Self-referencing biodetection method and patterned bioassays

1. A method of making a self-referencing colorimetric resonant optical biosensor comprising:

(a) immobilizing one or more specific binding substances in an immobilization volume to one or more assay regions of a self-referencing colorimetric resonant optical biosensor, wherein the biosensor comprises a liquid holding vessel comprising a colorimetric resonant optical biosensor as a surface, and a fluid-impermeable divider on the colorimetric resonant optical biosensor surface, wherein the fluid impermeable divider separates the liquid holding vessel into two or more assay regions, wherein the fluid-impermeable divider segregates an immobilization volume within each assay region, and wherein the fluid-impermeable divider allows a reaction volume to encompass all assay regions in the liquid-holding vessel; and

(b) preserving one or more assay regions as one or more reference regions, wherein the one or more reference regions do not contain immobilized specific binding substances.

2. The method of claim 1 , wherein when the biosensor is illuminated a resonant grating effect is produced on the reflected radiation spectrum and wherein the depth and period of a grating of the biosensor are less than a wavelength of the resonant grating effect.

3. The method of claim 1 , wherein a narrow band of optical wavelengths is reflected from the biosensor when the biosensor is illuminated with a broad band of optical wavelengths.

4. The method of claim 1 , wherein the liquid-holding vessel is selected from the group consisting of a microtiter plate well, a test tube, a Petri dish and a microfluidic channel.

5. The method of claim 1 , wherein the biosensor comprises two or more liquid-holding vessels.