Version 1
: Received: 27 December 2023 / Approved: 28 December 2023 / Online: 28 December 2023 (10:44:34 CET)
How to cite:
Segati, A.F.; Da Costa, H.H.M.; Prudencio, C.R.; Portilho, A.I.; De Gaspari, E. Evaluation of IgG Avidity of Sera from Mice Immunized with RBD of SARS-CoV-2 or OMVs of Neisseria meningitidis by Immunoblot and Dot-Blot. Preprints2023, 2023122180. https://doi.org/10.20944/preprints202312.2180.v1
Segati, A.F.; Da Costa, H.H.M.; Prudencio, C.R.; Portilho, A.I.; De Gaspari, E. Evaluation of IgG Avidity of Sera from Mice Immunized with RBD of SARS-CoV-2 or OMVs of Neisseria meningitidis by Immunoblot and Dot-Blot. Preprints 2023, 2023122180. https://doi.org/10.20944/preprints202312.2180.v1
Segati, A.F.; Da Costa, H.H.M.; Prudencio, C.R.; Portilho, A.I.; De Gaspari, E. Evaluation of IgG Avidity of Sera from Mice Immunized with RBD of SARS-CoV-2 or OMVs of Neisseria meningitidis by Immunoblot and Dot-Blot. Preprints2023, 2023122180. https://doi.org/10.20944/preprints202312.2180.v1
APA Style
Segati, A.F., Da Costa, H.H.M., Prudencio, C.R., Portilho, A.I., & De Gaspari, E. (2023). Evaluation of IgG Avidity of Sera from Mice Immunized with RBD of SARS-CoV-2 or OMVs of Neisseria meningitidis by Immunoblot and Dot-Blot. Preprints. https://doi.org/10.20944/preprints202312.2180.v1
Chicago/Turabian Style
Segati, A.F., Amanda Izeli Portilho and Elizabeth De Gaspari. 2023 "Evaluation of IgG Avidity of Sera from Mice Immunized with RBD of SARS-CoV-2 or OMVs of Neisseria meningitidis by Immunoblot and Dot-Blot" Preprints. https://doi.org/10.20944/preprints202312.2180.v1
Abstract
Several methods can be used to evaluate antibody avidity, one of which is the modified ELISA, in which a chaotropic agent is added to disrupt the binding between the antigen and the antibody. Although the avidity index provides relevant information about the functionality of antibodies, it does not allow us to characterize which antigens the antibodies are strongly bound to. To this end, we adapted the Immunoblot and Dot-Blot techniques by adding potassium thiocyanate (KSCN) after incubating nitrocellulose membranes with serum from mice immunized with Neisseria meningitidis outer membrane vesicles (OMVs) or serum from mice immunized with recombinant receptor-binding domain (RBD) of SARS-CoV-2, with different adjuvants for each antigen. In terms of results, when standardizing the incubation time of sera with KSCN, we observed that 5 minutes was the ideal time for both assays, however, when using different dilutions of serum from mice immunized with RBD, there was an influence on the strength of antigen-antibody binding. For the N. meningitidis assay, the antibodies maintained high avidity for the antigens present in the OMVs, PorA, NadA, and Opa, especially in the presence of the aluminum hydroxide adjuvant. The research showed that standardizing the Immunoblot and Dot-Blot in the same assay optimizes time and resources, allowing qualitative analysis for two different antigens, and resulting in more robust information for immunization studies against these pathogens.
Biology and Life Sciences, Immunology and Microbiology
Copyright:
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
,
