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Blotting Membranes and Filter Paper for Chromogenic, Chemiluminescent and Fluorescent Blotting Applications.
Thermo Scientific Blotting Membranes.Thermo Fisher Scientific offers a variety of solid supports for blotting applications in both precut and roll formats.
Nitrocellulose membranes are the most popular matrix used in protein blotting because of the high binding affinity for proteins and are compatible with a variety of detection methods. Some nitrocellulose membranes contain cellulose acetate, which inhibits binding; however, Thermo Scientific membranes contain 100% nitrocellulose for excellent binding, biological activity and sensitivity. All of the following nitrocellulose membranes are suitable for Western blotting. Most proteins can be successfully blotted using a 0.45 µm pore size membrane. For proteins of low molecular weight or peptides, a 0.2 µm pore size membrane is recommended.
PVDF (Polyvinylidene Difluoride) membranes
PVDF membranes are hydrophobic with a pore size of 0.2 or 0.45 µm. Proprietary alterations in the Pierce PVDF membrane make it more resistant to discoloration which often occurs with other commercially available PVDF membranes. These membranes have high binding affinity for proteins and nucleic acids and may be used for applications such as Western, Southern, Northern and dot blots. The PVDF membranes offer a better retention of adsorbed proteins than other supports including nitrocellulose.
In non-IR fluorescent blotting applications, the membrane will fluorescence to variying degrees depending on the excitation wavelenght used and sensitivity of the detector. In cases where the fluorescence of the membrane (not fluorescence due to nonspecific binding of the fluorophore conjugate) is too high to get an acceptable signal to noise ratio, use a low-fluorescence PVDF membrane. The Thermo Scientific Low-Fluorescent PVDF membrane has been optimized for use with fluorescent probes when excitation wavelenghts are used that cause most other membranes to produce higher levels of backgroud fluorscescence.
Charged Nylon (polyamide) membranes bind by ionic, electrostatic, hydrophobic interactions. Many of the charged nylons contain quanternary amines that form stable linkage to the nucleic acid under alkaline conditions. Fixation may not be required. Uncharged nylon relies on hydrophobic interaction from drying the nucleic acid on the membrane. Membranes with a positive zeta charge are recommended for blotting applications such as the LightShift Chemiluminescent EMSA Kit.
Western Blotting Filter Paper
Filter paper is used in Northern, Southern, and Western blots for quickly and efficiently transferring molecules from an electrophoresis gel to a membrane. These precut filter paper packs are a convenient companion for the membranes used in mini-gel sized blotting applications.
* Compatible with Chromogenic, chemiluminescent and fluorescent detection methods
o Low-fluorescence PVDF available for increased signal to noise ratio
* Available in pre-cut mini gel sizes or roll format
* Multiple pore sizes available
Important Usage Suggestions:
* Always pre-equilibrate membrane in transfer buffer prior to transfer.
* Always used powder-free gloves for handling membrane.
* Do not fold, crush, scratch or damage membrane. Membrane damage causes background.
* Always use clean labware and tweezers.
o Use disposable labware when possible in order to minimize potential for contamination.
o Note that rusty or oxidized metal implements can deposit metial ions on the membrane which will cause backgroung signal to occur when peroxide based enyzmatic substrates are used.
* Use quality grade filter papers surrounding membranes to minimize background potential.