In an initial study (Michael Schoof and coll.), 21 nanobodies binding to the spike protein were identified. There are 2 types: class 1 nanobodies, binding only with the ACE2 cell receptor binding domain of the spike protein, and class 2 nanobodies, not only binding to this domain but targeting other epitopes of the spike protein. Class 1 nanobodies are able to block spike-ACE2 binding entirely, unlike class 2, which allow only a moderate reduction in binding. It was also shown in this study that multimerising these nanobodies (binding two or three nanobodies) enabled greater efficiency in binding to the spike protein, thereby increasing their capacity to neutralize entry of the viral particle. Moreover, coupling the two categories of nanobodies enabled increased effectiveness in neutralizing the virus. Finally, the nanobodies’ stability means that they can be freeze-dried and administered using aerosols.
In a second study (Yufei Xiang and coll.), researchers immunized llamas with the region of the SARS-CoV-2 spike protein binding to the cell receptor. The serum of these llamas effectively neutralized the entry of viral particles into cells. Scientists analyzed the presence of specific nanobodies against the spike protein, in addition to more conventional neutralizing antibodies. After different tests on virus neutralization, 14 nanobodies were chosen to be used in complementary tests, notably physico-chemical tests, which confirmed the usefulness of nanobodies in therapeutic applications. It would seem that these nanobodies bind with the S1 region of the spike protein and block it in its conformation, making the binding domain to the cellular receptor inaccessible.