Journal
RSC ADVANCES
Volume 8, Issue 3, Pages 1511-1518Publisher
ROYAL SOC CHEMISTRY
DOI: 10.1039/c7ra12810j
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- Central Forensic Science Laboratory (CFSL), Chandigarh, India
- Department of Science and Technology (DST), New Delhi, India [ECR/000075]
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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 (V-H)- and light (V-L)-chain genes were connected via a glycine-serine linker isolated from murine immune repertoire and cloned into the expression vector pIT2. The scFv was produced in Escherichia coli 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(-1) with a limit of detection (LOD) of 5 ng mL(-1) under optimal conditions, and the IC50 value was 14 ng mL(-1) 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.
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