A new methodology toward biodefense and threat surveillance by water-dispersible, bactericidal, paramagnetic nanoparticles is presented. The nanoparticles consist of magnetite clusters (approximately 70 nm) modified by polyethyleneimine (PEI) and poly(hexamethylene biguanide) (PHMBG) and are prepared by a two-step procedure at an acceptable cost. The cationic nanoparticles are colloidally stable in water at pH < or = 10, where they sequester DNA or whole microbes. The nanoparticles and bound DNA are captured by high-gradient magnetic separation. Methods of efficient extraction and quantification of the DNA by real-time PCR are developed. Broad bactericidal activity of the nanoparticles is demonstrated at concentrations far below cytotoxicity levels for mammalian cells. The levels of the DNA detection sensitivity obtained in our experiments allow us to project the applicability of the developed method for the detection of DNA molecules of various germs.

Conjugates of linear and branched polyethyleneimine (PEI) and monoamine polyether Jeffamine M-2070 (PO/EO mol ratio 10/31, 2000 Da) were synthesized through polyether activation by cyanuric chloride followed by attachment to PEI and guanidinylation by 1H-pyrazole-carboxamidine hydrochloride. The resulting guanidinylated PEI-polyether conjugates (termed gPEI-Jeffamine) efficiently complexed plasmid DNA, and their polyplexes possessed enhanced colloidal stability in the presence of serum proteins. In vitro studies with mammalian CHO-1, 3T3, and Cos-7 cell lines demonstrated improved transfection efficiency of the pCMVbeta-gal plasmid/gPEI-Jeffamine polyplexes. The guanidinylation of the amino groups of PEI and the conjugation of PEI with the Jeffamine polyether enhanced the conjugates' interaction with genetic material and reduced the cytotoxicity of the polyplexes in experiments with the L929 cell line.