FabALACTICA® in a Quantitative IgG1 Clone Profiling

September 23, 2021 | Applications, References |

Our immune system protects us from pathogens and various diseases, and the immune response is dependent on a person’s repertoire of immune cells, antibodies and circulating plasma proteins. A detailed characterization of these plasma components is important for the understanding of how they affect each individual’s immune response.

Immunoglobulins are among the most important molecules in the human immune system and they are produced by plasma cells. Although humans can produce billions of IgG1 variants through recombination and hypermutation, the diversity of IgG1 clones in human blood plasma has long evaded direct characterization.
In this paper, an efficient and sensitive approach for quantitative IgG1 clone profiling is described. Several mass spectrometry-based methods were combined to enable personalized IgG1 profiling in plasma, and this approach was applied to a sample set of two healthy control donors and eight critically ill patients.
To map out and monitor the nature of the plasma IgG1 repertoire, a mass spectrometry-based Fab profiling was performed. All intact IgG were captured using affinity beads before they were subjected to digestion using the human IgG1-specific cysteine protease FabALACTICA (IgdE). This enzyme generates intact Fab and Fc fragments suitable for middle-level LC-MS analysis by digesting human IgG1 at a single site above the hinge.

By combining protein-centric (middle-down) and peptide-centric (bottom-up) approaches, it was revealed that the circulating IgG1 repertoire in human plasma is dominated by a limited number of distinct clones in healthy donors and septic patients, and that each donor’s IgG1 repertoire is unique. Also, it was observed that the IgG1 repertoire adapts to changes in physiology, such as septic episodes. Taken together, the authors show that individual plasma IgG1 clones can be identified, quantified and fully sequenced by using a combination of top-down, middle-down and bottom-up proteomics.


Bondt et al., 2021. Human plasma IgG1 repertoires are simple, unique, and dynamic. Cell Systems. https://doi.org/10.1016/j.cels.2021.08.008









FabALACTICA – Above hinge digestion of human IgG1