Description
HDL and LDL/VLDL Assay Kit (BA0119) (BA0119)
The HDL and LDL/VLDL Assay Kit (SKU: BA0119) provides a simple, direct and automation-ready colorimetric method for measuring cholesterol in high-density lipoprotein (HDL) and low-/very-low-density lipoprotein (LDL/VLDL) fractions. These cholesterol concentrations are strong predictors of coronary heart disease. The kit uses an improved PEG precipitation method to separate HDL from LDL/VLDL, then determines cholesterol using a cholesterol esterase/cholesterol dehydrogenase reagent in which NAD is reduced to NADH. The optical density of the NADH formed at 340 nm is directly proportional to the cholesterol concentration in the sample. The room-temperature procedure requires no 37C heater.
| Product Name: | HDL and LDL/VLDL Assay Kit (BA0119) |
| SKU: | BA0119 |
| Detection Method: | Colorimetric (340 nm) |
| Detection Range: | 5 - 300 mg/dL cholesterol (detection limit 5 mg/dL) |
| Sample Type: | Serum |
| Species Reactivity: | All |
| Assay Time: | 30 minutes |
| Kit Size: | 100 Assays |
| Equipment Required: | Microplate reader |
| Storage: | -20C |
| Shelf Life: | 6 months after receipt |
| Shipping: | Gel Pack |
A colorimetric assay that separates and quantifies HDL and LDL/VLDL cholesterol from serum via PEG precipitation and an NAD-coupled cholesterol reaction.
- Sensitive and accurate, requiring only a 20 uL serum sample, with a detection limit of 5 mg/dL and linearity up to 300 mg/dL cholesterol
- Convenient room-temperature assay with no 37C heater required
- Direct assay of HDL and LDL/VLDL cholesterol in serum samples from any species
- Evaluation of the effects of drugs on cholesterol metabolism
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 | Bring all reagents except the Enzyme Mix to room temperature. Use non-haemolysed serum. The procedure is designed for duplicate determinations. |
| 2 | Sample Preparation: mix 20 uL serum with 20 uL Precipitation Reagent, vortex and centrifuge 5 min at 9,500 g. Transfer 24 uL supernatant into a clean tube with 96 uL Assay Buffer (HDL). Resuspend the pellet in 40 uL PBS, transfer 24 uL of the mixture into a tube with 96 uL Assay Buffer (LDL/VLDL). Prepare a Total tube from 12 uL serum and 108 uL Assay Buffer, and a Standard tube from 12 uL of the 300 mg/dL standard and 108 uL Assay Buffer. |
| 3 | Assay: transfer 50 uL Assay Buffer (Blank), Standard, Total, HDL and LDL/VLDL into wells of a clear-bottom 96-well plate. |
| 4 | Prepare Working Reagent per well from 50 uL Assay Buffer, 18 uL NAD Solution and 1 uL Enzyme Mix. Transfer 60 uL to each reaction well and tap to mix thoroughly. |
| 5 | Incubate 30 minutes at room temperature and read OD values at 340 nm. |
[Total] = (ODTOTAL - ODBLANK) / (ODSTANDARD - ODBLANK) x 300 (mg/dL); [HDL] = (ODHDL - ODBLANK) / (ODSTANDARD - ODBLANK) x 300 (mg/dL); [LDL/VLDL] = (ODLDL/VLDL - ODBLANK) / (ODSTANDARD - ODBLANK) x 300 (mg/dL).
| Component | Quantity | Storage |
| PBS | 1.5 mL | -20C |
| Precipitation Reagent | 1.5 mL | -20C |
| Assay Buffer | 20 mL | -20C |
| Enzyme Mix | 120 uL | -20C |
| NAD Solution | 2 mL | -20C |
| Standard (300 mg/dL cholesterol) | 1 mL | -20C |
