Boro, Robin C and Singh, K Vikas and Suri, C Raman (2009) Characterization of hapten-protein conjugates: antibody generation and immunoassay development for chlorophenoxyacetic acid pesticides. Journal of AOAC International, 92 (6). pp. 1773-9. ISSN 1060-3271
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The generation of specific and sensitive antibodies against small molecules is greatly dependent upon the characteristics of the hapten-protein conjugates. In this study, we report a new fluorescence-based method for the characterization of hapten-protein conjugates. The method is based on an effect promoted by hapten-protein conjugation density upon the fluorescence intensity of the intrinsic tryptophan chromophore molecules of the protein. The proposed methodology is applied to quantify the hapten-protein conjugation density for two different chlorophenoxyacetic acid pesticides, 2,4-dichlorophenoxyacetic acid (2,4-D) and 2,4-dichlorophenoxybutyric acid (2,4-DB), coupled to carrier protein. Highly sensitive anti-2,4-D and anti-2,4-DB antibodies were obtained using these well-characterized hapten-protein conjugates. The generated antibodies were used in an immunoassay format demonstrating inhibitory concentration (IC50) values equal to 30 and 7 ng/mL for 2,4-D and 2,4-DB, respectively. Linearity was observed in the concentration range between 0.1-500 nglmL with LODs around 4 and 3 ng/mL for 2,4-D and 2,4-DB, respectively, in standard water samples. The proposed method was successfully applied for the determination of the extent of hapten-protein conjugation to produce specific antibodies for immunoassay development against pesticides.
| Item Type: | Article |
|---|---|
| Additional Information: | Copyright of this article belongs to AOAC International |
| Subjects: | Q Science > QD Chemistry |
| Depositing User: | Dr. K.P.S.Sengar |
| Date Deposited: | 08 Dec 2011 19:31 |
| Last Modified: | 09 Dec 2011 06:30 |
| URI: | http://crdd.osdd.net/open/id/eprint/533 |
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