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Reshaping nanobodies for affinity purification on protein a
- Source :
- New Biotechnology. 57:20-28
- Publication Year :
- 2020
- Publisher :
- Elsevier BV, 2020.
-
Abstract
- Nanobodies (Nbs) are 15 kDa recombinant, single-domain, antigen-specific fragments derived from heavy-chain only antibodies (HCAbs) occurring naturally in species of Camelidae. The beneficial properties of Nbs make them suitable tracers for diagnostic and therapeutic purposes. Whereas Nbs with a terminal hexa-histidine tag (His-tag) are easily purified via immobilized metal affinity chromatography, previous studies revealed a negative impact of the His-tag on the biodistribution of Nb-based tracers. Thus, it is important to develop alternative purification methods for Nbs without a His-tag. Protein A (SpA), a surface protein of Staphylococcus aureus, binds the Fc-region of IgG molecules and also to a lesser extent human heavy chain family-3 variable (VH) regions. Nbs also belong to this VH family, although many fail to be recognized by SpA. Here it is demonstrated that non-SpA binding Nbs can be mutagenized for purification by SpA affinity chromatography and that these Nb variants retain their thermostability and antigen affinity, while biodistribution remains unaffected.
- Subjects :
- 0106 biological sciences
Staphylococcus aureus
Biodistribution
Protein A
Bioengineering
Affinity chromatography
medicine.disease_cause
01 natural sciences
Chromatography, Affinity
law.invention
03 medical and health sciences
Antigen
law
010608 biotechnology
medicine
Staphylococcal Protein A
Molecular Biology
030304 developmental biology
Thermostability
Medicine(all)
0303 health sciences
biology
Chemistry
food and beverages
General Medicine
Single-Domain Antibodies
Biochemistry
embryonic structures
Nanobody
biology.protein
Recombinant DNA
Antibody
Biotechnology
Subjects
Details
- ISSN :
- 18716784
- Volume :
- 57
- Database :
- OpenAIRE
- Journal :
- New Biotechnology
- Accession number :
- edsair.doi.dedup.....e9de075c06f19c84f02c01d080a2719e