Protocol for evaluation and validation of TLR8 antagonists in HEK-Blue cells via secreted embryonic alkaline phosphatase assay

• DOI: 10.1016/j.xpro.2021.101061
• Publication Date: 2021-12-23T02:12:35Z
• Authors: Zhenyi Hu; Tuan Zhang; Shuangshuang Jiang; Hang Yin
• Keywords: 0303 health sciences; Science (General); Alkaline Phosphatase; High Throughput Screening; Cell Line; 3. Good health; Chemistry; Q1-390; 03 medical and health sciences; Molecular/Chemical Probes; Toll-Like Receptor 8; Protocol; Humans; Cell culture; Cell-based Assays

Toll-like receptor 8 (TLR8) is a pattern recognition receptor that senses RNA degradation products and initiates immune responses. TLR8 overactivation is associated with autoimmune diseases. Herein, we describe the evaluation and validation of TLR8 antagonists in HEK-Blue cells via secreted embryonic alkaline phosphatase (SEAP) assay, WST assay, ITC and immunoblotting. These assays can facilitate the development of TLR8 antagonists; this protocol can also be adapted to analyze agonists and antagonists for other TLRs. For complete details on the use and execution of this protocol, please refer to Hu et al. (2018).