Simultaneous detection of C-reactive protein and other cardiac markers in human plasma using micromosaic immunoassays and self-regulating microfluidic networks

Abstract

We show a proof-of-concept in which we combine our previously published concepts of micromosaic immunoassays (μMIAs) with self-regulating microfluidic networks (μFNs) to detect C-reactive protein (CRP) and other cardiac markers such as myoglobin (Mb) and cardiac Troponin I (cTnI). The μFNs are microfabricated in Si, have a well-defined surface chemistry, and are affixed to a bibulous material so as to self-regulate the displacement of an aliquot of liquid through the μFNs using capillary forces. An open section of the channels of the μFNs is covered with a hydrophobic poly(dimethylsiloxane) (PDMS) slab that acts as the substrate for a solid-phase immunoassay. Here, individual assays are conducted using independent channels. These assays are "sequential": series of samples, reagents, and buffers are displaced one after the other over the PDMS surface, and, as these assays are conducted under "microfluidic" conditions, they are fast to perform, very economical in their use of reagents, extremely integrated, and yield high-quality signals. The combinatorial character of μMIAs is exploited to optimize the assay parameters for detecting CRP. In particular, we found it optimal to deposit the capture antibody for CRP on PDMS at a concentration between 20 and 500μgml-1 in PBS in 1min and to detect captured CRP in 2min using a detection antibody having a concentration in PBS of 120μgml-1. With this method, CRP is quantitatively detected within 10min in one microliter of human plasma down to concentrations of 30ngml-1, which suggests the possibility to detect CRP at clinically relevant concentrations for the management of coronary heart disease (CHD) and systemic inflammation. © 2003 Elsevier B.V. All rights reserved.