Nano & Firefly-Glo Luciferase Reporter Assay Kit

Product Description

Reporter gene systems are commonly used in eukaryotic gene expression and cell physiology research, covering various aspects such as receptor activity, transcription factors, cell signaling, mRNA processing, and protein folding. “Dual reporter genes” are commonly utilized to enhance experimental accuracy by simultaneously expressing and measuring two independent reporter genes in one system. Typically, the reporter gene in the experimental group is associated with the effects of specific experimental conditions, while the co-transfected internal control group reporter gene serves as a calibration parameter and internal reference to establish a baseline for the response. Comparing the results of the experimental group to those of the internal control group helps minimize result fluctuations arising from variations in cell viability or transfection efficiency, as well as mitigates experimental errors stemming from sampling volume, cell lysis efficiency, and instrumental measurements. Consequently, dual reporter gene assays can generate more dependable experimental data by reducing external influences.

Nano&Firefly-Glo Luciferase Reporter Gene Detection System is a glow-type quantitative test kit with high sensitivity and a stable luminescent signal, which meets the needs of high-throughput testing. The kit detects Firefly luciferase with a new type of luciferase as substrate in the same sample, and then detects Nano-light luciferase with a new type of substrate, while quenching the fluorescent signal of Firefly luciferase, thus realizing the dual luciferase reporter gene detection.

This product has the following advantages: the use of a new luciferase detection of new substrates and Nano-light luciferase, with higher luminous intensity and better stability of the luminescence signal, the signal half-life of nearly 2 hours, which provides greater flexibility for the experimental design; high-throughput detection does not need to rely on the autosampler, and there is no need to discard the culture fluid, centrifugation and other steps to simplify the experimental process; the components of the kit to do a great deal of optimization, and compared with the traditional flash-type products, there is no obvious irritating odor. Nano&Firefly-Glo Dual luciferase Reporter Gene Assay Kit can be used in a variety of commonly used cell culture fluids: RPMI 1640, DMEM, MEM-α, F12, DMEM/F12, etc., with a half-life of about 2 hours (22℃). The half-life of the kit is about 2 hours (22℃), which meets the needs of most high-throughput experiments.

Performance

Figure 1. NANO&FIREFLYGLO luminescence quenching effect, comparing P company brands.

The results obtained by MeilunBio Nano-Light Luciferase Reporter Assay Kit maintained good consistency with P company brand. In the system of the same concentration of NANOLIGHT and Firefly luciferase, FIREFLYGLO and Stop&NANOLIGHT reaction solution were added in turn, and the luminescence intensity as well as the quenching of the two luciferasewere recorded respectively. As can be seen from the figure, the luminescence intensity of the two enzymes in the MeilunBio group is basically no different from that of the P company brand, and the quenching efficiency is higher than that of the P company brand.

Figure 2. NANOLIGHT/FIREFLYGLO luminescence stability compared to P company brand.

The results obtained by MeilunBio Nano-Light Luciferase Reporter Assay Kit maintained good consistency with P company brand. The absolute light intensity of Meilunbio’s NANOLIGHT/ FIREFLYGLO is very close to that of P company in a certain concentration range of luciferase; and the luminescence stability of Meilunbio’s NANOLIGHT is better than that of P company brand in the in vitro NANOLIGHT enzyme assay.

Figure 3. Validation of NANOLIGHT/ FIREFLYGLO linear range.

At a single moment in time, the Melun NANOLIGHT, FIREFLYGLO luciferase concentration-light intensity standard curves were linear and good (R2(NANOLIGHT)=0.999; R2(Firefly)=0.999)

Figure 4. NANOLIGHT/FIREFLYGLO luminescence detection in different media.

The data in the figure were normalized with the respective RPMI-1640 groups, and the MeilunBio system was able to luminesce normally in the commonly used media: MEMα, F12, DMEM/F12, DMEM, and RPMI-1640.

Figure 5. NANOLIGHT/FIREFLYGLO luminescence detection in different media.

The data in the figure were normalized with the respective RPMI-1640 groups, and the MeilunBio system was able to luminesce normally in the commonly used media: MEMα, F12, DMEM/F12, DMEM, and RPMI-1640.

Figure 6. Examples of NANOLIGHT /FIREFLYGLO Luminescence stability (A) and cell density- light intensity standard curve (B) in cell culture media.

NANOLIGHT and FIREFLYGLO luminescence was detected by in situ tracking in a cell culture medium. As can be seen from the figure, the NANOLIGHT and FIREFLYGLO luminescence stability and absolute light intensity are basically consistent with the P company brands; moreover, the cell density-light intensity standard curves under single moment conditions in the MeilunBio group are linear and good (R2=0.999 (NANOLIGHT); R2=0.996 (Firefly)).

Figure 7. TNFα gradient induced NF-κB signaling pathway detection effect, compared with P conpany brand.

HEK-293T cells were co-transfected with NANOLIGHT_PEST luciferase NF-κB response reporter and Firefly luciferase internal reference plasmid. After 12h post-transfection, the cells were induced by TNFα concentration gradient (0-20ng/ml) for 5h, and then incubated in cell culture system according to the steps in the instruction manual and detected the signal readings of the two enzymes using an Microplate Absorbance Reader respectively. And the NANOLIGHT and FIREFLYGLO light intensities were approximated (NANOLIGHT/Firefly). It is obvious from the figure that the NANOLIGHT /Firefly ratio is positively correlated with the concentration of the administered drug, and the detection effect of the MeilunBio group is not significantly different from that of the P company brand. (Instrument: BioTek HTX)

Shipping and Storage 

• Storage：The new kit is stored at -20℃ and is valid for one year;

  Dissolved and dispensed Firefly Luciferase Substrate is stored at -70℃ for one year protected from light, or -20℃ for short-term storage not exceeding one month.

• Shipment：

Usage Statement

Research Use Only (RUO)