Service E: Establishment of the purification protocol

Biomass either provided by you or alternatively prepared by us if using service D.

1. Preparation of the „Inclusion Bodies“ (IB)

• Disintegration of bacteria by lyses buffer and sonification .
• Accumulation of „IBs“ by several centrifugation steps .
• Check of „IB- preparation“ by SDS/PAGE and Comassie stain.

2. Solubilisation of „Inclusion Bodies“

• Solubilisation of „IBs“ .
• Check of „IBs“ by SDS/PAGE and Comassie stain

Note:  Further measurements concering the purification depend on the protein of interest and the information available.

The following features influence the subsequent proceeding.

1. existence of an established purification protocol,
2. the protein is fused to a “tag”,
3. the native protein is a dimer, or
4. there has to be established a new protocol.

In case of 3. the next step is the dimerization reaction followed by a gel filtration to separate monomers from dimers.

In case of 4. the process is continued with ion exchange chromatography (e.g. anion/cation).

• The total yield of purified protein.
• A detailed report sheet.

Dependent on the protocol used.