Articles tagged ”F(ab’)2”

mAb Deamidation Study using FabRICATOR® Digestion and HIC Separation


Hydrophobic interaction chromatography (HIC) is often used in characterization of therapeutic antibody products due to its ability to separate direct or indirect structural changes in the studied protein. Scientists at Alexion have published a study where FabRICATOR (IdeS) was used to generate Fc and F(ab’)2 fragments of an antibody to study conformational changes of a monoclonal antibody (King et al. 2018).


Separation of the intact antibody on HIC reveled two major peaks that were collected and subjected to FabRICATOR digestion. After digestion, the Fc and F(ab’)2 fragments separated well, and the heterogeneity was localized to the F(ab’)2 domain. Variations in the Fc were observed and attributed to oxidation modifications. Peptide mapping of the domains were carried out and a 1 Da difference was localized , indicating deamidation of Asn to either Asp or isoAsp in the complementarity-determining region (CDR) of the light chain. The observed difference in HIC separation pattern was also linked to changes in antigen binding, since the deamidation of the Asn residues reduced the binding of the antibody to its target antigen.


Taken together, this paper indicates that a single deamidation in the light chain changed the hydrophobicity profile of the antibody and impacted the antigen binding. The use of FabRICATOR (IdeS) digestion and HIC separation could serve as a quick screening assay to study deamidation changes in the F(ab’)2 domain.



King, C. et al., 2018. Characterization of recombinant monoclonal antibody variants detected by hydrophobic interaction chromatography and imaged capillary isoelectric focusing electrophoresis. Journal of Chromatography B, 1085, pp.96–103.


New reference of FabRICATOR in antibody based PET imaging

February 23, 2016 | References |

To develop an antibody based PET imaging for detection of amyloid beta in mouse models, Sehlin and colleagues  at the Rudbeck Laboratory in Uppsala, Sweden, used Fabricator to generate F(ab’)2 fragments. In the study, published in Nature Communications, Sehlin et al. generated F(ab’)2 fragments using FabRICATOR of an amyloid beta specific antibody and conjugated this to a transferrin receptor antibody for delivery across the blood brain barrier. Read more »