A chemiluminescence flow immunosensor based on a porous monolithic metacrylate and polyethylene composite disc modified with protein G.
A generic, fast, sensitive and new type of flow immunosensor has been developed. The basis is a monolithic porous poly(glycidyl methacrylate-co-trimethylolpropane trimethacrylate) polymer disc modified with protein G, placed in a fountain type flow cell compartment, in close proximity to a photomultiplier tube (PMT). Analyte and HRP labelled analyte derivative (tracer) compete for anti-analyte antibody binding sites. The mixture is then injected into the flow immunosensor system where the formed analyte- and tracer-antibody complexes are trapped by the monolithic protein G disc. The amount of bound tracer, inversely related to the concentration of analyte in the sample, is determined in a second step by injection of luminol, p-iodophenol and H2O2, generating enhanced chemiluminescence (CL) with horseradish peroxidase (HRP). A third and final step is need for regeneration of the protein G disc so that a new analysis cycle can take place. The performance of the disc immunosensor system was compared with a one step continuous flow injection immunoassay (FIIA) system, using the same reagents and a protein G column, in terms of assay sensitivity and influence of matrix effects from various water samples (millipore-, tap- and surface water). The detection limit for the analyte atrazine in PBS and surface water (SW) was 0.208 +/- 0.004 microg l(-1) (PBS) and 0.59 +/- 0.120 microg l(-1) (SW) for the FIIA and 0.033 +/- 0.003 microg l(-1) (PBS) and 0.038+/-0.003 microg l(-1) (SW) for the disc immunosensor. Statistical comparison of the two systems shows that the disc immunosensor results were significantly less influenced by the sample matrix, which is explained by the fact that the sample in the FIIA arrives simultaneously with the matrix to the detector, whereas these are separated in time in the disc immunosensor system.
Jain
Seema Rani
SR
Department of Analytical Chemistry, Lund University, PO. Box 124, Getingevägen 60, SE-221 00 Lund, Sweden.
Borowska
Ewa
E
Davidsson
Richard
R
Tudorache
Madalina
M
Pontén
Einar
E
Emnéus
Jenny
J
D000327:Adsorption; D001280:Atrazine / Q000032:analysis*; D004784:Environmental Monitoring / Q000295:instrumentation / Q000379:methods; D004867:Equipment Design; D019544:Equipment Failure Analysis; D017022:Flow Injection Analysis / Q000295:instrumentation* / Q000379:methods; D007118:Immunoassay / Q000295:instrumentation* / Q000379:methods; D008163:Luminescent Measurements; D008420:Manufactured Materials; D008567:Membranes, Artificial; D008768:Methylmethacrylates / Q000737:chemistry*; D009419:Nerve Tissue Proteins / Q000737:chemistry*; D010783:Photometry / Q000295:instrumentation* / Q000379:methods; D016062:Porosity; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014159:Transducers; D014867:Water / Q000737:chemistry*; D014874:Water Pollutants, Chemical / Q000032:analysis
C029152:G-substrate; D008567:Membranes, Artificial; D008768:Methylmethacrylates; D009419:Nerve Tissue Proteins; D014874:Water Pollutants, Chemical; C491289:poly(glycidyl methacrylate-co-trimethylolpropane trimethacrylate); D014867:Water; D001280:Atrazine