Antibodies

Recombinant antibodies within the ULTRA-DD project

Within work package 2 of the ULTRA-DD project one of the aims is to generate recombinant antibodies to selected target proteins to be used as research tools. A subset of these antibodies will be declared biological probes. These antibodies are targeting proteins present in the extracellular matrix and must be shown to have an effect on live cells in an assay format or to have a diagnostic use by detection of specific proteins in biological samples. The best antibody for each target protein will be made available through disclosing the sequence here. Selected ones will also be available through distributors, either in scFv or IgG format.

Antibody generation method

A core aim of the SGC labs has always been the ability to produce high-quality proteins. Over the years, SGC labs have been able to produce thousands of expression clones and protocols for medically relevant proteins. From this gold mine we can now cherry pick the proteins of highest importance to our partner projects and use them as antigens for antibody generation. The SGC KI team is collaborating closely with the Drug Discovery and Development platform (DDD) at the Science for Life Laboratory in Stockholm. DDD contributes with state-of-the-art combinatorial libraries of human antibody candidates where high-quality binders can be obtained through phage display technology. In contrast to traditionally used methods to generate antibodies that rely on immunizations of animals, phage display is animal-free and yields recombinant antibodies, which are completely renewable and can be converted to any desired format depending on downstream application.

Antibody validation method

Selected antibody candidates are first validated for their binding against their antigen using various immune-based assays in 96-well format. Binders that pass this initial validation are then subjected to cell-based validation, testing whether they can bind their target antigens produced endogenously in a human cell. This is done via immunoprecipitation followed by mass spectrometry, or IP-MS for short. The strength of this method is in its ability to detect not only the target protein that the antibody pulls down, but also its protein complex partners, and potential cross-reactivity targets. Compared to other traditional methods like western blot where you see only a band on a gel, IP-MS also provides the identity of each protein in the immunoprecipitate.

Antibody validation criteria:

• Binds its target antigen with an affinity of at least 20 nM
• Can immunoprecipitate its endogenous target protein from HEK293 cell lysate (or other relevant cell line or biological sample)
• Specific in IP-MS, or at least that any cross-reactivity is known

Additional for biological probes:

• Functional activity towards a target in the extracellular space measured in a cell-based assay
• or
• Diagnostic detection of proteins in biological samples

The table can be downloaded as an XLSX / CSV.
The single all-sequence FASTA for all entries can be downloaded here.