The Transfer Process During Electrophoresis: Nitrocellulose(NC) Membranes
What is NC Membrane?
Nitrocellulose membranes are a popular matrix used in protein blotting because of their high protein-binding affinity, compatibility with a variety of detection methods, and the ability to immobilize proteins, glycoproteins, or nucleic acids. The nitrocellulose membranes are pure nitrocellulose media that has a high binding capacity for proteins. It is easily wetted in water or transfer buffer and compatible with a wide range of protein detection systems.
Compared to other types of membrane:
| Reprobe characteristics | Binding interactions | Binding capacity | Advantages | Disadvantages | |
| Nitrocellulose | Can be stripped and reprobed | Hydrophobic and electrostatic | 80–100 µg/cm2 | Tendency to exhibit lower background | Can be brittle and fragile, which limits use in stripping and reprobing |
Applications:
- Transfer of:
- protein from SDS-PAGE gels to the membrane in preparation for western blotting
- a wide range of protein molecular weights and nucleic acids >500 bp (0.45 µm) / <500 bp (0.2 µm)
- nucleic acids in preparation for northern or Southern blotting
- Immunodetection of target proteins and probe hybridization of nucleic acids
Pore Sizes:
NuSep’s 0.45 µm pore–size nitrocellulose membrane is dense 100% nitrocellulose. Pure nitrocellulose is the most frequently used membrane for the transfer of both proteins and nucleic acids. It is a proven medium for western, northern, and Southern blotting that produces excellent signal-to-noise results. NuSep nitrocellulose is easy to handle, wets readily with water-based solutions, and resists the brittleness usually associated with nitrocellulose. The 0.45 µm pore size is recommended for most analytical blotting, including protein, ssDNA, and RNA transfers.
The 0.2 µm pore size is optimized for transferring nucleic acids and low molecular–weight proteins: the smaller pore size minimizes protein sample loss due to transfer through the membrane.
