Projects (2)

A microfluidic Biochip platform for dynamic real-time detection and quantification of proteins

Protein analytics based on immunoassays such as ELISA (enzyme linked immunosorbent assay) play an important role in today’s clinical diagnostics. However, the immanent restric-tion to mono parametric analytics resulting in large sample consumption, long incubation time and tedious handling impede applications in point of care units in close vicinity of the patient.

To overcome these limitations and their considerably high costs a novel microfluidic biochip platform is proposed providing the following: Dynamic real-time detection and quantification of a broad range of clinically relevant proteins in a multi parametric fashion.

Figure 4: competitive immunoassay in hydrogel, labeled antigen (yellow) is replaced by the same unlabeled species (grey)

The system is based on a competitive immunoassay (Figure 4). The capture antibodies are immobilized in hydrogel within the microfluidic serpentine channel structure (Figure 5). This surface attached polymer network provides an increased probe density over conventional immobilization strategies. The new inline principle permits multi parametric analysis of differ-ent proteins in one single experiment using one single sample volume. Real-time de-tection of analyte molecules resulting in a decrease of bound competitive labeled antigen is achieved by using total internal fluorescence spectroscopy. The dedicated fluidic dynamic design overcomes mass transport limitations due to diffusion with respect to protein kinetics. Several flow velocities and flow profiles are investigated yielding higher signal intensities and significantly reduced assay time. This new concept was demonstrated for proteins covering the relevant mass spectrum.

Figure 5: microfluidic protein biochip

Development of technical platforms for microarray applications

For a versatile microarray production and read-out technology, we develop a system using fluorescence excitation by an evanescent field.

Monochromatic light is coupled into a glass or plastic substrate, on which the microarray is printed. The substrate acts as a waveguide with an evanescent wave travelling along the surface.

Figure 6: Labeled analyte molecules bound to the surface are excited by the evanescent field.

Labeled analyte molecules bound to the surface are excited by the evanescent field. (Figure 6).

Chips can be placed in a reader system consisting of a laser, a flow cell and a CCD camera (Figure 7).

Figure 7: Monochromatic light (4) is coupled into the plastic microarray substrate (1), such that total internal reflection occurs. An evanescent field travels along the surface allowing fluorescence excitation. The substrate serves as a waveguide, which allows detection of the interaction between the immobilized probes (2) and the labelled DNA in the flow cell (3). A CCD-camera (7) with its filter (8) detects the fluorescence intensities. A cooling system (5) with a peltier element behind (6) regulates the temperature of the hybridization solution.

Using our 3D immobilization technology with an increased probe density in combination with the TIRF-based read-out gives us the possibility to study hybridization kinetics, to detect gene mutations (e.g. SNP, Single Nucleotide Polymorphism) or to perform on-chip PCR and on-chip NASBA (Nucleic Acid Sequence Based Amplification) as well as conventional DNA- and protein microarrays.