Biotechnology for immune cell detection and evaluation - Devices specifically designed to capture, detect or to evaluate alterations in leukocyte structure and function.

摘要

The advent of Nano-biotechnology has opened a whole new area of biological study that has allowed us to pattern, manipulate, separate, evaluate and detect biological structures and functions that were previously not possible. This dissertation will describe different types of micro-fabricated devices that were designed to quantify and/or evaluate the functional activities of cells of the immune system. The micro-fabricated devices were developed in collaboration with our colleagues at Princeton University as part of our Cellular Micro-dynamics Program of the Nanobiotechnology Center (NBTC) at Cornell University.;A micro-fluidic "bump array" was designed to either separate cells or biological components based on changes in "hydrodynamic" cell size or particulate size; thus enabling the device to separate white blood cells (leukocytes) and platelets. The distribution histograms generated with the micro-device were comparable to that of forward light scatter analysis obtained by conventional flow cytometry. Advantages of the chip separator include significantly lower blood volume requirements and reduced costs per assay performed. Using a fabricated in vitro capillary device, it was successfully demonstrated that leukocyte subsets show differential ability to roll, adhere, chemotax and diapedesis under flow conditions in a micro-fabricated device. Neutrophils from blood delivered into the device can be assessed across the above mentioned functions without further modifications typical of current in vitro methods. One exception is that lymphocytes require additional adhesive or surface substrate adhesion molecules in order to assess these same normal biological functions in the micro-fluidic device. A fabricated T-cell biosensor was able to capture, detect and quantify the number of CD4 + lymphocytes from a few micro-liters of blood. The design of this type of device provides support that microdevices can be used to develop new diagnostic platforms that can evaluate alterations in immunological cell structure and functions using single sample and lower volumes of blood. These devices were used in several experiments exploring the role of stress in altering an organism's susceptibility to environmental stressors.;It was demonstrated that stress alters blood and bone marrow (BM) leukocyte populations following Acute Cold Restraint Stress (ACRS). It was shown that mice undergo lymphopenia shortly after ACRS while simultaneously undergoing neutrophilia and that these alterations are differentially mediated through the beta-Adrenergic Receptor (beta-AR) pathway. This increase in blood neutrophils following ACRS is due to the activation/mobilization of BM neutrophils, which was mediated by the sympathetic nervous system through a beta1-AR mechanism. However, alterations in CD3+ T-Cell trafficking were mediated through beta2-AR, while CD19+ B-cell trafficking was modulated through a combination of beta1-AR and beta2-AR. It was postulated that early activation of neutrophils associated with stress may lead to BM neutrophil exhaustion, which could lead to decreased host resistance to Listeria monocytogenes (LM) following stress.;The merging of Micro-/Nano-technology with biology will allowed current and future researchers to manipulate, pattern and view biological structures and functions in a way not previously possible. These types of devices, which are currently being developed, will be the research tools, diagnostic assays, detection sensors and medical delivery systems of tomorrow. The limitation for Nanobiotechnological applications in science only relies on ones imagination.