Towards real-time detection of cellular cytokine secretion with photonic ring resonators in cellular bioreactors
Naar real-time detectie van cellulaire cytokine secretie met fotonische ring resonatoren in cellulaire bioreactoren
Ryken, Jef; R0264197
Cytokines are small cell signalling proteins that are important in the hosts immune system, specifically in the responses to infection, inflammation, trauma, etc. One of these cell signalling proteins is TNFa which is a pro-inflammatory cytokine that is secreted by a variety of cells such as macrophages, lymphocytes and neurons. TNFa is involved in apaptotic cell death, fever induction and inflammation and recent studies have shown that dysregulation in TNFa production is engaged in various human diseases such as Alzheimer's disease, cancer and diabetes. Although, cell signalling pathways are complex mechanisms that involve a reciprocity of many different cytokines in various time spans after activation, current cytokine detection techniques are often time consuming, expensive and lack the possibility of multiplex detection. Therefore, there is a need for cytokine detection techniques that allow a fast, easy and sensitive quantification and enable multiplex detection in real-time. A promising candidate is the silicon waveguide based ring resonator sensor which is an optical sensor that enables specific and sensitive protein detection based on refractive index sensing. Furthermore, ring resonators are CMOS compatible and have an unprecedented small size, making them very intresting for LOC applications.In this work, silicon waveguide ring resonators are used for the quantification of the TNFa levels secreted by U937 cells. SPR is used as a benchmark for the ring resonators. Therefore, home made SiO2-covered SPR substrates are characterized and compared with standard home made Au substrates in bulk sensitivity measurements. The functionality of the SiO2-covered SPR substrates for biosensing applications is proven and the substrates are used for TNFa bio-assay characterization. During this characterization, the different parameters such as primary antibody immobilization, non-specific adsorption reduction and signal enhancement factors such as secondary antibodies and beads are investigated. A standard sandwich assay with secondary antibodies resulted in the best LOD and this bio-assay is transferred onto the ring resonator sensors. It is observed that the same affinities are obtained with SPR and the ring resonators. However, the LOD acquired with the ring resonators is higher due to the increased noise in the ring resonator signal. This is also shown in the cell secretion quantification experiments where the ring resonators are able to detect a TNFa concentration of 140 ng/mL while ELISA and SPR even allow detection of concentrations down to10 ng/mL. The first steps towards real-time TNFa secretion detection are investigated, however, further work is advised in the optimization of the real-time detection process.To conclude, we investigated the utility of the ring resonator sensors for quantification of the TNFa level secreted by U937 cells. To do so, SiO2-covered SPR substrates are fabricated to be used as a benchmark and assay optimization. Moreover, this was the first report to describe the succesful use of such substrates for biosensing applications. We thus will show the potential of the ring resonators as real-time cell secretion detectors for LOC applications.