Technology
The simulated zero-reflectance condition at the core of the AIR sensor technology
AIR is a silicon chip based label-free biosensor platform that enables the detection of any probe/target pair of analytes by optically sensing molecular binding at the chip surface. By functionalizing the surface with highly specific probe molecules (ex. antibodies, complementary DNA, etc.) target binding can be detected and quantified directly with high sensitivity. As a label-free technique, the assay procedure requires <30 minute process time. Comparable labeled techniques require 2 or 3 additional chemistry steps after target binding, typically resulting in >2 hour process times and adding substantial reagent cost. The low complexity of our AIR method is also highly amenable to multiplexing, allowing potentially 100's of different probes to be arrayed on the chip surface for simultaneous detection without interference. This detection platform is unique in its combination of rapid testing, multiplex capability, and low cost, and has strong commercialization potential as it addresses many diagnostic needs in both research and patient care.
Figure 1. High Sensitivity detection using a zero reflectance condition on a coated silicon wafer. A) Thin film schematic and modeling variables. B) Simulated reflectance of a silicon dioxide layer on Silicon in an air ambient for 633 nm laser light – the zero condition exists for a 1419 Å thick film at 70.7° incidence. C) biodetection methodology – molecular binding causes a strong reflactance increase
AIR is based on the observation that the performance of a single layer antireflective (AR) coating is very sensitive to the thickness of the film. This sensitivity increases as the reflectance approaches zero. Therefore, by precisely engineering a substrate with an AR coating that approaches zero reflectance, a sensor with the capacity to detect nanoscale thickness changes on its surface is formed. As shown in figures 1A and 1B, a substrate meeting this criterion can be formed with ~1420 Å of silicon dioxide on a silicon wafer, when an s-polarized laser source is incident at 70.7°. This condition relies on precise Å-level control over the oxide layer thickness, which is why the well known Si/SiO2 materials system was chosen for our implementation. Advancements in semiconductor fabrication have made Å-level control and uniformity of this coating relatively easy to achieve with careful process development (as we have demonstrated). Functionality is achieved by immobilizing highly specific probe molecules on the oxide layer, thereby allowing specific target molecules to be detected when they bind to the probes (figure 1C). By measuring the resultant reflectance change, the target concentration in the sample can be quantitatively detected without the need for subsequent labeling chemistries that add time and cost to current assays.
Figure 2. Experimental AIR setup and reference images. A) The low-complexity AIR imaging reader. B) Reference image of a 264 spot photolithographically patterned array. C) Imaging reference chips are fabricated using photolithography and highly accurate etching to form arrays of spots slightly thicker than the remaining film. Images of arrays with spots heights of 700 pm and 10 pm are shown. D) Relative sizes of human serum albumin (HSA), FITC dye, and water, compared to a 10 pm step height. (i.e., smal fractions of a mollecular monolayer can be imaged).
In addition to simplified label-free chemistry, the AIR measurement is also low complexity, as illustrated in figure 2A. This basic laser reflectance measurement is implemented in an imaging format, allowing a large number of isolated array spots to be imaged simultaneously on the biochip, as demonstrated by the image of the photolithographically pattern thickness array in figure 2B. In our biosensing arrays, each of the >250 spots in this image could correspond to a different antibody. Because imaging uses a CCD camera, full analysis of a chip is accomplished in less than a second. Even though the device is deceptively simple, high sensitivity is routinely obtained, as demonstrated in figure 2C where spots with 700 pm and 10 pm thicknesses are clearly observed. This is far smaller than the physical size of proteins and other molecules, as shown schematically in figure 2D.
