Detection of catalytic monoclonal antibodies

Dan S. Tawfik*, Bernard S. Green, Zelig Eshhar

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

13 Scopus citations

Abstract

Several laboratories have now shown that monoclonal antibodies having enzyme-like properties can be generated. The generation of catalytic antibodies makes use of the same basic procedures that have been used for the generation of binding monoclonal antibodies, yet the process involves an additional crucial step: screening for catalytic activity. In this paper we address the unique problems involved in the detection of inefficient catalytic activity that is accompanied by uncatalyzed background reaction. An analysis that allows optimization of assay conditions and estimation of the minimal antibody concentration required to observe catalysis is presented. The results indicate that the structure of the substrate should be optimized to increase its affinity (i.e., decrease its Km) and reduce its concentration to pseudo-first-order conditions (So ≪ Km) so that the signal observed in the presence of a catalytic antibody (ΔPcat) is significantly higher than that of the background (ΔPuncat). Other factors involved in the screening procedures, e.g., sensitivity of the assay, solubility and reactivity of the substrate, and purity of the antibody preparation, are also discussed. The effect of these assay parameters on the ability to detect catalytic activity is demonstrated with p-nitrophenyl ester-hydrolyzing antibodies.

Original languageEnglish
Pages (from-to)35-39
Number of pages5
JournalAnalytical Biochemistry
Volume202
Issue number1
DOIs
StatePublished - Apr 1992

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