TY - JOUR
T1 - Epitope mapping of nanobodies binding the Alzheimer's disease receptor SORLA
AU - Monti, Giulia
AU - Vincke, Cécile
AU - Lunding, Melanie
AU - Jensen, Anne Mette G
AU - Madsen, Peder
AU - Muyldermans, Serge
AU - Kjolby, Mads
AU - Andersen, Olav M
N1 - Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.
PY - 2023/9/20
Y1 - 2023/9/20
N2 - Reduced levels of the Sortilin-related receptor with A-type repeats (SORLA) in different brain regions as well as in the cerebrospinal fluid have been associated with Alzheimer's disease. Methods and reagents to develop reliable detection assays to quantify SORLA and its specific isoforms are therefore much needed. Nanobodies (Nbs) are unique biomolecules derived from the blood of camelids that display advantageous physicochemical and antigen affinity properties, making them attractive tools with great relevance to both diagnostic and therapeutic applications. Here, we purified and characterized eight Nbs that were isolated from the blood of an alpaca immunized with the recombinant extracellular domain of SORLA. The selected Nbs showed high affinity to SORLA in the low nanomolar range as observed by surface plasmon resonance. For mapping of the Nbs' epitopes within the antigen, we transiently transfected HEK293 cells with a panel of SORLA deletion constructs, and developed a protocol of immunostaining by applying fluorescent dye conjugated Nbs. With this method, we showed that the selected Nbs specifically recognize a part of SORLA containing Fibronectin-type III domains, representing promising tools not only for disease clarifying research, but also for translational medicine as candidates for clinical diagnostic purposes.
AB - Reduced levels of the Sortilin-related receptor with A-type repeats (SORLA) in different brain regions as well as in the cerebrospinal fluid have been associated with Alzheimer's disease. Methods and reagents to develop reliable detection assays to quantify SORLA and its specific isoforms are therefore much needed. Nanobodies (Nbs) are unique biomolecules derived from the blood of camelids that display advantageous physicochemical and antigen affinity properties, making them attractive tools with great relevance to both diagnostic and therapeutic applications. Here, we purified and characterized eight Nbs that were isolated from the blood of an alpaca immunized with the recombinant extracellular domain of SORLA. The selected Nbs showed high affinity to SORLA in the low nanomolar range as observed by surface plasmon resonance. For mapping of the Nbs' epitopes within the antigen, we transiently transfected HEK293 cells with a panel of SORLA deletion constructs, and developed a protocol of immunostaining by applying fluorescent dye conjugated Nbs. With this method, we showed that the selected Nbs specifically recognize a part of SORLA containing Fibronectin-type III domains, representing promising tools not only for disease clarifying research, but also for translational medicine as candidates for clinical diagnostic purposes.
UR - http://www.scopus.com/inward/record.url?scp=85169003768&partnerID=8YFLogxK
U2 - 10.1016/j.jbiotec.2023.08.005
DO - 10.1016/j.jbiotec.2023.08.005
M3 - Article
C2 - 37634829
VL - 375
SP - 17
EP - 27
JO - Journal of Biotechnology
JF - Journal of Biotechnology
SN - 0168-1656
ER -