuurbier K.R., Sivaprakasam K., Shirazi S., Chuah A., Arneaud S.L., Konopka G., Qian D, & Douglas P.M. (2021). Glycolytic preconditioning in astrocytes mitigates trauma-induced neurodegeneration. eLife, 10, e69438.

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Oral FITC-Dextran Absorption Assay

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Mice were starved over-night and then orally administered through gavage with $400 mg/Kg FITC-Dextran (4 kDa; Sigma-Aldrich, # 46944-500MG-F). Blood was collected from the heart after four hours and the concentration of FITC-Dextran in plasma samples was measured as fluorescence intensity (Clariostar Plus Microplate Reader; BMG Labtech).

Bertocchi A., Carloni S., Ravenda P.S., Bertalot G., Spadoni I., Lo Cascio A., Gandini S., Lizier M., Braga D., Asnicar F., Segata N., Klaver C., Brescia P., Rossi E., Anselmo A., Guglietta S., Maroli A., Spaggiari P., Tarazona N., Cervantes A., Marsoni S., Lazzari L., Jodice M.G., Luise C., Erreni M., Pece S., Di Fiore P.P., Viale G., Spinelli A., Pozzi C., Penna G, & Rescigno M. (2021). Gut vascular barrier impairment leads to intestinal bacteria dissemination and colorectal cancer metastasis to liver. Cancer cell, 39(5).

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Organoid Harvesting, Viability Assays

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For the harvesting, mouse and human organoids were collected and centrifuged at 200 RCF for 5 minutes at 4 C. The Matrigel was removed by incubation in Matrisperse Cell Recovery Solution (Corning) for 20 minutes on ice and the organoids were then pelleted by centrifugation. For immunolabeling, the organoid pellet was suspended in 2% PFA and fixed overnight at 4 C. For molecular analyses, the pellet was frozen at À80 C followed by RNA and protein isolation.
MTT assay was performed in organoids seeded in 96-well plates. MTT (500 mg/mL, Sigma-Aldrich) was added to 70 mL of culture medium and incubated for 1 hour at 37 C. After incubation the medium was discarded and the Matrigel solubilized by adding 20 mL of 20% SDS. After 1-hour incubation at 37 C, 100 mL of DMSO (Sigma) were added to each well and the plate was incubated for 90 minutes at 37 C in the dark. Absorbance measurement at 560 nm was performed in a CLARIOstar Plus Microplate reader (BMG Labtech).
Assessment of organoid viability by live imaging was performed by adding Hoechst 33342 (50 mg/mL) and propidium iodide (PI; 1 mg/mL, SIGMA) to the culture medium. After 10-minute incubation, the analysis of fluorescence was performed on a Zeiss AxioVert inverted microscope.

Frau C., Jamard C., Delpouve G., Guardia G.D.A., Machon C., Pilati C., Nevé C.L., Laurent-Puig P., Guitton J., Galante P.A.F., Penalva L.O., Freund J.N., de la Fouchardiere C, & Plateroti M. (2021). Deciphering the Role of Intestinal Crypt Cell Populations in Resistance to Chemotherapy. Cancer research, 81(10).

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Dexamethasone Induces Apoptosis in LSECs

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LSECs, 0.1×10⁶ per well, were seeded in 96 well plates (Corning Costar® Cat. No 3610). Cells were washed after 40 min, then incubated with 0.1, 1, or 10 μg/ml of Dex, or without Dex for 24 h. The positive control was established by replacing the medium in non-treated cultures at 20 h with medium containing 100 ng/ml of recombinant human Fas Ligand protein (rhsFasL, Abcam, Cat. No ab157085), then incubation was continued for 4h. At 24h, Caspase-Glo® 3/7 Assay reagents (Promega Corporation, Cat. No G8090) were added to all wells and results read by CLARIOstar Plus microplate reader (BMG Labtech GMbH, Ortenberg, Germany) after 1 h. The experiment was done in triplicate and repeated in 4 biological replicates.

Li R., Bhandari S., Martinez-Zubiaurre I., Bruun J.A., Urbarova I., Smedsrød B., Simón-Santamaría J, & Sørensen K.K. (2022). Changes in the proteome and secretome of rat liver sinusoidal endothelial cells during early primary culture and effects of dexamethasone. PLoS ONE, 17(9), e0273843.

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Quantifying Sit1-Venus Levels During Germination

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To determine the level of Venus-tagged Sit1 protein during germination, the previously described strain Sit1^C-Venus (Sit1 tagged C-terminally with the yellow fluorescence protein derivative Venus with expression of the tagged sit1 alleles under the control of the endogenous promoter) (10 (link), 24 (link)) was used. The protein level of sit1^C-Venus during germination was measured by inoculating 200 μL of AMM with 5 × 10⁶ conidia and incubating at 37°C for 10 h in a 96-well plate (catalog no. 167008; Nunc MicroWell 96 well; Thermo Fisher Scientific Inc., Waltham, MA, USA). For iron-deplete conditions, iron was omitted, and for iron-replete conditions, FeSO₄ was added to a final concentration of 0.03 mM. After incubation, fluorescence was measured at 497 ± 15 nm (excitation, 497 ± 15 nm; dichroic mirror, auto, 517.2 nm; emission, 540 ± 20 nm; top excitation and detection) using CLARIOstar Plus microplate reader (BMG Labtech GmbH, Ortenberg, Germany). To visualize the increase of Venus-mediated fluorescence during germination, the unspecific WT background fluorescence of the respective time points, as well as the fluorescence of the 0-h time point, was subtracted from each time point. Three biological triplicates of each reporter strain were analyzed. The mean value and standard deviation were then calculated and displayed.

Happacher I., Aguiar M., Alilou M., Abt B., Baltussen T.J., Decristoforo C., Melchers W.J, & Haas H. (2023). The Siderophore Ferricrocin Mediates Iron Acquisition in Aspergillus fumigatus. Microbiology Spectrum, 11(3), e00496-23.

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Intracellular ATP Quantification Assay

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Intracellular ATP levels were measured using the CellTiter-Glo^® Luminescent Cell Viability Assay (PROMG7571, Promega (Madison, WI, USA) according to manufacturer instructions. Briefly, cells were seeded and treated with MitoBENs, as previously described in the Section 4.3, using white opaque 96-well plates (Costar-reference CORN3917). At the end of the compounds’ incubation period the growth media was replaced by 40 μL of fresh room temperature equilibrated growth media, and 40 μL of CellTiter-Glo^® reagent mix was added to each well to promote cell lysis and the luciferase-catalysed oxidation of luciferin to oxyluciferin. The reaction occurred for 10 min. The luminescent signal generated is proportional to the amount of ATP and measured using a CLARIOstar Plus microplate reader (BMG Labtech, Ortenberg, Germany).

Fernandes C., Cagide F., Simões J., Pita C., Pereira E., Videira A.J., Soares P., Duarte J.F., Santos A.M., Oliveira P.J., Borges F, & Silva F.S. (2022). Targeting Hydroxybenzoic Acids to Mitochondria as a Strategy to Delay Skin Ageing: An In Vitro Approach. Molecules, 27(19), 6183.

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Quantifying Exo-PG Activity with DNS Assay

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The exo-PG activity was measured by quantifying reducing groups expressed as GalA units according to the method described by Miller [21] (link). A volume of 0.5 mL of enzyme solution (0.5 μg/mL TMA01 and μg/mL BLI04) was added to 0.5 mL of 0.5% (w/v) polygalacturonic acid in 20 mM sodium phosphate buffer at pH 6.5. After 15 min of incubation at 50 • C (Thermomixer™ C, Eppendorf, UK), 0.5 mL of 3,5-Dinitrosalicylic acid (DNS) reagent were added. The mixture was heated at 100 • C for 15 min and then centrifuged at 10,000g for 5 min. The reducing groups were quantified in the supernatant at 540 nm (CLAR-IOstar Plus Microplate Reader, BMG LabTech, Germany) in 96-well plates. A blank reaction was carried out following the procedure describe above using only buffer instead of enzyme solution. The molar extinction coefficient (ε) of 0.1266 mM -1 cm -1 was calculated using GalA as standard (SM Fig. A.2). One unit (U) of exo-PG activity was defined as the amount of enzyme that releases 1 μmol of GalA equivalent per min at pH 6.5 and 50 • C.

Flores-Fernández C.N., Cárdenas-Fernández M., Lye G.J, & Ward J.M. (2022). Synergistic action of thermophilic pectinases for pectin bioconversion into D-galacturonic acid. Enzyme and microbial technology, 160.

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Glutathione Reductase Activity Assay

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The GR activity assay was based on the reduction of GSSG to GSH in the presence of NADPH. GSH then reacts with 5,5′‐dithiobis‐(2‐nitrobenzoic acid) (DTNB) to generate 5‐thio‐(2‐nitrobenzoic acid) (TNB), which is associated with an increase in the absorbance at 412 nm; the rate of this increase is directly proportional to the GR activity in the sample. The extinction coefficient (ε) for TNB is 14.15 mM⁻¹ cm⁻¹. Briefly, the assay was performed with 1 mM GSSG (Sigma), 0.05 mM NADPH (Sigma), 1.5 mM DTNB, and zebrafish lysate (after using the Bio‐Rad Bio‐Spin 6 to remove endogenous GSH) or with GR (Sigma) as a positive control in 0.1 M potassium phosphate buffer with 1 mM ethylene diamine tetraacetic acid, pH 7.5 (final volume, 200 µL). Absorbance was read at 412 nm with the CLARIOstar Plus microplate reader (BMG LABTECH).

Park K., Gooz M., Ye Z., Zhang J., Beeson G.C., Rockey D.C, & Kim S. (2021). Flavin Adenine Dinucleotide Depletion Caused by electron transfer flavoprotein subunit alpha Haploinsufficiency Leads to Hepatic Steatosis and Injury in Zebrafish. Hepatology Communications, 5(6), 976-991.

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