This thesis presents the development of a micro-aspirator chip using vacuum expanded microchannels for mechanical characterization of single cells. Mechanical properties of cells can offer valuable insights into the pathogenic basis of diseases and can serve as a biomarker to identify cells depending on disease state, and thus have the potential for use in human disease diagnostic applications. Micropipette aspiration and atomic force microscopy (AFM) are the most commonly used techniques for measuring mechanical properties of single cells. Though powerful and versatile, both methods have two drawbacks. First, micromanipulation of glass micropipettes and AFM tips require expertise and extensive operator skills. Second, the serial manipulation process severely limits the throughput. Although recently reported microfluidic micropipette device showed the potential of microfluidic chip type micropipette aspiration, difficulty in cell trapping and unnatural cell deformation remain to be solved. In order to address these limitations, a high-throughput micro-aspirator chip, which can deliver, trap, and deform multiple cells simultaneously with single-cell resolution without skill-dependent micromanipulation was developed. The micro-aspirator chip is composed of 20 arrays of cell traps and aspiration channels. The principle of cell trapping is based on differences in flow resistance inside the microfluidic channels. Once the first cell trap is filled with a cell, the next cell coming in passes by the trap and is captured in the next trap. After all traps are filled with cells, negative pressure can then be applied to the integrated aspiration channels using hydrostatic pressure. The aspiration channels are positioned at the center of a trapped cell both in vertical and horizontal directions to obtain a good seal just like a traditional micropipette, a design made possible through a vacuum expanded raised microfluidic channel fabrication technique. Device operation was demonstrated using HeLa cells. The cell trapping efficiency was almost 100 percent. Using this device, Young's modulus of 1.3 ± 0.8 kPa (n = 54) was obtained for HeLa cells. Device to device variation was less than 15.2 percent (n = 3), showing good repeatability of the device. No dependence of the Young's modulus on the cell diameter was found.

No / The aim of the present work is to evaluate the similarity between PDMS membranes and human skin in vitro in permeation study by linear free energy relationship (LFER) analyses. The values of the permeability coefficient log Kp (cm/s) under reliable experimental conditions were collected from the literature for a set of 94 compounds including both neutral and ionic species, which cover a broad range of structural diversity. The values of log Kp (cm/s) have been correlated with Abraham descriptors to yield an equation with R2 = 0.952 and SD = 0.38 log units. The established LFER model for log Kp (cm/s) across PDMS membranes showed no close analogy with that through human skin in vitro. A further critical analysis of the coefficients of the LFER models confirmed that the PDMS permeation system is a very poor model for human skin permeation.

Sepsis, commonly referred to as blood poisoning, is a serious medical condition characterized by a whole-body inflammatory state caused by microbial infection. Rapid treatment is crucial, however, traditional culture-based diagnostics usually takes 2-5 days.  The overall aim of the thesis is to develop microfluidic based sample preparation strategies, capable of isolating bacteria from whole blood for rapid sepsis diagnostics.  Although emerging technologies, such as microfluidics and “lab-on-a-chip” (LOC) devices have the potential to spur the development of protocols and affordable instruments, most often sample preparation is performed manually with procedures that involve handling steps prone to introducing artifacts, require skilled technicians and well-equipped, expensive laboratories.  Here, we propose the development of methods for fast and efficient sample preparation that can isolate bacteria from whole blood by using microfluidic techniques with potential to be incorporated in LOC systems. We have developed two means for high throughput bacteria isolation: size based sorting and selective lysis of blood cells. To process the large blood samples needed in sepsis diagnostics, we introduce novel manufacturing techniques that enable scalable parallelization for increased throughput in miniaturized devices. The novel manufacturing technique uses a flexible transfer carrier sheet, water-dissolvable release material, poly(vinyl alcohol), and a controlled polymerization inhibitor to enable highly complex polydimethylsiloxane (PDMS) structures containing thin membranes and 3D fluidic networks. The size based sorting utilizes inertial microfluidics, a novel particles focusing method that operates at extremely high flow rates. Inertial focusing in flow through a single inlet and two outlet, scalable parallel channel devices, was demonstrated with filtration efficiency of >95% and a flowrate of 3.2 mL/min. Finally, we have developed a novel microfluidic based sample preparation strategy to continuously isolate bacteria from whole blood for downstream analysis. The method takes advantage of the fact that bacteria cells have a rigid cell wall protecting the cell, while blood cells are much more susceptible to chemical lysis. Whole blood is continuously mixed with saponin for primary lysis, followed by osmotic shock in water. We obtained complete lysis of all blood cells, while more than 80% of the bacteria were readily recovered for downstream processing. Altogether, we have provided new bacteria isolation methods, and improved the manufacturing techniques and microfluidic components that, combined offer the potential for affordable and effective sample preparation for subsequent pathogen identification, all in an automated LOC format. / QC 20120611

No description available.

A spin-cast silicone membrane has been successfully bonded between two injection-molded microstructured plastic discs. This sandwich structure creates a useful platform for mass production of microfluidic systems, provided that the bonds are leakproof. The bonds were achieved by a silicon dioxide coating deposited on the plastic discs by evaporation. This investigation is concerned with the process and the result only, no theory is discussed.