Functionalized diethylenetriaminetetraacetic acid-based metal chelates for chemical modification of proteins and small biomolecules

• Published in: Russian journal of bioorganic chemistry 2015-11, Vol.41 (6), p.607-616
• Main Authors: Kuprienko, O. S., Dubovskaya, L. V., Shabunya, P. S., Fatykhava, S. A., Sviridov, O. V.
• Format: Article
• Language: English
• Subjects: Biochemistry; Biomedical and Life Sciences; Biomedicine; Bioorganic Chemistry; Life Sciences; Organic Chemistry
• ISSN: 1068-1620; 1608-330X
• DOI: 10.1134/S1068162015060072

Diethylenetriaminetetraacetic acid-based bifunctional reagents containing a bound Eu 3+ ion and a reactive group capable of interacting with proteins or low-molecular-weight compounds have been synthesized. These reagents are intended for use in lanthanide immunofluorimetric assay with a dissociation-enhancing solution (DELFIA). A complexonate of Eu 3+ and N l -(2-aminoethylamide) of diethylenetriaminepentaacetic acid (DTPA) has been obtained by acylation of N -Boc-1,2-ethylenediamine with DTPA dianhydride with subsequent unblocking of the amino group and chelation of Eu 3+ ions. The metal chelate was used to synthesize conjugates with horseradish peroxidase and 17α-hydroxyprogesterone 3-( O -carboxymethyloxime). The conjugate of 17α-hydroxyprogesterone 3-( O -carboxymethyloxime) with a metal chelate has been shown to retain affinity for the respective antibodies in an immunochemical DELFIA system. A complexonate of Eu 3+ and DTPA N l -[2-( p -(succinimidylcarboxy)benzoylamino)ethylamide] has been obtained by acylation of the aminocomplexonate by p -phtalic acid di- N -succinimide ester and used to synthesize conjugates with human serum albumin and certain immunoglobulins G. The amount of Eu 3+ bound to the modified proteins was inferred from the fluorescence of the conjugates dissolved in a dissociation-enhancing solution. Fluorescence intensity, background fluorescence, sensitivity, and specificity achieved using the monoclonal antibody conjugates obtained according to the procedure proposed were sufficient for use of the conjugates in DELFIA.