Development Of An Immunoassay Based On Nano-magnetic Beads For The Rapid Detection Of Anthracene In Water

Polycyclic aromatic hydrocarbons (PAHs) mainly come from the pyrolysis or incompletecombustion of organics. They are considered to be persistent organic pollutants widely exists innatural environment with carcinogenicity and mutagenicity. In recent years, it reported that manyrivers, lakes and seas had been detected to contain PAHs pollutants with serious ecological risk.Currently, the main test methods of PAHs relies on instrumental detection, cellular detection andimmunological detection. Among them, the immunoassay with its high efficiency, sensitivity,accuracy, easy operation and many other advantages, has been increasingly applied to theexamination of PAHs in water.In this paper, anthracene (ANT), the research object, is a hapten with small molecular whichcould be used as complete antigen after coupling with bovine serum albumin (BSA) orOvalbumin (OVA). Using the ANT-BSA to immune Balb/c mice,2specific hybridoma cell line(D3and H8) were screened out after cell fusion between popliteal lymph node cells or spleencells from the immunized mice with SP2/0myeloma cells. The ascites gained by induce ascitesmethod were purified by affinity chromatography. The purified anti-ANT monoclonal antibodywas used to established enzyme label plate-indirect competitive enzyme-linked immunosorbentassay (ic-ELISA) and immunomagnetic bead (IBM)-ic-ELISA, and the relevant conditions of thetwo detection methods were optimized.The equation of linear regression with ANT in enzyme label plate-ic-ELISA method was Y=0.4253x-0.1883, and the correlation coefficient was0.9901with a linear detection range from4.76ng/mL to362.6ng/mL. It showed an IC50value of41.53ng/mL with a detection limit of3.653ng/mL. The Cross-reactivities of this McAb with other15PAHs were less than1%, exceptfor Pyrene (5.80%). The results of reproducibility showed that the average coefficient ofvariation was range from5.95%~12.86%. The recovery rate was between100.07%and111.08%with the detection of spiked sample. The detection of ANT in water samples from changchun, daqing, jinzhou, beijing is5.63ng/mL,13.78ng/mL,4.42ng/mL,4.65ng/mL, respectively.Tapwater was not checked out.The equation of linear regression with ANT in IBM-ic-ELISA method was Y=0.3128x+0.0383, and the correlation coefficient was0.9761with a linear detection range from1.575ng/mL to568.585ng/mL. It showed a detection limit of2.431ng/mL. The results of repro-ducibility showed that coefficient of variation was range from8.28%~13.44%. The recoveryrate was between101.59%and106.77%with the detection of spiked sample. The detection ofANT in above samples is4.28ng/mL,12.42ng/mL,4.17ng/mL,5.13ng/mL, respectively.