A gold nanoparticle-single-chain fragment variable antibody as an immunoprobe for rapid detection of morphine by dipstick.

Gold nanoparticle (AuNP)-based optical assays are of significant interest since the molecular phenomenon can be examined easily with change in the color of AuNPs. Herein, we report the development of a dipstick using a AuNP-labeled single-chain fragment variable (scFv) antibody for the detection of morphine. The scFv antibodies for morphine were developed using phage display-based antibody library. Immunoglobulin variable regions of heavy ( )- and light ( )-chain genes were connected a glycine-serine linker isolated from murine immune repertoire and cloned into the expression vector pIT2. The scFv was produced in HB2151, yielding a functional protein with a molecular weight of approximately 32 kDa. The morphine scFv was labeled with gold nanoparticles and used as an optical immunoprobe in a dipstick. The competitive dipstick assay characterized the ability of the scFv antibody to recognize free morphine. The detection range was 1-1000 ng mL with a limit of detection (LOD) of 5 ng mL under optimal conditions, and the IC value was 14 ng mL for morphine. The developed optical dipstick kit of scFv antibody was capable of specifically binding to free morphine and its analogs in a solution in less than 5 min and could be useful for on-site screening of a real sample in blood, urine, and saliva.