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United States Department of Agriculture

Agricultural Research Service

Title: Comparison of Immunoaffinity Column Retention Patterns of Pcdds/pcdfs on Columns Generated with Different Monoclonal Antibody Clones and Polyclonal Antibodies Produced Using Different Hapten Conjugates.

item Shelver, Weilin
item Shen, Guomin - UC-DAVIS
item Gee, Shirley - UC-DAVIS
item Stanker, Larry - USDA ARS WEST REG RES CTR
item Hammock, Bruce - UC-DAVIS

Submitted to: Analytica Chimica Acta
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: January 2, 2002
Publication Date: February 2, 2002
Citation: Shelver, W.L., Shen, G., Gee, S.I., Stanker, L.H., Hammock, B.D. 2002. Comparison of immunoaffinity column recovery patterns of polychlorinated dibenzo-p-dioxins/polychlorinated dibenzofurans on columns generated with different monoclonal antibody clones and polyclonal antibodies. Analytica Chimica Acta 457:199-209.

Interpretive Summary: Analysis of the contaminant, polychlorinated dibenzo-p-dioxins (PCDDs), is very expensive and time consuming. The high costs prevent comprehensive surveys to ensure food safety in order to protect the general public. Our laboratory, with collaboration with Dr. Stanker and Dr. Hammock's group, has developed a procedure whereby antibodies produced by mice and goat are used to specifically bind the dioxins in bovine serum samples. This procedure allows the analysis to be carried out by a shorter, faster that reduces the use of toxic solvent. The overall results will be a decrease in cost and a faster turnaround time for the analysis of dioxins.

Technical Abstract: Evaluation of immunoaffinity columns (IACs) for dioxin serum sample cleanup requires a determination of the recovery of various congeners. Comparison of the IAC performance of anti-dioxin antibodies derived from various sources revealed different recovery patterns. Some of the recovery patterns could be rationalized by the design of the hapten used to produce the antibodies, but others seem to be related to differences in antibody function (binding). Each monoclonal clone produced a unique antibody molecule different from other monoclonal clones even though they may have similar gross binding properties. Polyclonal antibodies produced in different animals and antisera from different bleeds of the same animal have different antibody constituents. In general, the polyclonal antibody based IACs evaluated had lower recovery for furans and highly chlorinated dioxin congeners, but were more specific toward 2,3,7,8-TCDD. The resemblance of the hapten to 2,3,7,8-TCDD appeared to played a clear role, but as long as the hapten had the chlorines in the 2, 3, and 7 positions, recovery from the IAC was moderately good. Recovery of 2,3,7,8-TCDD from the IAC correlated roughly with the affinity constant (Ka) as determined by accelerator mass spectrometry (AMS) or with an IC50 determined from an ELISA analysis. Finally, IACs prepared from four monoclonal antibodies (Mabs) each from a different mouse but each derived from a common hapten showed differences in their retention patterns of PCDDs/PCDFs. Monoclonal antibody DD3, appeared to be the most broad-spectrum of the Mabs, and showed good recoveries of a number of furan and dioxin congeners.

Last Modified: 4/22/2015
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