Feasibility of High-Performance Immunochromatography as an Isolation Method for PCBs and Other Dioxin-Like Compounds

Abstract

A high performance immunochromatographic procedure to isolate polychlorinated biphenyls (PCBs) and other dioxin-like compounds from a sample is shown. Development of the procedure includes (i) synthesis of the hapten, binding it to the spacer arm and to the carrier protein to make the immunizing molecule; (ii) raising and purification of anti-PCB antibodies; (iii) building of the immunocolumn; (iv) selection of the binding, rinsing, and elution conditions adequate for these highly lipophilic compounds; (v) study of the influence of the concentration and volume of sample on recovery; and (vi) study of the selectivity of the immunosystem for dioxins, furans, PCBs, and several insecticides of different toxicity. Evaluation of the method is carried out by analyzing the fractions retained and nonretained in the immunocolumn by GC/MS. The immunochromatographic system that is developed shows itself to be feasible as cleanup and isolation steps carried out prior to GC/MS analyses. When compared to classical cleanup and isolation methods traditionally used for analysis of PCBs in water, the immunochromatographic method is > 20x faster and uses 100x less organic solvents, and its selectivity is enormously enhanced. Good recoveries are obtained with both kinds of methods. The immunochromatographic procedure fulfils the acceptance criteria indicated by the EPA, even for sub-parts-per-billion concentrations.

Differences in retention of dioxin‐like compounds and organochlorinated insecticides on an immunochromatographic column. Interpretation and applicability

“…Synthesis of the PCB hapten and immunogen, raising of the polyclonal anti-PCB antibodies, as well as their immobilization onto epoxy-silica beads and packing into an empty stainless steel HPLC column (50 mm length62.1 mm ID) to prepare the high performance IAC (HPIAC) column were described previously [10].…”

“…PBS was used as mobile phase [10]. Twenty lL of working solution (WS1 or WS2) was injected through the valve V2 into the IAC column pre-equilibrated for 20 min with PBS.…”

“…One of these new techniques is immunochromatography. It has been applied in high performance [10] or low performance [11] systems as isolation, concentration, and clean-up steps for dioxin-like compounds prior to their gas chromatography-mass spectrometry determination (GC-MS).…”

“…In a recent work, we developed an immunoaffinity chromatography (IAC) column made with polyclonal antibodies raised against an antigen that mimicked the 3,39,4,49-tetrachloro substitution pattern and coplanarity characteristics of the most toxic PCBs [10]. We showed that this anti-PCB column was able to capture the most toxic PCB congeners.…”

“…C labeled compounds 98% purity). Their structures and that of the hapten synthesized in the laboratory [10] and used to raise antibodies are shown in Figure 1. For the sake of simplicity only one PCB of each chlorination degree is shown.…”