Desalting Protein Solutions prior to Sequencing

Product: 7216-06 BAKERBOND spe™ Wide-Pore Butyl (C4) Disposable Extraction Columns

Sample Preparation: No sample pretreatment is required. Filtration may be necessary.

Column Conditioning: Slowly aspirate or force two column volumes (12ml) of methanol immediately followed by one column volume (6ml) of water through the columns. Do not allow the columns to dry during this stage or before the samples are applied.

Sample Addition: With the vacuum off, add the samples to the columns. Slowly aspirate or force the samples through the columns at approximately 5 ml/min.

Column Wash: Aspirate or force three-column volumes (18ml) water through the columns.

Protein Elution: Elute with two or more void volumes (one void volume=1.0-1.2 ul per 1mg sorbent) of elution solvent. Elution solvent: H2O/Isopropanol/TFA (0.1%) in the appropriate ratio depending on the hydrophobicity of the protein.

Note: The large quantity of packing in the 6 ml, 500mg column may result in poor recoveries when nanomoles of protein are desalted. A 1 ml column packed with 100mg is more ideal. This configuration is not commercially available, but can be prepared by packing a 1 ml filtration column (7121-01) with BAKERBOND Wide-Pore™ Butyl(C4) (7283-02). Wash and elution volumes should be reduced accordingly. Recoveries from normal pore butyl are predictably poor due to the hydrophobic surface and small pore size. In many cases, both properties contribute to poor recoveries.