Diffractive Fluorescence Multiplex Assay

A 14-member team from the University of Southampton and from the Rutherford Appleton Laboratory in Didcot, both in the UK, has combined microscopic bar codes and fluorescent biomolecules in a rapid, noninvasive assay with high encoding capacity. The technique could be useful for immunodiagnostics and single nucleotide polymorphism genotyping.

The serrated-edge micro bar-code particle diffracts the laser beam, and a CCD camera captures and analyses the diffraction pattern and reads the code. Courtesy of Joseph K. She, University of Southampton.

The researchers spin-coated Chestech Ltd.’s photoactive epoxy SU-8 onto silicon wafers and marked bar-coded diffraction gratings on them photolithographically. Superimposing gratings with different spacings in two dimensions enabled a 100-μm-long particle to carry up to 10¹⁸ unique codes. Ring-opening of residual surface epoxide groups of SU-8 allowed fluorescent probe molecules to bond to the diffractive microparticles. A custom-built optical system carried out simultaneous diffraction and fluorescence measurements. A HeNe laser at 633 nm and a diode-pumped solid-state laser at 532 nm excited fluorescence in Cy5 and Cy3 markers, and CCD cameras captured the images. Both wavelengths interacted with the micro bar codes to form transmission diffraction patterns, recorded by another CCD camera and analysed by dedicated computer software, taking less than 1 ms to read the code on a particle.

(Analytical Chemistry, 15 March 2008, pp. 1902-1909)