Dendrimer-Activated Solid Supports for Nucleic Acid and Protein Microarrays

Abstract

The generation of chemically activated glass surfaces is of increasing interest for the production of microarrays containing DNA, proteins, and low-molecular-weight components. We here report on a novel surface chemistry for highly efficient activation of glass slides. Our method is based on the initial modification of glass with primary amino groups using a protocol, specifically optimized for high aminosilylation yields, and in particular, for homogeneous surface coverages. In a following step the surface amino groups are activated with a homobifunctional linker, such as disuccinimidylglutarate (DSG) or 1,4-phenylenediisothiocyanate (PDITC), and then allowed to react with a starburst dendrimer that contains 64 primary amino groups in its outer sphere. Subsequently, the dendritic monomers are activated and crosslinked with a homobifunctional spacer, either DSG or PDITC. This leads to the formation of a thin, chemically reactive polymer film, covalently affixed to the glass substrate, which can directly be used for the covalent attachment of amino-modified components, such as oligonucleotides. The resulting DNA microarrays were studied by means of nucleic acid hybridization experiments using fluorophor-labeled complementary oligonucleotide targets. The results indicate that the novel dendrimer-activated surfaces display a surface coverage with capture oligomers about twofold greater than that with conventional microarrays containing linear chemical linkers. In addition, the experiments suggest that the hybridization occurs with decreased steric hindrance, likely a consequence of the long, flexible linker chain between the surface and the DNA oligomer. The surfaces were found to be resistant against repeated alkaline regeneration procedures, which is likely a consequence of the crosslinked polymeric structure of the dendrimer film. The high stability allows multiple hybridization experiments without significant loss of signal intensity. The versatility of the dendrimer surfaces is also demonstrated by the covalent immobilization of streptavidin as a model protein.

A comparison of mono and multivalent linkers and their effect on the colloidal stability of nanoparticle and immunoassays performance

“…This dye has a quantum efficiency of 23 %, a molar absorptivity of 187,000 L mol -1 cm -1 and fluorescence excitation and emission wavelengths of 672 nm and 694 nm, respectively, in isopropanol [28].…”

“…Multivalent dendrimers are attracting a great deal of interest for applications in biomedical diagnostics [27][28][29][30], pharmacology / drug delivery [31,32] [33], cancer research [34], and nanomedicine [35]. Their use in research is still unabated now, 30 years after their first reported synthesis by Vogtle [36].…”

Flame treatment of low-density polyethylene: Surface chemistry across the length scales

“…Fluorescent labeling, on the other hand, is a promising method because of its high sensitivity, ease of operation and in situ applicability. It has been widely used in biological applications [46,47], polymer chemistry [48][49][50][51], and study of self assembled monolayers [52][53][54][55].…”

Diels–Alder Ligation of Peptides and Proteins

“…The presence of the dendrimeric interface leads to amplification of the signal intensity as well as to improvement of the physical-chemical stability and homogeneity of the biochips. [117,118] The functionalization of the dendrimeric slides with maleimido groups was performed by Maja Köhn The degree of protein immobilization was evaluated by screening the slide for fluorescence.…”