Spectramax id5 microplate reader

Manufactured by Molecular Devices

Sourced in United States

The SpectraMax iD5 microplate reader is a flexible, high-performance instrument designed for a wide range of absorbance-based applications in life science research and drug discovery. It features a monochromator-based optical system that can measure absorbance across a wavelength range of 190 to 850 nm.

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Lab products found in correlation

Dcf da probe, by Abcam (2 mentions) Heparanase, by R&D Systems (1 mentions) Streptavidin xlent, by PerkinElmer (1 mentions) Biotin heparan sulfate eu cryptate, by PerkinElmer (1 mentions) White 96 well plate, by SPL Life Sciences (1 mentions) Renilla luciferase lysis buffer, by Promega (1 mentions) Te2000 e, by Nikon (1 mentions) Ace2 sars cov 2 spike inhibitor screening assay kit, by BPS Biosciences (1 mentions) Lasx 3, by Leica (1 mentions) Dm il led, by Leica (1 mentions) Dfc3000g ccd camera, by Leica (1 mentions) Epigenase hdac activity inhibition direct assay kit, by Epigentek (1 mentions) Protein extraction kit, by BestBio (1 mentions) Ub amc, by R&D Systems (1 mentions) Dp72 ckx41, by Olympus (1 mentions) Heat inactivated fetal bovine serum, by Thermo Fisher Scientific (1 mentions) Dmem medium, by Thermo Fisher Scientific (1 mentions) Cls3824, by Corning (1 mentions) Mtase glo detection solution, by Promega (1 mentions) Tmb substrate solution, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

Microplate reader (2507 citations) SpectraMax iD5 (292 citations) SpectraMax microplate reader (255 citations) SpectraMax iD5 Multi-Mode Microplate Reader (59 citations) SpectraMax iD5 microplate (3 citations) SpectraMax iD5 reader (2 citations) Nanodrop Spectrometer (Spectramax iD5 multi-mode microplate reader; (2 citations) Multimode Microplate Reader Spectramax iD5 (2 citations)

40 protocols using spectramax id5 microplate reader

3WJ-Induced Transcription Amplification

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The 3WJ-induced transcription amplification reaction was performed in two steps: DNA amplification followed by transcription amplification. For DNA amplification, solution A containing 4 μL of 250 nM 3WJ_template, 8 μL of 250 nM 3WJ_primer, 1 μL of 10 mM (each 2.5 mM) deoxyribonucleotide triphosphates (dNTPs), 10 μL of 10 mM (each 2.5 mM) nucleotide triphosphates (NTPs), and 4 μL of 10× Cutsmart^® buffer was incubated with the target DNA at different concentrations for 5 min at 37 °C. To this mixture, solution B containing 0.4 μL of 10 U/μL Klenow DNA polymerase (exo-) and 0.8 μL of 10 U/μL Nt.AlwI was added and incubated for 1 h at 37 °C. For transcription amplification, 1.2 μL of 50 U/μL T7 RNA polymerase was added to a total reaction volume of 40 μL and incubated for 1 h at 37 °C. Finally, 10 μL DFHBI (50 μM) was added to interact with the light-up RNA aptamer and generate a fluorescence signal, which was measured at an excitation wavelength of 452 nm and an emission wavelength of 506 nm using a SpectraMax iD5 microplate reader (Molecular Devices, Sunnyvale, CA, USA).

Kim J.H., Kim S., Hwang S.H., Yoon T.H., Park J.S., Lee E.S., Woo J, & Park K.S. (2021). Three-Way Junction-Induced Isothermal Amplification with High Signal-to-Background Ratio for Detection of Pathogenic Bacteria. Sensors (Basel, Switzerland), 21(12), 4132.

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Heparanase Activity Assay Protocol

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42 μL of HS trisaccharide solution in Milli-Q water (0.0038–500 μM) or just Milli-Q water (as a control), and 42 μL of heparanase (5.3 nM, R&D Systems) solution in pH 7.5 triz buffer (consisting of 20 mM TrisHCl, 0.15 M NaCl and 0.1% CHAPS) or just buffer as blank were added into microtubes and pre-incubated at 37 °C for 10 min bringing the [heparanase] to 0.5 nM. Next, 84 μL of biotin-heparan sulfate-Eu cryptate (Cisbio, Cat #: 61BHSKAA) (58.6 ng in pH 5.5 0.2 M NaOAc buffer) was added to the microtubes, and the resulting mixture was incubated for 60 min at 37 °C. The reaction mixture was stopped by adding 168 μL of Streptavidin-XLent! (Cisbio, Cat #: 611SAXLA) (1.0 μg/ml) solution in pH 7.5 dilution buffer made of 0.1 M NaH₂PO₄, 0.8 M KF, 0.1% BSA. After the mixture had been stirring at room temperature for 15 min, 100 μL (per well) of the reaction mixture was transferred to a 96 well microplate (Corning #3693 96 well, white polystyrene, half-area) in triplicates and HTRF emissions at 616 nm and 665 nm were measured by exciting at 340 nm using a SpectraMax iD5 Microplate Reader (Molecular Devices).

Zhu S., Li J., Loka R.S., Song Z., Vlodavsky I., Zhang K, & Nguyen H.M. (2020). Modulating Heparanase Activity: Tuning Sulfation Pattern and Glycosidic Linkage of Oligosaccharides. Journal of medicinal chemistry, 63(8), 4227-4255.

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Luciferase Activity Quantification in EV and Cell Lysates

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For the luciferase assay, HEK-293 cells and COS-7 cells were transfected for luciferase assay with expression vectors. The following protocols measured the luciferase activity from isolation of EVs and cell lysates. According to manufacturer’s instructions, the samples were resuspended with 150 μL Renilla luciferase lysis buffer (Promega). The 20 μL total sample volume was dissolved in 100 μL luciferase assay reagents containing substrate. Immediately, the sample was loaded into a white 96 well plate (SPL Life sciences). Measurements were performed using SpectraMax iD5 Microplate Reader (Molecular devices, Silicon Valley, CA, USA).

Kim R, & Kim J.H. (2023). Engineered Extracellular Vesicles with Compound-Induced Cargo Delivery to Solid Tumors. International Journal of Molecular Sciences, 24(11), 9368.

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Anoikis Assay in Prostate Cancer Cells

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Anoikis was measured with a CytoSelect 96-Well Anoikis Assay Kit (Cell Biolabs, Inc., San Diego, CA, USA) using MTT colorimetric and Calcein AM/EthD-1 fluorometric detection according to manufacturer’s instructions. Briefly, NC or miR-214 mimic transfected and wild-type or miR-214KO PCa cells (2 × 10⁴ cells/well) were seeded in a Poly-HEMA coated anchorage-resistant 96-well plate. After culturing for 48 h, MTT reagent was added to each well for 4 h at 37 °C, then detergent was added. The plate was incubated for another 4 h in the dark at room temperature, 150 μL was transferred to the 96-well plate, and absorbance was measured at 570 nm on the Fluostar Omega plate reader. Calcein AM (C-AM) and ethidium homodimer-1 (EthD-1) fluorescence was also detected following the manufacturer’s protocol, monitored under a Nikon TE-2000-E fluorescence microscope, and quantitatively measured with a Spectramax iD5 microplate reader (Molecular Devices) (C-AM, Ex: 485 nm and Em: 515 nm).

Cagle P., Smith N., Adekoya T.O., Li Y., Kim S., Rios-Colon L., Deep G., Niture S., Albanese C., Suy S., Collins S.P, & Kumar D. (2021). Knockdown of microRNA-214-3p Promotes Tumor Growth and Epithelial-Mesenchymal Transition in Prostate Cancer. Cancers, 13(23), 5875.

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SARS-CoV-2 Spike Inhibitor Screening Assay

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The ACE2: SARS-CoV-2 Spike Inhibitor Screening Assay Kit (Cat: 79936) was purchased from BPS Biosciences (San Diego, US). Briefly, Fc-RBD was added to ACE2-His-coated test wells in the presence of 25–200 μM compounds, and to negative control wells containing no compound. Blank wells were left compound-free and mFc-RBD-free. Anti-Fc-horseradish peroxidase substrate was added to each well followed by mixed ELISA ECL Substrate A and B. Chemiluminescence was read with a SpectraMax iD5 Microplate Reader (Molecular Devices, San Jose, US).

Zhang D., Hamdoun S., Chen R., Yang L., Ip C.K., Qu Y., Li R., Jiang H., Yang Z., Chung S.K., Liu L, & Wong V.K. (2021). Identification of natural compounds as SARS-CoV-2 entry inhibitors by molecular docking-based virtual screening with bio-layer interferometry. Pharmacological Research, 172, 105820.

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Heparin-Antithrombin III Inhibition of Factor Xa

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All the reagents for this assay were reconstituted and prepared according to the manufacturer’s instructions and incubated at 37 °C for 15 min. Different concentrations of heparin (0.002‐500 nM; 40 μL) or trisaccharide 1α (3.9‐4000 nM; 40 μL) and ATIII (0.04 IU; 40 μL) were added to a deep‐well block (Nunc 96 DeepWell 1.0 mL/well, clear), mixed, and incubated at 37 °C for 2 min. To the reaction mixture, FXa (0.32 μg; 40 μL) was added by multichannel pipette and was incubated at 37 °C for another 2 min (stage 1), then FXa specific chromogenic substrate (0.048 mmol; 40 μL) was added. The reaction was stopped by adding citric acid (240 μL; 20 g/L) exactly after 2 min. A 100 μL solution was then transferred to a clear 96‐well microplate in triplicate, and absorbance at 405 nm was measured with a SpectraMax iD5 Microplate Reader (Molecular Devices). The sample blank was measured by mixing the reagents in reverse order from that of the test, i.e. citric acid, FXa substrate, FXa, ATIII, and sample. The sample blank value was deducted from the absorbance measured for the corresponding assay.

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Heparin and Trisaccharide 1α Inhibition of Thrombin

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All the reagents for this assay were reconstituted and prepared according to the manufacturer’s instructions and incubated at 37 °C for 15 min. Different concentrations of heparin (0.002‐500 nM; 40 μL) or trisaccharide 1α (3.9‐4000 nM; 40 μL) and ATIII (0.01 IU; 40 μL) were added to a deep‐well block (Nunc 96 DeepWell 1.0 mL/well, clear), mixed, and incubated at 37 °C for 2 min. To the reaction mixture, FIIa (1.2 nkat; 40 μL) was added by multichannel pipette and was incubated at 37 °C for another 2 min (stage 1), then FIIa specific chromogenic substrate (0.048 mmol; 40 μL) was added. The reaction was stopped by adding citric acid (240 μL; 20 g/L) exactly after 2min. 100 μL was transferred to a 96‐well microplate in triplicate, and absorbance at 405 nm was measured with SpectraMax iD5 Microplate Reader (Molecular Devices). The sample blank was measured by mixing the reagents in reverse order from that of the test, i.e. citric acid, FXa substrate, FXa, ATIII, and sample. The sample blank value was deducted from the absorbance measured for the corresponding assay.

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Titration ELISA for Measuring scFv Binding to GLP-1R

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Titration ELISA was used to measure binding of purified scFv to GLP-1R. GLP-1R immobilization in a 96-well plate and blocking was carried out as described for phage ELISA. A purified scFv was prepared at varying concentrations (from 1 nM to 3 μM) in 2% MPBS. 100 μl scFv aliquot of each concentration was added to a well and incubated at room temperature for 1.5 h. The plate was then washed with TPBS and PBS three times each. Primary anti His-tag Mouse mAb (Cell signaling Technology, Inc.) was diluted 1:250 in 2% MPBS and added into each well. After 1.5 h incubation, wells were washed and secondary anti-mouse (goat-anti mouse-HRP; Santa Cruz Biotechnology) diluted 1:1000 in 2% MPBS was added and incubated for 1 h. After washing, TMB substrate solution was added for color development (15 to 30 min) and 2M sulfuric acid was added to stop the reaction. The absorbance was read at 450 nm, using SpectraMax iD5 microplate reader (Molecular Devices, LLC). The dissociation constant K_D was calculated according to a published protocol (35 (link)).

Jeong S.L., Zhang H., Yamaki S., Yang C., McKemy D.D., Lieber M.R., Pham P, & Goodman M.F. (2022). Immunoglobulin somatic hypermutation in a defined biochemical system recapitulates affinity maturation and permits antibody optimization. Nucleic Acids Research, 50(20), 11738-11754.

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Quantification of Monoclonal Antibodies and ScFv Fusion Proteins

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Cell culture medium was clarified by centrifugation. ETE mAb LC and
DTE mAb LC were quantified using Kappa and Lambda Human Immunoglobulin
Free LC ELISAs (BioVendor, UK) following the manufacturer’s
protocol. Both IgG1 mAbs and ScFv fusion protein titers were quantified
using ValitaTitre (ValitaCell, Ireland). ValitaTitre measurements
were done in accordance with the manufacturer’s protocol. The
commercially available purified kappa IgG1 mAb (Merck, Germany) and
lambda IgG1 mAb (Merck, Germany) were used for quantification of the
ETE IgG1 mAb and DTE IgG1 mAb, respectively. The purified ScFv fusion
protein (AstraZeneca, UK) was used for quantification of the ScFv
fusion protein. All assays were read using a SpectraMax iD5 microplate
reader (Molecular Devices, USA).

O’Neill P., Mistry R.K., Brown A.J, & James D.C. (2023). Protein-Specific Signal Peptides for Mammalian Vector Engineering. ACS Synthetic Biology, 12(8), 2339-2352.

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Quantifying Advanced Glycation End-Products

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For the measurements of MGO-induced AGEs accumulation, EpiDermFT skin models or HDF cultures were washed in phosphate-buffered saline (PBS) and the 6-well plates were transferred to a microplate reader (SpectraMax iD5 Microplate Reader, Molecular Devices). The intensities of the fluorescence signals were recorded at Ex 370 nm/Em 440 nm and Ex 335 nm/Em 385 nm on days 0, 1, 4 and 7 and the relative fluorescence signal (RFI) was normalized to control.
For the visualization of the auto-fluorescence, the samples were observed under a fluorescence microscope (Leica DM IL LED, DAPI channel) and photographed using CCD DFC3000G Camera and LAS X 3.6.0.20104 software (Leica Microsystems).

Markiewicz E., Jerome J., Mammone T, & Idowu O.C. (2022). Anti-Glycation and Anti-Aging Properties of Resveratrol Derivatives in the in-vitro 3D Models of Human Skin. Clinical, Cosmetic and Investigational Dermatology, 15, 911-927.

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