Streptavidin-biotinylated IgG conjugates: a simple procedure for reducing polymer formation

Renato B. Del Rosario, Lee Ann Baron, Richard G. Lawton, Richard L. Wahl

Research output: Contribution to journalArticle

Abstract

Disulfide links of the IgG2ak anti-ovarian carcinoma antibody, 5G6.4, were site-specifically biotinylated [≈2 biotins/ IgG2a] using a novel crosslinking procedure using the biotin derivatized ETAC (equilibrium transfer alkylation crosslink reagent) 1a. Complexation of ETAC 1a biotinylated 5G6.4 on a column of immobilized protein A at high dilution, followed by passage of [125I]streptavidin, washing and pH change leads to elution of a streptavidin-free product with a molecular mass in the 200-300 kDa range. By contrast, direct mixing with [125I]streptavidin rapidly gave larger oligomers of ≫669 and ≈440-669 kDa molecular mass, respectively. The biodistribution of the 200-300 kDa complex showed significantly diminished liver, kidney and spleen uptake as well as higher blood activity than the 440-669 kDa complex. The methodology represent the first application of ETAC chemistry to disulfide-bond directed biotinylation of antibodies and the synthesis of streptavidin antibody conjugates which minimizes their polymerization.

Original languageEnglish (US)
Pages (from-to)417-421
Number of pages5
JournalInternational Journal of Radiation Applications and Instrumentation.
Volume19
Issue number3
DOIs
Publication statusPublished - 1992
Externally publishedYes

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