QBT Fatty Acid Transporter Assay
The QBT™ Fatty Acid Uptake Assay is a single-step, homogeneous, fluorescent assay for monitoring the activity of fatty acid transport proteins.
Conventional protocols for monitoring fatty acid uptake using radioactivity often require cell lysis and processing at very low temperature, making them expensive, slow and unsuitable for high throughput screening. Other fluorescence-based protocols generally require the use of low throughput fluorescence activated cell sorter (FACS) instrumentation, or require cell washing, which can severely compromise the integrity of the fragile adipocytes used in these assays.
The QBT™ Fatty Acid Uptake Assay is a homogeneous assay amenable to high-throughput screening. It uses a BODIPY® -dodecanoic acid fluorescent fatty acid analog coupled with Molecular Devices proprietary quench technology. The BODIPY label provides an ideal long chain fatty acid analog that behaves much like natural fatty acids and is a known substrate for fatty acid transporters. The elimination of radioactive compounds results in easier reagent handling, reduced disposal costs and eliminates safety risks associated with radiolabel assays.
Benefits of the QBT™ Fatty Acid Uptake Assay Kit
- Replaces cumbersome and time-consuming radioactive assays, reducing disposal costs and eliminating safety risks associated with radioactive assays.
- One-step, homogeneous assay format enables high-throughput screening.
- Fluorescent assay with proprietary quench technology allows users to monitor real-time uptake kinetics in live cells.
QBT Fatty Acid Uptake Assay Kit includes a loading buffer that contains fluorescent fatty acid analog, along with a quenching dye to greatly reduce fluorescence in the extracellular space. When loading buffer is added to cells, transport of fluorescent fatty analog into the cells results in an increase in fluorescence signal that is monitored in real time using a bottom-reading fluorescence microplate reader. Inhibition of fatty acid transport by test compounds results in reduced fluorescent signal relative to controls.
- Prepare differentiated adipocytes from 3T3-L1 fibroblasts, or other FATP-expressing cell type as desired (for details on cell preparation, please refer to the QBT Fatty Acid Uptake Assay Kit product insert).
- Plate cells so that a confluent monolayer forms.
- Serum deprive the cells if necessary.
- Treat cells with test compounds.
- Add the recommended volume of loading buffer to wells.
- Transfer the plates immediately to a fluorescence microplate reader and set the microplate reader to read wells every 20 seconds for 30-60 minutes.
For more details, please refer to the QBT™ Fatty Acid Uptake Assay Kit product insert, which may be accessed through the Customer Portal.