GeneBLAzer® Substrate Loading Kits
In addition to the dual-color readout of positive and negative cells, FRET-based substrates offer the advantage of ratiometric data analysis. Ratiometric methods eliminate well-to-well differences in cell number and substrate concentration caused by variations in hand pipetting or liquid handling instrumentation. Ratiometric methods also reduce or eliminate variations caused by excitation path length, fluorescence detectors or volume changes. These variations contribute to experimental noise and can mask the underlying biological response of interest.
Our broad range of substrates and assay formats provide you with the optimal choice for your experimental objectives and preferences.
Recommended GeneBLAzer® Substrate Loading Kit By Application
Our broad range of substrates and assay formats provide you with the optimal choice for your experimental objectives and preferences.
Recommended GeneBLAzer® Substrate Loading Kit By Application
LiveBLAzer™ - FRET B/G Substrate Loading Kits
Ratiometric, live cell β-lactamase quantitation
LiveBLAzer™-FRET B/G Loading Kits provide reliable, ratiometric detection of beta-lactamase activity in live cells. The blue/green ratio provides a normalized reporter response with smaller %CVs than single channel readouts. Each substrate consists of two fluorophores that form an efficient FRET pair, which in the absence of beta-lactamase activity results in a green fluorescence signal at 518 nm. In the presence of beta-lactamase activity, cleavage of the substrate disrupts FRET, resulting in a blue fluorescence signal at 447 nm. The blue signal can be readily observed under a microscope and can also be detected on bottom-read fluorescent microplate readers.
The LiveBLAzer™-FRET B/G Loading Kit:
This live cell microscopy image shows a population of CHO-K1 cells expressing β-lactamase as measured by LiveBLAzer™- FRET B/G Substrate.
LiveBLAzer™-FRET B/G Loading Kits provide reliable, ratiometric detection of beta-lactamase activity in live cells. The blue/green ratio provides a normalized reporter response with smaller %CVs than single channel readouts. Each substrate consists of two fluorophores that form an efficient FRET pair, which in the absence of beta-lactamase activity results in a green fluorescence signal at 518 nm. In the presence of beta-lactamase activity, cleavage of the substrate disrupts FRET, resulting in a blue fluorescence signal at 447 nm. The blue signal can be readily observed under a microscope and can also be detected on bottom-read fluorescent microplate readers.
The LiveBLAzer™-FRET B/G Loading Kit:
- Reduces false hits by minimizing experimental noise with the ratiometric blue/green readout
- Enables rapid development of stable cell lines using flow cytometry
- Facilitates miniaturization and measurement of small biological responses due to high sensitivity
- Minimizes sample handling and increases throughput with a homogeneous format
This live cell microscopy image shows a population of CHO-K1 cells expressing β-lactamase as measured by LiveBLAzer™- FRET B/G Substrate.
ToxBLAzer™ DualScreen Loading Kits
Ratiometric, live cell beta-lactamase quantitation with cytotoxicity indicator
The ToxBLAzer™ DualScreen Assay combines the advantages of LiveBLAzer™-FRET B/G with a cytotoxicity indicator in a single assay format. The readout allows you to identify false positives due to cytotoxic compounds, which is critical in screening applications which down-regulate transcription or when there is a lack of viable cells (Figure 20). The cytoxicity indicator fluorescence is red-shifted to ensure compatibility with the blue and green emission of the LiveBLAzer™-FRET B/G Substrate, allowing detection of both beta-lactamase activity and the cytotoxicity readout in the same well (Figure 21).
The ToxBLAzer™ DualScreen Assay:
A. This live cell microscopy image shows a population of cells expressing beta-lactamase as measured by LiveBLAzer™-FRET B/G Substrate.
B. The same population of cells analyzed with beta-lactamase reporter activity are shown with the red emission
The ToxBLAzer™ DualScreen Assay combines the advantages of LiveBLAzer™-FRET B/G with a cytotoxicity indicator in a single assay format. The readout allows you to identify false positives due to cytotoxic compounds, which is critical in screening applications which down-regulate transcription or when there is a lack of viable cells (Figure 20). The cytoxicity indicator fluorescence is red-shifted to ensure compatibility with the blue and green emission of the LiveBLAzer™-FRET B/G Substrate, allowing detection of both beta-lactamase activity and the cytotoxicity readout in the same well (Figure 21).
The ToxBLAzer™ DualScreen Assay:
- Provides rapid identification of false positives due to toxicity or lack of viable cells
- Reduces false hits by minimizing experimental noise with the ratiometric blue:green reporter readout
- Eliminates need for follow-up cytotoxicity assays by multiplexing the reporter readout with a cytoxicity readout
A. This live cell microscopy image shows a population of cells expressing beta-lactamase as measured by LiveBLAzer™-FRET B/G Substrate.
B. The same population of cells analyzed with beta-lactamase reporter activity are shown with the red emission
LyticBLAzer™ Loading Kits
LyticBLAzer™ Substrates provide an end-point method for analyzing beta-lactamase activity in cell lysates using any fluorometer. By combining the components for lysis and detection in a single reagent addition, the LyticBLAzer™ Substrates provide a very simple assay. They are available in two detection formats: ratiometric (FRET) or fluorescence intensity (FI) based on the BODIPY® dye. Complete LyticBLAzer™ Substrate Kits are available in homogeneous as well as non-homogeneous assay formats, each optimized for a different assay condition (Figure 22).
LyticBLAzer™ Substrate Kits:
A. Starting with cells in cell culture medium, simply add 2X lysis and detection solution. There is no requirement for plate shaking or mixing. Following an incubation at room temperature, read the results on a fluorescent plate reader.
B. Starting with cells in cell culture media, simply aspirate the medium and add the 1X lysis and detection solution. There is no requirement for plate shaking or mixing. Following an incubation step, read the results on a fluorescence plate reader.
LyticBLAzer™ Substrate Kits:
- Enable quantitation of beta-lactamase activity on top-read or bottom-read fluorescence instrumentation
- Reduce false hits by minimizing experimental noise using LyticBLAzer™ -FRET B/G Ratiometric (blue:green) reporter readout
- Minimize fluorescence interference caused by sample contaminants such as dust and lint using LyticBLAzer™ -BODIPY® FL dye
- Provide long read times with stable fluorescence
A. Starting with cells in cell culture medium, simply add 2X lysis and detection solution. There is no requirement for plate shaking or mixing. Following an incubation at room temperature, read the results on a fluorescent plate reader.
B. Starting with cells in cell culture media, simply aspirate the medium and add the 1X lysis and detection solution. There is no requirement for plate shaking or mixing. Following an incubation step, read the results on a fluorescence plate reader.
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