Epitope mapping of nanobodies binding the Alzheimer's disease receptor SORLA

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.

PY - 2023/9

Y1 - 2023/9

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.

KW - Alzheimer's disease

KW - Epitope mapping

KW - Nanobodies

KW - SORLA

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 - Journal article

C2 - 37634829

AN - SCOPUS:85169003768

SN - 0168-1656

VL - 375

SP - 17

EP - 27

JO - Journal of Biotechnology

JF - Journal of Biotechnology

ER -