Abstract

Optical assays are of significant interest that can be examined easily with change in the color of gold nanoparticle (AuNPs). We report the development of AuNPs based dipstick assay for the detection of morphine using single-chain fragment variable (scFv) antibody. Phage display-based antibody library was developed for morphine. Variable regions of heavy (VH)- and light (VL)-chain genes of immunoglobulins were connected via a glycine–serine linker isolated from murine immune repertoire and cloned into the expression vector pIT2. The scFv was produced approximately 32 kDa yielding a functional protein in Escherichia coli HB2151. Anti-morphine scFv was conjugated with AuNPs to develop an optical immunochromatographic dipstick assay. The developed dipstick assay showed the limit of detection (LOD) of 5 ng/mL with IC50 value 14 ng/mL for morphine. The developed dipstick kit was capable of specifically recognize and bind to free morphine and its analogs in less than 5 min and could be useful for on-site screening of a real sample in blood, urine, and saliva.