Modified surfaces for the detection of biomolecules at the single molecule level

A surface composition comprising:

• a substrate,
• a functionalized layer disposed on a surface thereof, and
• an absorption layer disposed on the functionalized layer,
• where the functionalized layer is adapted to bond or absorb the absorption layer,
• where the absorption layer include sites capable of absorbing or binding a first molecule, molecular complex, or molecular assembly or a plurality of first molecules, complexes, assemblies or mixtures or combinations thereof,
• where each first molecule, complex and assembly includes a first fluorescent dye or fluorophore, and
• where the composition increases fluorescent dye or fluorophore lifetime relative to a polyelectrolyte surface.

A method for preparing surface compositions comprising the steps of:

• cleaning a substrate to reduce or eliminate fluorophores from surfaces of the substrate or from the substrate itself, and
• contacting the substrate with an absorbent under conditions to form an absorption layer on a surface of the substrate,
• where the absorption layer include sites capable of absorbing or binding a first species selected from the group consisting of a molecule, molecular complex, or molecular assembly or a plurality of first species selected from the group consisting of molecules, complexes, assemblies or mixtures or combinations thereof,
• where each first species includes a first fluorescent dye or fluorophore, and
• where the substrate increases fluorescent dye or fluorophore lifetime relative to a polyelectrolyte surface.

A method for preparing surface compositions comprising the steps of:

• 2 cleaning a substrate to reduce or eliminate fluorophores from surfaces of the substrate or from the substrate itself,
• placing the cleaned substrate on a support rack in a bottle including a cap having an aperture and a bottom having a plurality of aperture,
• placing a tube within the bottle containing a functionalizing agent,
• capping the bottle,
• inserting a pipette through the cap aperture into the tube above or below a surface of the functionalizing agent,
• connecting the pipette to a source of an inert gas,
• supplying a flow of the inert gas to the bottle via the pipette at a rate sufficient to evaporate or entrain the functionalizing agent in the inert gas flow and sufficient for a pressure in the bottle to be maintained at a desired pressure below the rupture pressure of the bottle, and
• continuing the flow for a time sufficient to achieve a desired degree of substrate functionalization to form a substrate having a functionalized layer formed one or all surfaces of the substrate,
• where the functionalized layer is adapted to bond, absorb or support the absorption layer, or to absorb or bond a first species selected from the group consisting of a molecule, molecular complex, or molecular assembly or a plurality of first species selected from the group consisting of molecules, complexes, assemblies or mixtures or combinations thereof,
• where each first species includes a first fluorescent dye or fluorophore, and
• where the composition increases fluorescent dye or fluorophore lifetime relative to a polyelectrolyte surface.

A method for immobilizing molecules, molecular complexes or molecular assemblies comprising the steps of:

• providing a substrate comprising an absorption layer disposed on a surface thereof, an absorption layer disposed on a functionalized layer disposed on a surface thereof, or a functionalized layer disposed on a surface thereof,
• contacting the surface with a solution comprising a plurality of species selected from the group consisting of molecules, molecular complexes, molecular assemblies or mixtures or combinations thereof, where some or all of the species include a first detectable label, where the contacting is under conditions sufficient to immobilize the species on the functionalized layer or the absorption layer so that a majority of the immobilized species are separately or individually detectable,
• placing the resulting composition in a detection system, and
• detecting the labels.