แนวคิดหลัก
A new antibody screening system that directly links antigen-binding function with the encoding gene enables efficient isolation of broadly reactive antibodies against influenza virus.
บทคัดย่อ
The authors developed a new antibody screening system that directly links the antigen-binding function of membrane-expressed immunoglobulins (Igs) with their encoding genes. This system utilizes a dual-expression vector and Golden Gate cloning to rapidly generate an Ig library and express membrane-bound Igs on mammalian cells.
The key highlights are:
- The dual-expression vector links the heavy and light chain genes, reducing the time and effort required for plasmid preparation.
- The membrane-bound Ig expression enables direct selection of antigen-binding clones through flow cytometry, bypassing the laborious steps of conventional cloning-based methods.
- The authors demonstrated the efficiency of this system by isolating broadly reactive antibodies against influenza virus hemagglutinin (HA) antigens from an experimental mouse model.
- Several of the isolated antibodies showed broad reactivity, binding to HA from different influenza strains, including group 1 and group 2 HAs, as well as the highly pathogenic H5N1 strain.
- One of the broadly reactive antibodies, A6p4, was found to compete with the classic broadly neutralizing antibody C179, suggesting it may target the conserved HA stem region.
- This technology can be applied to the rapid isolation of therapeutic and diagnostic antibodies, particularly during pandemics when timely development of countermeasures is crucial.
สถิติ
The affinities (Kd) of the isolated antibodies for different HA antigens ranged from 500 to 100 nM, with the highest affinity of 5.66 × 1010 M for the A/California/2009 (X-179A) [H1N1] Pdm09 strain.
Seven out of the nine isolated mAbs bound to the HA from the highly pathogenic avian influenza strain H5N1.
Six of the antibodies that bound to H3 HA were categorized as group 2 influenza viruses.
คำพูด
"Our technology can also be applied to human antibody screening and represents a new line of mAb screening that accelerates the isolation of therapeutic and diagnostic mAbs."
"Compared with droplet-based experimental systems, well-based systems are limited in the number of cells they can process. Furthermore, experiments involving infectious bacteria and viruses have imposed limitations on human experimentation. To solve these problems, the automation of experiments will become important in the future."