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What is the benefit of using the TurboLuc Luciferase One-Step Glow Assay Kit over the Luciferase Glow Assay kits? Product FAQ

Answer

TurboLuc Luciferase One-Step Glow Assay Kit measures activity in mammalian cells with a single reagent-addition step, making it ideal for high-throughput screening (HTS) applications. Several features of the TurboLuc Luciferase One-Step Glow Assay Kit make it suitable for luminometers without injectors and for HTS applications. The convenient, one-step, homogenous protocol minimizes handling steps to support the use of automation. Simply combine the supplied substrate solution and assay buffer to make a single working solution, and then add it to microplate containing the transfected, treated cells. The stable signal output (glow enzyme kinetics) of this system provides flexibility with regard to lead time before read. When used with Thermo Scientific TurboLuc 16 Luciferase vectors, the TurboLuc Luciferase One-Step Glow Assay Kit provides a highly sensitive bioluminescent reporter assay system for the detection of promoter or pathway activity.

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Answer Id: E15650

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What are the advantages of one mode of detection over another for protein and enzyme activity assays? Product FAQ

Answer

The first consideration in using a specific mode of detection is availability. What modes of detection are available for an assay? Some assays are not available in one mode or another due to the chemistry of the reaction or availability of reagents for the reaction.
Secondly, does the experimental sample have color (absorbance) or autofluorescence? Some natural materials are naturally colored or fluorescent. Check your samples to see if they have a strong absorbance or absorbance over a broad range of wavelengths. This may limit your ability to use a colorimetric assay. If you are interested in a fluorescence-based assay, examine your samples, media, etc. for autofluorescence (fluorescence that is inherent to the sample due to various natural components) using all filters sets or channels available. Autofluorescent components include fluorescent proteins (GFP, etc.), porphyrins, dyes in media, some vitamins, natural pigments, etc. If a sample has high autofluorescence in a broad range of wavelengths, a fluorescence-based assay may not be compatible.
Time-resolved FRET (TR-FRET) and luminescence-based assays are desirable because endogenous sample absorbance and autofluorescence can be avoided in the detected signal. TR-FRET involves exciting the sample with defined wavelengths, turning the excitation light off and then reading emission within 50 to 100 µsec after the light has been turned off.
Luminescence-based assays, either bioluminescent or chemiluminescent assays, do not require that a sample be exposed to any excitation light; light is generated by the biological or chemical modification of the substrate.
Ratiometric assays reduce the effects of well-to-well and day-to-day variations.

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Answer Id: E15830

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Protein-Based Ca2+ Indicators—Section 19.5 Molecular Probes Handbook

Detecting Enzymes That Metabolize Phosphates and Polyphosphates—Section 10.3 Molecular Probes Handbook

Viability and Cytotoxicity Assay Kits for Diverse Cell Types—Section 15.3 Molecular Probes Handbook

Firefly and bacterial luminescence: basic science and applications. Citations & References

  • Authors: McElroy WD, DeLuca MA
  • Journal: J Appl Biochem
  • PubMed ID: 6680120
Catalog #

Viability and Cytotoxicity Assay Reagents—Section 15.2 Molecular Probes Handbook

Substrates for Microsomal Dealkylases, Acetyltransferases, Luciferases and Other Enzymes—Section 10.6 Molecular Probes Handbook

Lysophosphatidic acid-induced membrane ruffling and brain-derived neurotrophic factor gene expression are mediated by ATP release in primary microglia Citations & References

  • Authors: Fujita, R; Ma, Y; Ueda, H
  • Journal: JOURNAL OF NEUROCHEMISTRY

Thiol-Reactive Probes Excited with Visible Light—Section 2.2 Molecular Probes Handbook

Fluorescence Fundamentals Molecular Probes Handbook

Recombinant Gaussia luciferase. Overexpression, purification, and analytical application of a bioluminescent reporter for DNA hybridization. Citations & References

  • Authors: Verhaegent M, Christopoulos TK
  • Journal: Anal Chem
  • PubMed ID: 12236345
Catalog #

Expression hybridization assays combining cDNAs from firefly and Renilla luciferases as labels for simultaneous determination of two target sequences. Citations & References

  • Authors: Laios E, Obeid PJ, Ioannou PC, Christopoulos TK
  • Journal: Anal Chem
  • PubMed ID: 10994960
Catalog #

A review of bioluminescent ATP techniques in rapid microbiology. Citations & References

  • Authors: Stanley PE
  • Journal: J Biolumin Chemilumin
  • PubMed ID: 2678922
Catalog #

An enzymatic cycling method using pyruvate orthophosphate dikinase and firefly luciferase for the simultaneous determination of ATP and AMP (RNA). Citations & References

  • Authors: Sakakibara T, Murakami S, Eisaki N, Nakajima M, Imai K
  • Journal: Anal Biochem
  • PubMed ID: 10036167
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