Luciferase enzymes were diluted in 1x passive lysis buffer from the assay kit tested. Promega luciferase assay systems an extremely sensitive reagent for rapid quantitation of firefly luciferase. The bioglonl luciferase assay system provides a highly sensitive, robust, homogeneous reagent for the detection of nanoluc luciferase reporter gene expression in promega bioassays. The system is a convenient alternative to firefly photinus pyralis reporter systems and is designed to yield reliable, linear results for a concentration range over 7 orders of magnitude. Dualluciferase reporter assay system technical manual pdf. Luminometer in combination with promega s dualglo luciferase assay kit provides a convenient, rapid, and sensitive procedure for quantifying gene expression.
For kits e1500, e1501, e4030, e4530 and e4550, prepare luciferase assay reagent lar by adding. Consecutive detection of firefly and renilla luciferase activities in a single sample. Luciferase assay system firefly luciferase and 100fold greater sensitivity than a published method for assaying renilla luciferase. The assay of this genetic reporter is rapid, sensitive and quantitative. The renillaglo luciferase assay system is a singleaddition reagent that generates a glowtype signal with renilla luciferase. The dualglo luciferase assay system is designed to work in growth media commonly used for mammalian cells with or without added serum. The dualglo luciferase assay system is a homogeneous reagent system that enables fast and simple quantitation of a stable luminescent signal from two reporter genes in a single sample. In the presence of nadh a proluciferin reductase substrate is converted by reductase to luciferin, which is then used in a luciferase reaction to produce. Dualluciferase reporter assay system kirschner lab. The halflife of these reagents remains the same with or without use of glb, 5 hours for steadyglo reagent and 24 minutes for bright. Can be used for assays in singletube luminometers or in multiwellplate luminometers with autoinjectors. The assay measures cellular atp as a surrogate measure of host cell viability. Promega manufacturing and delivery systems continue to be fully operational during the covid19 outbreak.
In 384well mode, a trituration step was added after pipetting each reagent increasing linearity at lower concentrations of luciferase. The firefly luciferase reporter is measured first by adding luciferase assay reagent. Firefly luciferase, a monomeric 61kda protein, cat. Luciferase assays cell viability assay kits biotium. Email promega technical services if you have questions on use of this system. The renilla luciferase assay system is formulated with a.
In addition, the luciferase reporter vectors contain numerous features aiding in the structural characterization of the putative regulatory sequences under investigation. Monitoring of renilla luciferase activities invitro and invivo the expression of a novel enduren and viviren renilla luciferase reporter gene promega in vitro and in vivo is explored. Renilla luciferase assay system is designed to provide a fast, sensitive method of detecting luciferase from sea pansy renilla reniformis. For automated applications, the dlr assay is performed directly in the multiwell plate. Light is produced by converting the chemical energy of luciferin oxidation through an electron transition, forming the product molecule oxyluciferin. Luciferase reporter assay for deciphering gpcr pathways. The convenient addmixmeasure protocol generates both firefly and renilla luciferase luminescence signals from cells that have not been preconditioned or. Our teams are in regular contact with suppliers and distributors worldwide to manage inventory of raw materials to ensure continued availability. For an overview and information on the development of the dualglo luciferase assay system, see sections 6. Luciferase assay systems quick protocol card, fb026 promega. Dual luciferase reporter assay chemistry continued in the dlr assay chemistry, the kinetics of the renilla luciferase reaction provide a stabilized luminescent signal that.
Luciferase assay system, 10pack e1501 from promega. The promoterdriven firefly control vectors are commonly cotransfected with experimental nanoluc or renilla luciferase vectors for use in dualreporter assays such as the dualluciferase or dualglo reporter assay systems. The viral toxglo assay is a simple, quantifiable method of determining viralinduced cytopathic effects cpe in host cells caused by lytic virions. To see articles that cite the use of the dualglo luciferase assay system, visit.
Chemistry of luciferase enzyme assay diagram courtesy of promega corp. Product components and storage conditions product size cat. Signal detection may become saturated when measuring very high lightemitting samples in a luminometer or scintillation counter. Dualluciferase reporter assay system technical manual. Nih3t3 cells expressing the luciferase gene from rous sarcoma virus were lysed with 1x cell culture lysis. Therenilla luciferase assay has a linear range covering six orders of magnitude and allows for the detection of approximately. The reactions are started by the injection of 200 l of the 0. Bioglo assay reagent is functionally tested for performance in the adcc reporter bioassay and is intended for use with this or other bioassays. Luciferase assays can be used to investigate a variety of cellular processes using bioluminescent light output as a sensitive measure of biological activity. Riiah adcp reporter bioassay, core kit 5x 1 each g9995 not for medical diagnostic use.
Luciferase reactions were performed according to manufacturers. The nanoglo dual luciferase reporter nanodlr assay system is a homogeneous reagent system that allows you to sequentially detect the activities of firefly photinus pyralis luciferase and nanoluc luciferase nluc from a single sample. The dualluciferase reporter dlr assay system provides an efficient. In the dlr assay, the activities of firefly photinus pyralis and renilla renilla reniformis, also known as sea pansy luciferases are measured sequentially from a single sample. Promegas luciferase assay systema,b is substantially improved over. Renilla luciferase assay system quick protocol, 9fb066 promega. When reconstituted, it has the capacity to lyse cells, reduce the autoluminescence of the coelenterazine substrate, and produce a stable signal halflife greater than 60 minutes at 22c. Following singlecopy integration of the fluc2anlucp biocircuit into a gene locus relevant to parkinsons disease, hts using nanodlr easily distinguished compounds affecting one of the. Luciferase assay buffer 10ml for e152a and 100ml for. Light is produced by converting the chemical energy of luciferin oxidation through an electron transition, forming the. Nanoglo dualluciferase reporter assay system nextgen. It is widely used because it is convenient, relatively inexpensive, and gives quantitative measurements instantaneously. Aug 26, 20 the luciferase reporter assay is commonly used as a tool to study gene expression at the transcriptional level. Optimization of the luciferase assay kit may be necessary for use with your equipment or samples.
The oneglo luciferase assay system provides a sensitive, robust, homogeneous assay for detection of firefly luciferase reporter gene expression in mammalian cells. The firefly luciferase reporter assay is initiated by adding an aliquot of lysate to luciferase assay reagent ii. Luciferase enzyme is regularly used to study a wide range of. Affordable luciferase reporter assay for cellbased highthroughput screening ellen siebringvan olst1 christie v, ermeulen 2, renee x. Nih3t3 cells expressing the luciferase gene from rous sarcoma virus were lysed with 1x cell culture lysis reagent 48 hours after infection. The luciferase assay system also can be used for quantitation in plant and bacterial cells, but only cell culture lysis reagent is. Glo lysis buffer is fully compatible with brightglo, steadyglo, oneglo and renillaglo luciferase assay reagents and the luciferase assay reagent for analysis of firefly luciferase expression. See the product label for the expiration date of cat. The luciferase reporter assay is commonly used as a tool to study gene expression at the transcriptional level. A new luciferase substrate has enabled promega to create a novel reagent that is more robust and stable when reconstituted, and less aromatic than standard reagents created with luciferin. Linear results are seen over at least eight orders of magnitude of enzyme concentration, and less than 10 20 moles of luciferase can be measured under optimal conditions. Dlrz assay system, the linear range of the firefly luciferase assay is seven orders of magnitude, providing detection sensitivity of.
The approximately 2hour signal halflife provides greater flexibility in assay design. Renilla luciferase assay system technical manualpdf promega. Oneglo ex luciferase assay system provides both the high sensitivity and longlived luminescence required to batchprocess multiple 96, 384 or 1,536well plates. Dualluciferaser reporter assay system tm046 promega. Dualluciferase reporter assay system protocol promega. The nanoglo dualluciferase reporter nanodlr assay system allows sensitive detection of firefly and nanoluc luciferases. Reporter gene systems are used widely for studying gene regulation in response to gpcrs. Promega corp priority date the priority date is an assumption and is not a legal conclusion. Impact of the synthetic vectors with renilla luciferase assay reagent. Robust firefly luciferase assay with improved performance for high and ultrahighthroughput applications. The dualluciferase reporter dlr assay system provides an efficient means of performing two reporter assays. Multiplexing assays in the same sample well allows complementary measures of cell health to correct for errors in culturing, plating or assay chemistry interferences. Bioglo assay reagent contains a new luciferase substrate, resulting in a reagent that is more stable and more tolerant to sample components than standard luciferase assay reagents.
It has broad applications across various fields of cell and molecular biology wherever you want to measure or track expression of a cloned gene. This listings of restriction sites for the pgl2 luciferase reporter vectors are provided in sections 7. In the dlr assay, the activities of firefly and renilla luciferases are measured sequentially from a single sample. The oneglo luciferase assay system employs new assay chemistry to provide a robust, homogeneous assay. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed. The lactateglo assay couples lactate oxidation and nadh production with a bioluminescent nadh detection system. Promega s luciferase assay systema,b is substantially improved over conventional assay methods in both sensitivity and simplicity 2,68.
Nanoglo dualluciferase reporter assay protocol promega. If this occurs, dilute your sample with 1x lysis buffer and repeat the assay. Dualluciferase reporter assay system harvard university. Promegas luciferase assay systema,b is substantially improved over conventional assay methods in both sensitivity and simplicity 2,68. Product components and storage conditions continued product size cat. Bioglonl assay reagent contains a new luciferase substrate, resulting in a reagent that is brighter, more stable and more tolerant to sample components. Renilla and firefly luciferase luminescence were compared in renilla luciferase assay reagent rlar or luciferase assay reagent lar, respectively, over a titration range of 1. A veritas microplate luminometer method for promega. Luciferase assay systems instructions for use of products e1483, e1500, e1501, e1531, e4030, e4530 and e4550. Allard and kevin kopish, promega corporation luciferase reporter technology gives the researcher unparalleled sensitivity, dynamic range, versatility and ease of use when investigating questions that involve gene regulation. Reagent cclr, also can be used for reporter quantitation in plant and bacterial cells.
Promega glomax 2020 single tube luminometer using the dual glo program with integration time of 1 second. Luciferase assay system protocol promega corporation. For kits e1500, e1501, e4030, e4530 and e4550, prepare luciferase assay reagent lar by adding luciferase assay buffer 10ml for e152a and 100ml for e152b to the vial of lyophilized luciferase assay substrate. Promega corporation 2800 woods hollow road madison, wi 537115399 usa toll free in usa 8003569526 phone 6082744330 fax 6082772516. Luminescent signals generated in the dualluciferase reporter assay system by firefly and renilla. Our teams are in regular contact with suppliers and distributors worldwide and are taking all steps necessary to address both demands for diagnostic tools and reliable delivery of all products as quickly as possible. We offer a full range of luciferase assay products and tools for your research. Comparison of the promega luciferase assay system to the conventional luciferase assay method. Compatibility of the pierce bca protein assay with promega lysis. Linear results are seen over at least 8 orders of magnitude of enzyme concentration, and the patented technology incorporated into this assay system allows measurement of less than 1020 moles of luciferase under optimal conditions. Generally, 100fold greater sensitivity can be achieved over the chloramphenicol acetyltransferase cat assay. A protocol for reporter quantitation in mammalian cells. Promega s kinaseglo assay kit is a homogenous method for virtually any kinase and substrate combination using a nonradioactive luciferase assay.
For use as a genetic reporter, multiple forms of nanoluc luciferase have been configured to meet differing experimental objectives. There are many reporter genes available, including alkaline phosphatase. The kinase substrate can be a peptide, protein, or lipid. Promega luciferase assay systems fisher scientific. Quantitation of luminescent signal from each of the luciferase reporter enzymes may be performed immediately following lysate preparation without the need for dividing samples or performing additional treatments.
Renilla luciferase in the existing homogenous fireflyrenilla dualluciferase assay dualglo, allowing both luciferases to be dynamic reporters. The shaded area represents the light typically lost in measurements where the cell. The luciferase assay system was developed for reporter quantitation in mammalian cells. Luciferase reactions were performed according to manufacturers instructions using titration curves of purified recombinant quantilum recombinant firefly luciferase promega or purified recombinant renilla luciferase ray biotech. The luciferase assay system is an extremely sensitive and rapid reagent for quantitation of firefly luciferase. Brightglo luciferase assay system technical manual pdf 383 kb english. Promega offers the pgl4 series of firefly and renilla luciferase vectors designed for use with the dlr assay systems. Reporter assay and dualluciferaser promega corporation. Transcriptional regulation, coupled to the expression of a luciferase reporter gene, is regularly used to. To perform the luciferase assay, 100 l of each plate lysate is added to 360 l of luciferase assay buffer table i in a luminometer vial. E3971 can be used with the luciferase assay system for reporter quantitation in mammalian cells.