NITROCELLUOSE

Proteins separated by SDS-PAGE and transferred to Nitrocellulose can be used as antigens in a custom antibody project. Nitrocellulose is especially practical when small (<10kDa) proteins are used. The size and purity of the isolated proteins can be assessed by using a reversible stain such as Ponceau S. Generally, a band that can be visualized by Ponceau S contains enough protein for use in a custom antibody project.

Note: It is important to use pure Nitrocellulose as Nitrocellulose reinforced with acetate or plastics can interfere with processing steps used by Quality Bioresources, Inc. and, therefor, the titer and affinity of the resulting antibodies.

Isolating Protein Bands From Nitrocellulose*

Rinse nitrocellulose with deionized water following transfer of proteins from an SDS-PAGE gel.
Stain nitrocellulose with 0.1% Ponceau S (w/v) in 5% acetic acid (ex. Sigma P 7170)
Rinse with deionized water until bands are readily visible
Excise desired band with a clean razorblade
Rinse excised nitrocellulose strip until the band(s) are no longer distinguishable from the background.
Allow nitrocellulose strip to dry completely, then seal in an appropriately sized container (eppendorff tube, cryo-vial, etc.).

* Adapted from Harlow & Lane. Antibodies a Laboratory Manual (1988)