In this study, we propose a fast, simple method to biofunctionalise microfluidic systems for cellomic investigations based on micro-fluidic protocols. Many available processes either require expensive and time-consuming protocols or are incompatible with the fabrication of microfluidic systems. Our method differs from the existing since it is applicable to an assembled system, uses few microlitres of reagents and it is based on the use of microbeads. The microbeads have specific surface moieties to link the biomolecules and couple cell receptors. Furthermore, the microbeads serve as arm spacer and offer the benefit of the multi-valent interaction. Microfluidics was adapted together with topology and biochemistry surface modifications to offer the microenvironment for cellomic studies. Based on this principle, we exploit the streptavidin-biotin interaction to couple antibodies to the biofunctionalised microfluidic environment within 5 h using 200 μL of reagents and biomolecules. We selected the antibodies able to form complexes with the MHC class I (MHC-I) molecules present on the cell membrane and involved in the immune surveillance. To test the microfluidic system, tumour cell lines (RMA) were rolled across the coupled antibodies to recognise and strip MHC-I molecules. As result, we show that cell rolling performed inside a microfluidic chamber functionalised with beads and the opportune antibody facilitate the removal of MHC class I molecules. We showed that the level of median fluorescent intensity of the MHC-I molecules is 300 for cells treated in a not biofunctionalised surface. It decreased to 275 for cells treated in a flat biofunctionalised surface and to 250 for cells treated on a surface where biofunctionalised microbeads were immobilised. The cells with reduced expression of MHC-I molecules showed, after cytotoxicity tests, susceptibility 3.5 times higher than normal cells. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
KAUST Repository Item: Exported on 2020-10-01
Acknowledgements: This work was partially supported by the project PON "Nuove strategie nanotecnologiche per la messa a punto di farmaci e presidi diagnostici diretti verso cellule cancerose circolanti" (code: PON01_02782) and the project FIRB "Rete Nazionale di Ricerca sulle Nanoscienze ItalNanoNet" (cod. RBPR05JH2P_010, CUP B41J09000110005) granted to the nanotechnology laboratory of the Department of Experimental Medicine of the University of Magna Graecia of Catanzaro. Ennio Carbone is Wenner Gren Foundation senior research fellow and was supported by grant from Italian Association for Cancer Research (AIRC IG 10189). Rossana Tallerico is awarded by triennial fellowships "Luciana Selce" FIRC. The authors have declared no conflict of interest.
- Clinical Biochemistry