Lancaster Environment Centre, Lancaster University, Lancaster, UK.
Methods Mol Biol. 2024;2790:405-416. doi: 10.1007/978-1-0716-3790-6_21.
Antibodies are a valuable research tool, with uses including detection and quantification of specific proteins. By using peptide fragments to raise antibodies, they can be designed to differentiate between structurally similar proteins, or to bind conserved motifs in divergent proteins. Peptide sequence selection and antibody validation are crucial to ensure reliable results from antibody-based experiments. This chapter describes the steps for the identification of peptide sequences to produce protein- or isoform-specific antibodies using recombinant technologies as well as the subsequent validation of such antibodies. The photosynthetic protein Rubisco activase is used as a case study to explain the various steps involved and key aspects to take into consideration.
抗体是一种有价值的研究工具,可用于检测和定量特定蛋白质。通过使用肽片段来产生抗体,可以设计它们来区分结构相似的蛋白质,或者结合在不同蛋白质中保守的模体。肽序列的选择和抗体的验证对于确保基于抗体的实验的可靠结果至关重要。本章描述了使用重组技术鉴定产生蛋白质或同工型特异性抗体的肽序列的步骤,以及随后对这些抗体的验证。光合蛋白 Rubisco 激活酶被用作案例研究来解释所涉及的各个步骤和需要考虑的关键方面。