(72h) Dielectrophoretic Separation of Nano-Particle Conjugated Bacterial Cells within Micro-Scale Architecture

The detection and counting of bacterial cells can provide a tremendous impetus to rapid diagnostics and clinical applications. The traditional culture, growth and colony counting techniques, although well-established and reliable, are time-consuming (may need a few hours to days) and thus, cannot provide a fast and accurate means of quantitation which may be vital in some emergency situations. In addition, these traditional methods need extra effort and investment for maintenance of sterility for accurate and reproducible results. The utilization of chip-based techniques has obvious advantages in terms of enhanced speed of detection, automation, and point-of-care use. For this purpose a concentration step is normally added prior to the detection process. One of the most widely used concentration techniques in fluidic microchips is dielectrophoresis (DEP). This refers to the interaction of non-uniform electric field with polarization charges induced in any electrically neutral particle which results in differential forces and selective trapping over the electrodes causing the non homogenous electric field. Dielectrophoresis has emerged as a very useful tool for selective trapping and sample pre-concentration. For alternating fields the selectivity towards a certain particle of a specific dielectric constant depends on the operating frequency of such an alternating field. We are developing a novel cell sorting method using nanoparticle conjugates of bacterial cells by actively capturing conjugated cells over a set of interdigitated electrodes and then also measuring the capture efficiency. We start with a mixture of E.Coli (BL21) and Psuedomonas Aerogenosa cells. Goat anti mouse IgG conjugated gold nano-particles (10nm) (Ted Pella Inc., Redding, CA, USA) are used to form conjugates with E.Coli (BL21) cells using monoclonals of Anti E. Coli antibodies. Finally, the gold conjugated cells in the mixture sample are flown through a hybrid micro-fluidic platform with an upper die of PDMS, containing a micro-channel and a lower die of silicon with imprinted interdigitated electrodes. The electrodes are fed with an A/C frequency which is varied temporally until the first capture begins to happen. In order to visualize the capture optically fluorescent dies are used. Figure below shows the microscopic image of a micro-channel with agglomerates of captured E. Coli (BL21) cells dyed with Acridine Orange (AO). We intend to separate the nano-particle conjugated E. Coli (BL21) cells from the mixture using this method. The capture yields will be verified by fluorescence measurements and the selectivity will be evaluated by running a multiplex PCR over the retentate solution.