Development of an Item Unique Identification strategy for the legacy components of the US Marine Corps M1A1 Abrams Tank

MBA Professional ReportThe Office of the Secretary of Defense (OSD) has mandated that every component that meets certain cost and management criteria in the Department of Defense's (DoD) tangible inventory must have a valid Item Unique Identification (IUID) mark by December 2010. The IUID program is expected to increase force readiness, enhance the lifecycle management of assets, and provide more accurate asset valuation to achieve unqualified audit opinions on DoD financial statements. One of the challenges of the program is to achieve IUID-marking saturation throughout in-use, or legacy, inventories with minimal interruption of operational readiness. The purpose of this project is to propose an effective implementation strategy for the legacy items that meet the DoD's requirements for IUID marking in the US Marine Corps M1A1 Abrams tank community. This plan could potentially serve as a model for other communities facing the challenge of IUID implementation. This project examines different implementation alternatives and identifies and develops the determined best course of action. In addition, an estimation of the costs to implement the chosen plan will be provided for comparison and decision-making purposes.http://archive.org/details/developmentofnit1094510296Outstanding ThesisApproved for public release; distribution is unlimited

Nucleofection, an efficient nonviral method to transfer genes into human hematopoietic stem and progenitor cells.

Contains fulltext : 49545.pdf (publisher's version ) (Open Access)The targeted manipulation of the genetic program of single cells as well as of complete organisms has strongly enhanced our understanding of cellular and developmental processes and should also help to increase our knowledge of primary human stem cells, e.g., hematopoietic stem cells (HSCs), within the next few years. An essential requirement for such genetic approaches is the existence of a reliable and efficient method to introduce genetic elements into living cells. Retro- and lentiviral techniques are efficient in transducing primary human HSCs, but remain labor and time consuming and require special safety conditions, which do not exist in many laboratories. In our study, we have optimized the nucleofection technology, a modified electroporation strategy, to introduce plasmid DNA into freshly isolated human HSC-enriched CD34(+) cells. Using enhanced green fluorescent protein (eGFP)-encoding plasmids, we obtained transfection efficiencies of approximately 80% and a mean survival rate of 50%. Performing functional assays using GFU-GEMM and long-term culture initiating cells (LTC-IC), we demonstrate that apart from a reduction in the survival rate the nucleofection method itself does not recognizably change the short- or long-term cell fate of primitive hematopoietic cells. Therefore, we conclude, the nucleofection method is a reliable and efficient method to manipulate primitive hematopoietic cells genetically

Primitive human hematopoietic cells give rise to differentially specified daughter cells upon their initial cell division.

Contains fulltext : 50393.pdf (publisher's version ) (Closed access)It is often predicted that stem cells divide asymmetrically, creating a daughter cell that maintains the stem-cell capacity, and 1 daughter cell committed to differentiation. While asymmetric stem-cell divisions have been proven to occur in model organisms (eg, in Drosophila), it remains illusive whether primitive hematopoietic cells in mammals actually can divide asymmetrically. In our experiments we have challenged this question and analyzed the developmental capacity of separated offspring of primitive human hematopoietic cells at a single-cell level. We show for the first time that the vast majority of the most primitive, in vitro-detectable human hematopoietic cells give rise to daughter cells adopting different cell fates; 1 inheriting the developmental capacity of the mother cell, and 1 becoming more specified. In contrast, approximately half of the committed progenitor cells studied gave rise to daughter cells, both of which adopted the cell fate of their mother. Although our data are compatible with the model of asymmetric cell division, other mechanisms of cell fate specification are discussed. In addition, we describe a novel human hematopoietic progenitor cell that has the capacity to form natural killer (NK) cells as well as macrophages, but not cells of other myeloid lineages

High-resolution video monitoring of hematopoietic stem cells cultured in single-cell arrays identifies new features of self-renewal

To search for new indicators of self-renewing hematopoietic stem cells (HSCs), highly purified populations were isolated from adult mouse marrow, micromanipulated into a specially designed microscopic array, and cultured for 4 days in 300 ng/ml Steel factor, 20 ng/ml IL-11, and 1 ng/ml flt3-ligand. During this period, each cell and its progeny were imaged at 3-min intervals by using digital time-lapse photography. Individual clones were then harvested and assayed for HSCs in mice by using a 4-month multilineage repopulation endpoint (>1% contribution to lymphoid and myeloid lineages). In a first experiment, 6 of 14 initial cells (43%) and 17 of 61 clones (28%) had HSC activity, demonstrating that HSC self-renewal divisions had occurred in vitro. Characteristics associated with HSC activity included longer cell-cycle times and the absence of uropodia on a majority of cells within the clone during the final 12 h of culture. Combining these criteria maximized the distinction of clones with HSC activity from those without and identified a subset of 27 of the 61 clones. These 27 clones included all 17 clones that had HSC activity; a detection efficiency of 63% (2.26 times more frequently than in the original group). The utility of these characteristics for discriminating HSC-containing clones was confirmed in two independent experiments where all HSC-containing clones were identified at a similar 2- to 3-fold-greater efficiency. These studies illustrate the potential of this monitoring system to detect new features of proliferating HSCs that are predictive of self-renewal divisions