Description
Parallel Artificial Membrane Permeability Assay-Skin Kit (BA0262) (BA0262)
The Parallel Artificial Membrane Permeability Assay-Skin Kit (SKU: BA0262) is designed to aid in evaluating skin membrane permeability. Membrane permeability is an important characteristic when evaluating compounds as potential drug candidates, since drugs often need to cross membranes to reach their target of action. The skin, and in particular the stratum corneum, is a complex barrier that can be mimicked, and rapid, early screening of compounds for skin penetration is highly desirable in drug discovery. While permeability can be evaluated by cell-based methods, these are often expensive and time-consuming; Parallel Artificial Permeability Assays (PAMPA) instead offer a quick, inexpensive means of evaluating the permeability of test compounds. This kit provides all the necessary components to run a PAMPA plate for skin permeability studies, including donor and acceptor plates, a ready-to-use skin mimic solution and high-, medium- and low-permeability controls. The procedure is straightforward, low-cost and readily automated for high-throughput screening.
| Product Name: | Parallel Artificial Membrane Permeability Assay-Skin Kit (BA0262) |
| SKU: | BA0262 |
| Detection Method: | Parallel Artificial Membrane Permeability Assay (PAMPA); UV absorbance (200-500 nm) |
| Sample Type: | ['Test compounds'] |
| Species Reactivity: | All |
| Assay Time: | 18 hours incubation (typically 16-24 hours) |
| Kit Size: | 96 Assays |
| Equipment Required: | Microplate reader (UV/absorbance spectrum capable) |
| Storage: | Store Permeability Controls and Skin Mimic Solution at -20°C; store Donor Plate, Acceptor Plate and Working Tray at room temperature |
| Shelf Life: | 12 months |
| Shipping: | Room Temperature |
PAMPA kit providing all components required to run a 96-well artificial membrane permeability assay for the quantitative determination of skin membrane permeability.
- Convenient: includes all necessary equipment to run a PAMPA plate.
- Simple and low-cost: procedure is easy to follow and more affordable than cell-based permeability assays.
- High-throughput: readily automated as a 96-well plate assay for thousands of samples per day.
- Direct assessment of skin membrane permeability of test compounds.
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 | Equilibrate all components to room temperature and briefly centrifuge tubes before opening. Bring the Skin Mimic Solution to room temperature and ensure it is completely dissolved and homogeneous (it comes ready to use). |
| 2 | Prepare 10 mM test compound stock solutions in DMSO (the supplied Permeability Controls are provided as 10 mM DMSO solutions). |
| 3 | In separate tubes, prepare 500 µL of 500 µM Test Compound (25 µL 10 mM compound + 475 µL PBS); dilute Permeability Controls to 500 µM similarly. |
| 4 | Prepare 200 µM Equilibrium Standards for each compound and control (80 µL of 500 µM + 120 µL PBS), and a Blank Control (5 µL DMSO + 245 µL PBS); set aside for next-day analysis. |
| 5 | Add 300 µL PBS to wells in the acceptor plate. |
| 6 | With the donor plate still in its tray, add 17 µL Skin Mimic Solution directly to the well membranes, taking care not to puncture them. |
| 7 | Add 200 µL of each 500 µM Test Compound and Permeability Control to duplicate wells of the donor plate (run all variables at least in duplicate). |
| 8 | Carefully place the donor plate into the acceptor plate wells and incubate at RT or 37°C for 18 hours (or 16-24 hours as desired). |
| 9 | Carefully remove the donor plate and collect the liquid in the acceptor plate wells (Acceptor Solution). |
| 10 | Add 100 µL of Acceptor Solution, Equilibrium Standards and Blank Control to wells of a UV plate (Cat # P96UV). |
| 11 | Read the absorbance spectrum from 200 nm to 500 nm in 10 nm intervals to determine peak absorbance; peak absorbance for High, Medium and Low Permeability Controls are 260 nm, 260 nm and 270 nm respectively. Alternatively, analysis can be performed by HPLC, MS or other quantification methods. |
Determine the Permeability Rate (Pe) using Pe = C × -ln(1 - ODA/ODE) cm/s, where ODA is the absorbance of the Acceptor Solution minus Blank and ODE is the absorbance of the Equilibrium Standard minus Blank. For an 18-hour incubation, C = 7.72×10^-6. For other incubation times, calculate C = (VD × VA) / ((VD + VA) × Area × time) cm/s, where Donor Volume (VD) = 0.2 cm3, Acceptor Volume (VA) = 0.3 cm3, Membrane Area = 0.24 cm2, and time = 64,800 s for 18 hours.
| Component | Quantity | Storage |
| Donor Plate | 1 Plate | Room temperature |
| Acceptor Plate | 1 Plate | Room temperature |
| Working Tray | 1 Plate | Room temperature |
| Skin Mimic Solution | 2.0 mL | -20°C |
| High Permeability Control | 120 µL | -20°C |
| Medium Permeability Control | 120 µL | -20°C |
| Low Permeability Control | 120 µL | -20°C |
