Microfluidic chips antibody immobilization

Adhesion-based strategies make use of the protein markers expressed by cells in order to immobilize them to specific points within microfluidic devices. This has been identified as a robust and reliable method for POC applications. Typically an antibody with (specific) affinity for the protein marker of interest (e.g., CD4 in HIV diagnostics or EpCAM in CTC testing) is immobilized on the surface of a microfluidic device (Fig. 1). The capture area can be defined in various shapes, patterns, and locations, e.g., to optimize binding and/or detection. This approach was used by Nagrath et al. [3] to isolate CTCs using a microfluidic chip which consisted of an array of 78,000 microposts which were coated with anti-EpCAM antibodies. The posts increased the inner surface area of the chip and thus enhanced the efficiency of CTC capture. Up to 5.1 ml of whole blood, acquired from 116 patients with... 

The development of a cellular microenvironment in a microfluidic chip starts with device fabrication. The most commonly used material for fabrication is polydimethylsiloxane (PDMS). One of the most important components to develop such platforms is the extracellular matrix (ECM), which is the 3D cellular microenvironment. Different approaches have been followed to pattern the ECM inside the microfluidic device, which will significantly affect the arrangement of cells on the chip. Following this, the target cells are seeded and cultured. Such cell culture techniques would be common for drug screening as well as the fundamental research. But for CTCs detection, usually the antibodies will be immobilized on the chip before introducing the cells. 

An example of the immobilization of antibodies on channel surfaces was presented by Eteshola and Leckband [395]. A microfluidic sensor chip was developed to quantify a model analyte (sheep IgM) with sensitivities down to 17 nM. This was achieved by first immobilizing a layer of bovine serum albumine (BSA) onto the channel wall, followed by specific adsorption of protein A to which the primary antibody for IgM was coupled covalently. This antibody could capture IgM, which was detected with the secondary antibody, labeled with horseradish peroxidase (Scheme 4.91). This enzyme catalyzes the conversion of the fluorogenic substrate 3-(p-hydroxyphenyl)propioni c acid into a fluorophore, which was quantified off-chip with a spectrofluorometer. The measured fluorescence signal was proportional to the analyte concentration in the test sample. 

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