For the top one docking pose, SAL bound between the H chain and L chain, forming contacts with residues from CDR loops (Figure 2A). In addition, the selected scFv was fused with alkaline phosphatase and expressed in to develop a quick and low-cost one step ELISA to detect SAL. Keywords: phage display, docking, salbutamol, scFv, ELISA Graphical Abstract The flowchart for discovery of anti-SAL scFvs. By using molecular docking approach and phage display, anti-SAL scFvs with high affinity were recognized and fused with alkaline phosphatase for one-step ELISA salbutamol detection. 1. Introduction Salbutamol (SAL) is usually a 2 adrenergic receptor agonist, which is usually widely used to treat bronchial asthma (Price and Clissold, 1989). In the mean time, it can promote protein synthesis, increase animal lean meat rate, and improve feed conversion rate. S-Gboxin It is often illegally used as a feed additive in animal husbandry (Baker et al., 1984; Dalrymple et al., 1984; S-Gboxin Jones et al., 1985). Excessive intake of SAL can cause myalgia, headache, dizziness, nervousness, tachycardia, nausea, vomiting, and even cause liver and kidney damage, and its residues pose a serious hazard to human health (Wang and Shen, 2007; Khamta et al., 2009; Sheu et al., 2009). Therefore, SAL has been purely banned as a feed additive by many countries, but due to its economic incentives, many farms still use SAL extensively (Kearns et al., 1985; Garssen et al., 1995). Illegal addition of SAL can cause environmental pollution and affect public health via the food chain (Wang et al., 2015). Studies have shown that SAL has the possibility of entering the ecological environment through animal feces and urine. While causing environmental pollution, it then enters the human body through indirect channels (Fang et al., 2019). SAL has already been a common environmental pollutant (Depaolini et al., 2016). At present, SAL residues have been found in natural waters around the world, including tap water, wastewater, treated sewage, and river water (Yamini et al., 2006; Lei et al., 2015a). Even though concentration of SAL in some water bodies has reached 470 ng/L (Bound and Voulvoulis, 2006), you will find few reports focusing on environmental problems caused by SAL (Liu et al., 2018). Therefore, it is imperative to establish a sensitive method to monitor SAL. The analytical methods currently used to detect SAL include gas chromatographyCmass spectrometry (GC-MS) (Black and Hansson, 1999), high-performance liquid chromatography (HPLC) (Rosales-Conrado et al., 2013), and high-performance liquid chromatographyCmass spectrometry (HPLC-MS) (Zhang et al., 2012). Because these methods require cumbersome sample preparation before instrumental analysis (Liu Z. J. et al., 2016), it is difficult to meet the requirements for high-throughput and quick screening of a large number of environmental samples. Immunoassay is a fast, low-cost, and Rabbit Polyclonal to FGFR1 Oncogene Partner high-throughput method, and it is becoming a reliable tool for the analysis of environmental pollutant residues. So far, many immunoassays for S-Gboxin detecting SAL have been successfully developed. Among them, ELISA is the commonly used method for SAL detection (Degand et al., 1993; Lei et al., 2008, 2015b). Chemiluminescence and electrochemiluminescence assay, time-resolved immunofluorescence technique, and lateral chromatography technique (colloidal platinum) have been developed for SAL and other -agonist detection (Cai et al., 2015; Xu et al., 2015, 2022; Liu B. et al., 2016; Li et al., 2017; Gu et al., 2020). Immunoassay methods also have some problems. For example, most of the currently available anti-SAL antibodies are polyclonal antibodies from sheep and rabbits (Degand et al., 1993; Lei et al., 2008; Wu et al., 2014), and their specificity is usually poor. For polyclonal antibodies, the heterogeneity of antibody preparations usually prospects to cross-reactions with highly comparable antigens. For example, polyclonal anti-SAL antibody shows significant cross-reactivity to clenbuterol (Degand et al., 1993; Lei et al., 2008; Wu et al., 2014). Compared with polyclonal antibodies, monoclonal antibodies have the advantages of high specificity, and a relatively simple production process has been widely used in immunoassays. Adam et al. obtained an anti-salbutamol antibody by immunizing mice. S-Gboxin