Single-domain antibody screening by isPLA-seq
Yueyuan Yin1, Fei Yan1, Ruimin Zhou1
1Tianjin Key Laboratory of Medical Epigenetics, Key Laboratory of Immune Microenvironment and Disease (Ministry of Education), Department of Immunology, Biochemistry and Molecular Biology, School of Basic Medical Sciences, Tianjin Medical University, Tianjin, China.
Life Science Alliance
|October 22, 2021
Summary
We developed a high-throughput method combining in situ proximity ligation assay (isPLA) and sequencing (isPLA-seq) to screen single-domain antibody (sdAb) libraries for specific proteins at the single-cell level.
Area of Science:
- Biotechnology
- Molecular Biology
- Immunology
Background:
- Single-domain antibodies (sdAbs) offer versatile strategies for research and translational applications.
- Developing high-throughput methods for sdAb screening is crucial for identifying novel binders.
Purpose of the Study:
- To adapt and validate the in situ proximity ligation assay (isPLA) coupled with sequencing (isPLA-seq) for high-throughput screening of sdAb libraries.
- To enable the identification of sdAbs targeting specific proteins at subcellular and single-cell resolution.
Main Methods:
- Adaptation of the in situ proximity ligation assay (isPLA) for sdAb library screening.
- Integration of sequencing (isPLA-seq) for high-throughput analysis of sdAb-protein interactions.
- Recombinant sdAb production based on complementarity-determining region 3 (CDR3) sequences.
Main Results:
- Demonstrated a method for screening sdAb libraries with high sensitivity and throughput.
- Enabled the characterization of sdAbs at subcellular and single-cell resolution.
- Facilitated the identification of functional sdAbs, their epitopes, and potential applications.
Conclusions:
- The developed isPLA-seq method provides a general approach for identifying functional sdAbs.
- This technique allows for the characterization of sdAb-epitope interactions and investigation of cellular processes.
- The method supports diverse research and translational applications of sdAbs.
