Ultra-sensitive detection of biomarkers and applications in pharmaceutical discovery and development. Example application to rat cardiac troponin I assay via nanoparticle probes

e14594 Although specific to heart, rat cardiac troponin I (cTnI) is an example of an important biomarker for assessing drug-induced cardiotoxicity in animal models used in various phases of drug discovery and development. Current commercially available assays can only detect 10 ∼ 100 pg/mL in serum at the lowest limits. To improve the sensitivity of rat cTnI assay, we have developed a generically applicable, microarray based nano-probe test. Our rat cTnI assay algorithm uses a multi-step robotic process, which relies on non- isotropically oriented antibodies on functionalized glass as multiplexed microarrays to capture cTnI from serum. Functionalized, 130 angstrom diameter gold nano-probes (measured by static light scattering, 5 nm S.D.) also bind to the troponin through a molecular-scale complex containing antibodies. The troponin-bound molecular complex is then quantified through silver enhancement of the functionalized gold. Assays in this format can be rapidly configured and implemented for a wide array of potential biomarkers. For cTnI we have demonstrated a robust and ultra-sensitive assay with an LOD of less than 500 femtograms of rat cTnI per mL serum, and an overall CV of less than 20%. The assay also shows very low background, a broad dynamic range and over 3 logs of linear dose response. As an example of the potential of high sensitivity, the nanoparticle-based rat cTnI assay could significantly increase the effectiveness of measuring drug-induced heart damage at very low drug dosages and early times. Such sensitive and early measurements can improve examination of the safety of drug candidates while correspondingly reducing drug development time and cost. [Table: see text]