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Updated: Jul 23, 2026
A Protocol for Phage Display and Affinity Selection Using Recombinant Protein Baits
Published on: February 16, 2014
Identification of a novel anti-ROR1 nanobody through phage display and its biochemical characterization
Li Kang1,2, Yingkui Dong1,2, Wanxue Wang2,3
1Institute of Physical Science and Information Technology, Anhui University, Hefei, Anhui, China.
Abstract:
In this study, we aimed to develop nanobodies targeting receptor tyrosine kinase-like orphan receptor 1 (ROR1) for cancer diagnosis and therapy. We immunized alpacas with ROR1, extracted RNA from their blood, and converted it to complementary DNA (cDNA) to amplify the VHH (variable domain of heavy-chain antibodies) sequence. This sequence was used to construct a phage library with a capacity of 8 ×108. Screening identified a high-affinity nanobody, HCAbs1, which binds effectively to ROR1. ELISA and surface plasmon resonance analyses revealed HCAbs1's binding affinities to ROR1 at 4.42 and 12.9 nM, respectively. Functional tests showed HCAbs1 could reduce extracellular signal-regulated kinase (ERK) phosphorylation levels induced by Wnt5a in ROR1-transfected cells. Our findings highlight the potential of HCAbs1 nanobodies in diagnosing and treating cancers through targeting ROR1.
Insights
Researchers developed HCAbs1 nanobodies targeting receptor tyrosine kinase-like orphan receptor 1 (ROR1) for cancer applications. These nanobodies show high affinity and functional activity, offering potential for cancer diagnosis and therapy.
Area of Science:
- Biotechnology
- Immunology
- Oncology
Background:
- Receptor tyrosine kinase-like orphan receptor 1 (ROR1) is implicated in various cancers.
- Targeting ROR1 presents a promising strategy for cancer diagnosis and therapy.
Purpose of the Study:
- To develop high-affinity nanobodies targeting ROR1.
- To evaluate the diagnostic and therapeutic potential of these nanobodies.
Main Methods:
- Immunization of alpacas with ROR1 and construction of a phage display library.
- Screening and characterization of ROR1-specific nanobodies, including HCAbs1.
- Assessment of binding affinity using ELISA and surface plasmon resonance, and functional evaluation in cell-based assays.
Main Results:
- A high-affinity nanobody, HCAbs1, was identified with nanomolar binding affinities to ROR1.
- HCAbs1 demonstrated functional activity by reducing Wnt5a-induced ERK phosphorylation in ROR1-transfected cells.
- The nanobody effectively binds to ROR1, indicating its potential for targeted applications.
Conclusions:
- HCAbs1 nanobodies are effective ROR1 binders with therapeutic potential.
- These nanobodies could serve as valuable tools for cancer diagnosis and treatment strategies.
- Targeting ROR1 with nanobodies offers a novel approach in oncology.
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