Description
Cholesterol Assay Kit (Fluorometric) (BA0021) (BA0021)
The Cholesterol Assay Kit (Fluorometric) (SKU: BA0021) offers a straightforward, non-radioactive means of measuring cellular cholesterol uptake directly in a 96-well plate. Cholesterol is a sterol present in cell membranes and transported in the bloodstream, where it contributes to membrane structure, hormone synthesis and cell signalling. Dysregulated cholesterol handling has been linked to obesity, heart disease and the elevated uptake seen in highly proliferative cancer cells. The assay relies on the cellular uptake of a fluorescently tagged cholesterol probe, with the fluorescence measured at λex/em = 485/535 nm being proportional to the amount of cholesterol taken up. Cells are treated directly in the assay plate, making the method convenient and readily adaptable to high-throughput screening. It is well suited to studying cholesterol metabolism and to screening compounds that modulate cholesterol uptake.
| Product Name: | Cholesterol Assay Kit (Fluorometric) (BA0021) |
| SKU: | BA0021 |
| Detection Method: | Fluorometric, cell-based cholesterol uptake (λex/em = 485/535 nm) |
| Sample Type: | Adherent cells cultured in 96-well plates |
| Species Reactivity: | All |
| Kit Size: | 100 Assays |
| Equipment Required: | Microplate reader |
| Storage: | Store all components at -20°C upon receipt. |
| Shelf Life: | 6 months after receipt. |
| Shipping: | Room Temperature |
A fluorometric, cell-based assay for the quantitative determination of cholesterol uptake by adherent cells. The kit uses a fluorescently tagged cholesterol probe; the fluorescence intensity measured at λex/em = 485/535 nm is proportional to the quantity of cholesterol taken up by the cells.
- Convenient: cells are treated directly in a 96-well fluorescent plate.
- Safe: non-radioactive assay.
- High-throughput: can be readily automated for thousands of samples per day.
- Measurement of cholesterol uptake by adherent cells.
- Screening of cholesterol uptake inhibitors.
- Evaluation of the effect of drugs on cholesterol uptake.
Note: The below protocol is a sample protocol. Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
| Step | Procedure |
| 1 | Prepare Tracer Medium by diluting Fluorescent Tracer 1:50 in serum-free or low-percentage FBS (<1%) medium. |
| 2 | Using a black flat-bottom 96-well plate, seed control wells at the desired density in 100 µL Tracer Medium. For the positive control, dilute the 2.5 mM Positive Control 1:1000 in Tracer Medium (final 2.5 µM) and seed cells in 100 µL. For each treatment, spike the compound into Tracer Medium and seed cells in 100 µL. |
| 3 | Allow cells to propagate for 24 to 72 hours or to the desired confluence. |
| 4 | Carefully aspirate the culture medium from all wells. |
| 5 | Rinse all wells twice with 100 µL 1x PBS, removing all PBS when finished. |
| 6 | Add 100 µL Assay Reagent to all wells. |
| 7 | Read fluorescence at λex/em = 485/535 nm immediately. |
Compare the fluorescence intensity of each treatment relative to the controls. Wells with greater fluorescence indicate an increase in cholesterol uptake; wells with lower fluorescence indicate a decrease.
| Component | Quantity | Storage |
| Assay Reagent | 12 mL | -20°C |
| Fluorescent Tracer | 250 µL | -20°C |
| Positive Control (2.5 mM) | 20 µL | -20°C |
