Handheld imaging photonic crystal biosensor for multiplexed, label-free protein detection.

We present a handheld biosensor system for the label-free and specific multiplexed detection of several biomarkers employing a spectrometer-free imaging measurement system. A photonic crystal surface functionalized with multiple specific ligands forms the optical transducer. The photonic crystal slab is fabricated on a glass substrate by replicating a periodic grating master stamp with a period of 370 nm into a photoresist via nanoimprint lithography and deposition of a 70-nm titanium dioxide layer. Capture molecules are coupled covalently and drop-wise to the photonic crystal surface. With a simple camera and imaging optics the surface-normal transmission is detected. In the transmission spectrum guided-mode resonances are observed that shift due to protein binding. This shift is observed as an intensity change in the green color channel of the camera. Non-functionalized image sections are used for continuous elimination of background drift. In a first experiment we demonstrate the specific and time-resolved detection of 90.0 nm CD40 ligand antibody, 90.0 nM EGF antibody, and 500 nM streptavidin in parallel on one sensor chip. In a second experiment, aptamers with two different spacer lengths are used as receptor. The binding kinetics with association and dissociation of 250 nM thrombin and regeneration of the sensor surface with acidic tris-HCl-buffer (pH 5.0) is presented for two measurement cycles.

Quantitative measurement of human anti-HCV Core immunoglobulins on an electrical biochip platform.

The importance of early diagnosis devices increased continuously in the last two decades and plays an important role in medical care. Early stage diagnosis of e.g. ovarian cancer, HCV-infection or HIV-infection increased the survival rate of patients significantly. In parallel there is a trend leaving centralized diagnostic laboratories in order to get closer to the patient to perform analysis of even complex parameters in the field. This often saves time, increases the prognosis of the patient significantly and is cheaper in many cases. In this study we employ a rapid and cost-effective detection system based on electrical biochip technology for decentralized detection of anti-HCV Core immunoglobulins (HCV antibodies). In this system the qualitative and quantitative detection of virus-specific antibodies is done by an ELISA directly on a gold electrode array utilizing HCV Core as capture antigen. The biochip allows antibody detection within 20 min. Signal amplification was done by enzyme labelling and by "Single Electrode Redox Cycling". This method enhances current signals up to 40-fold in comparison to simple oxidation. The sensitivity of this approach is therefore comparable to a standard microtiter plate based ELISA with a 9-fold saving of assay time. This biochip system allows serum or whole blood analysis with no signal loss or increasing background caused by the red blood cells. Fields of application can be hospital emergency units where only single detections have to be conducted in a quick manner or by the general practitioner.

Electrical microarrays for highly sensitive detection of multiplex PCR products from biological agents.

For the sensitive detection of amplicons derived from diagnostic PCR, a novel electrical low-density microarray is applied and compared to state-of-the-art quantitative real-time PCR. The principle of the electrochemical method and the effective use for analysis are described. Interdigitated array gold electrodes (IDA-E) embedded into a silicon chip are the core technology of the fully automated compact biosensor system, basing on enzyme coupled electrochemical detection. The biointerface is built up with thiol-modified capture oligonucleotides on gold and mediates the specific recognition of hybridised target DNA amplified with uniplex or multiplex PCR. In here we show the potential of the designed electrical microarray to function as an advanced screening method for the parallel detection of a panel of the four pathogens Bacillus anthracis, Yersinia pestis, Francisella tularensis and ortho pox viruses (genus), which are among the most relevant biowarfare agents. PCR products, generated from 10 to 50 gene equivalents, have been detected reproducibly. The experiments with varying pathogen amounts showed the good reliability and the high sensitivity of the method, equivalent to optical real-time PCR detection systems. Without PCR the total assay time amounts to 27 min. The advantage of the combination of multiplex-PCR with electrical microarray detection avoiding intensive PCR probe labelling strategies is illustrated.