We are interested in the label-free sensing of large numbers of molecules quantitatively to enable the profile of a complex system. Ideally we would like to do this form whole blood so that we have a real point-of-care device. Current array technologies are based on fluorescence and have reached very high capacities in terms of numbers of assays. However, each of the assays has to be labelled with a fluorescent marker such as a dye or a quantum dot and this provides a significant overhead in the preparation of the array. Further, fluorescence is very hard to quantify confidently and the resulting data are at best semi-quantitative and the return on the preparation investment is poor.
Aims and Objectives
We have developed a nanoparticle based label-free screening technology that produces a quantitative assessment of the concentrations of target molecules in complex fluids such as blood.
The nanoparticle particle biophotonic array can screen molecules with a sensitivity of 1 ng ml-1 and an accuracy of 15% for antibodies and the detection limit scales with mass. We can screen 100 assays in 10 minutes although we have printed and imaged 6000 spot arrays but there is no limit to the array size.
Using the analysis of blood we can profile up to 100 components of the blood proteome targeting a system’s level view of diseases. We are currently working on:
- Allergy and Intolerance to common allergens including food
- Profiling the immune system response to diseases
- Differential diagnosis of Lung Diseases
The technology was developed as part of an EPSRC Basic Technology grant for screening label-free arrays. Seed nanoparticles are printed into the array configuration using and Arrayjet inkjet printer. The seeds are then removed and grown into larger scattering centres. Returning the array to the print allows the array spots to be functionalised with proteins, allergens or antibodies for detecting target molecules in the complex medium.
The array reader is a small form reader that interrogates the arrays in an internal configuration arrangement looking at scattered light normal to the surface. The light is collected in real time on a video camera from which a kinetic analysis may be performed.
The project is now moving from the developmental stage to the applications looking for research targets within the blood proteome. Allergy is becoming increasingly prevalent in western society both as allergy and intolerance. Interestingly allergy is hardly known in Africa. The current tests look for IgE in blood for a specific allergen and if the concentration is larger than 170 ng ml-1 then allergic rhinitis symptoms become apparent. However, in some peanut allergic patients the IgE allergy to peanut makes up approximately one third of the total IgE. In addition, binding to the peanut allergen is dominated, by approximately 2:1, by binding from IgG. The project is looking into the theories of allergy and intolerance with a full epitope screening of blood.
All diseases have an effect on the blood proteome whether as the immune system response to the specific disease or pathogen or the presence of new disease specific proteins or biomarkers in the blood. The approach is to look both the biomarkers but also the systemic response to the disease condition. We are exploring these ideas with differential diagnosis of lung diseases with Pfizer and the immune response post-operatively.
1. Human Serum Albumin Interference on Immuno-kinetic Assay for Antibody Screening in Model Blood Sera. Bernard van Vuuren, Thomas Reed, Rouslan V. Olkhov and Andrew. M. Shaw, Analytical Biochemistry (2010 In Press, doi:10.1016/j.ab.2010.05.015).
2. Localized Surface-Plasmon Resonance Biosensing: a Comparison of Position and Intensity Monitoring. W. Andrew Murray, Rouslan V. Olkhov, Andrew M. Shaw and William L. Barnes (Submitted, J Phys. Chem C, 2010)
3. Plasmon Biophotonic Arrays for Multi-Analyte Biosensing in Complex Media. R. V. Olkhov1, A. Jerdev, A. M. Shaw1 and W. L. Barnes (Book Chapter ?)
4. Quantitative label-free screening for antibodies using scattering biophotonic microarray imaging. Olkhov RV, Shaw AM., Analytical Biochemistry. 2010 Jan 1;396(1):30-5. Epub 2009 Aug 11.