Question 1

Will the reducing conditions required for digestion impact the interchain disulfide bond integrity?

Felicia Mejare · Accepted Answer

Depending on what reducing agent and what concentration you use you can obtain intact Fab and not reduce the interchain disulfide bond. The light chain (LC) and heavy chain (HC) will not dissociate under native conditions due to hydrogen/hydrophobic forces. Before you analyze your samples with SDS-PAGE you must remove the cysteine otherwise the Fab will be reduced to Fd and LC during sample preparation for the SDS-PAGE. This can be done by a simple buffer exchange.

Question 2

Will the reducing conditions required for digestion impact the interchain disulfide bond integrity?

Felicia Mejare · Accepted Answer

Depending on what reducing agent and what concentration you use you can obtain intact Fab and not reduce the interchain disulfide bond. The light chain (LC) and heavy chain (HC) will not dissociate under native conditions due to hydrogen/hydrophobic forces. Before you analyze your samples with SDS-PAGE you must remove the cysteine otherwise the Fab will be reduced to Fd and LC during sample preparation for the SDS-PAGE. This can be done by a simple buffer exchange.

Question 3

Will the reducing conditions required for digestion impact the interchain disulfide bond integrity?

Felicia Mejare · Accepted Answer

Depending on what reducing agent and what concentration you use you can obtain intact Fab and not reduce the interchain disulfide bond. The light chain (LC) and heavy chain (HC) will not dissociate under native conditions due to hydrogen/hydrophobic forces. Before you analyze your samples with SDS-PAGE you must remove the cysteine otherwise the Fab will be reduced to Fd and LC during sample preparation for the SDS-PAGE. This can be done by a simple buffer exchange.