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Lateral flow immunostrips for the sensitive and rapid determination of 8-hydroxy-2′-deoxyguanosine using upconversion nanoparticles

Wu Nengying,Wei Yuxi,Pan Lanlan,Yang Xiaolin,Qi Honglan,Gao Qiang,Zhang Chengxiao

Lateral flow immunostrips were newly designed and a sensitive and rapid fluorometric method for the determination of 8-hydroxy-2′-deoxyguanosine (8-OHdG) as a model target of small biomarker molecules was developed. The upconversion nanoparticles (UCNPs, NaYF4:Yb/Er core, and polyacrylic acid (PAA)-modified shell, size ~ 39 nm, excitation wavelength = 980 nm; emission wavelength = 540 nm) were employed as fluorescence signal material. The 8-OHdG antibody (Ab) was taken as the recognition probe while UCNP-labeled Ab was taken as the signal probe. Bovine serum albumin (BSA) was designed as carrier protein for 8-OHdG to form 8-OHdG-BSA conjugate as the capture probe. The lateral flow immunostrips were prepared by laminating a sample pad (glass fiber membrane), a test pad (nitrocellulose membrane), and adsorption pad (filter paper) on PVP backing. The capture probe was immobilized on the test zone while an IgG antibody taken as the control probe was immobilized on the control zone. When the signal probe and the sample were in sequence loaded on the sample pad, 8-OHdG analyte bound with the signal probe, and then the excess of the signal probe move along the strip and is collected by the capture probe on the test zone while the remnant signal probe is collected by the control probe on the control zone. The signal probe and capture probe were synthesized and characterized. The fluorescence intensity on the test zone was inversely proportional to the concentration of 8-OHdG for the quantitative determination while the fluorescence emission on the control zone was observed to validate the assay. The developed method showed a wide linear range from 0.10 to 10 nM, a quite low detection limit of 0.05 nM, small sample volume requirement (100 μL), short assay time (15 min), and good method reproducibility (RSD = 4.4%, nine immunostrips). Schematic illustration of the configuration and measurement principle of lateral flow fluorescence immunostrip for 8-OHdG: (a) configuration; (b) preparation: load of capture probe (BSA-8-OHdG, 2 μL) on test zone; load of control probe (IgG Ab, 2 μL) on control zone; load of signal probe (UCNP-Ab, 16 μL) on sample pad; (c) measurement: load of sample (8-OHdG, 100 μL) on sample pad, collection, and measurement.