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THE USE AND DEVELOPMENT OF LASER MICRODISSECTION TO SEPARATE SPERMATOZOA FROM EPITHELIAL CELLS FOR STR ANALYSIS
Title
THE USE AND DEVELOPMENT OF LASER MICRODISSECTION TO SEPARATE SPERMATOZOA FROM EPITHELIAL CELLS FOR STR ANALYSIS.
Creator
Sanders, Christine, Ballantyne, Jack, University of Central Florida
Abstract / Description
Short Tandem Repeat (STR) analysis has become a valuable tool in identifying the source of biological stains, particularly from the investigation of sexual assault crimes. Difficulties in analysis arise primarily in the interpretation of mixed genotypes when cell separation of the sexual assailant's sperm from the victim's cells is incomplete. The forensic community continues to seek improvements in cell separation methods from mixtures for DNA typing. This report describes the use of laser...
Show moreShort Tandem Repeat (STR) analysis has become a valuable tool in identifying the source of biological stains, particularly from the investigation of sexual assault crimes. Difficulties in analysis arise primarily in the interpretation of mixed genotypes when cell separation of the sexual assailant's sperm from the victim's cells is incomplete. The forensic community continues to seek improvements in cell separation methods from mixtures for DNA typing. This report describes the use of laser microdissection (LMD) for the separation of pure populations of spermatozoa from two-donor cell mixtures. In this study, cell separation was demonstrated by microscopic identification of histologically stained spermatozoa and female buccal cell mixtures, and STR analysis of DNA obtained from the separated sperm cells. Clear profiles of the male donor were obtained with the absence of any additional alleles from the female donor. Five histological stains were evaluated for use with LMD and DNA analysis: hematoxylin/eosin, nuclear fast red/picroindigocarmine, methyl green, Wright's stain, and acridine orange. Hematoxylin/eosin out-performed all other stains however nuclear fast red/picroindigocarmine could be used satisfactorily with STR analysis. In addition, three DNA isolation methods were evaluated for LMD collected cells: QIAamp (Qiagen), microLYSIS (Microzone Ltd.) and Lyse-N-Go (Pierce Chemical Co.). MicroLYSIS performed poorly, yielding low levels of PCR product. Lyse-N-Go extraction was effective for the recovery of DNA from LMD collected sperm cells while QIAamp isolation performed best for the recovery of DNA from LMD collected epithelial cells. LMD is shown to be an effective, low-manipulation separation method that enables the recovery of sperm while excluding epithelial cell DNA.
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Date Issued
2005
Identifier
CFE0000876, ucf:46652
Format
Document (PDF)
PURL
http://purl.flvc.org/ucf/fd/CFE0000876
Design, Development, and Testing of a Miniature Fixture for Uniaxial Compression of Ceramics Coupled with In-Situ Raman Spectrometer
Title
Design, Development, and Testing of a Miniature Fixture for Uniaxial Compression of Ceramics Coupled with In-Situ Raman Spectrometer.
Creator
Jordan, Ryan, Orlovskaya, Nina, Kwok, Kawai, Ghosh, Ranajay, University of Central Florida
Abstract / Description
This thesis is about the design, development and integration of an in-situ compression stage which interfaces through the Leica optical microscope coupled with a Renishaw InVia micro-Raman spectrometer. This combined compression stage and Raman system will enable structural characterization of ceramics and ceramic composites. The in-situ compression stage incorporates a 440C stainless steel structural components, 6061 aluminum frame, a NEMA 23 stepper motor. Two load screws that allow to...
Show moreThis thesis is about the design, development and integration of an in-situ compression stage which interfaces through the Leica optical microscope coupled with a Renishaw InVia micro-Raman spectrometer. This combined compression stage and Raman system will enable structural characterization of ceramics and ceramic composites. The in-situ compression stage incorporates a 440C stainless steel structural components, 6061 aluminum frame, a NEMA 23 stepper motor. Two load screws that allow to apply compressive loads up to 14,137 N, with negligible off axis loading, achieving target stresses of 500 MPa for samples of up to 6.00 mm in diameter. The system will be used in the future to study the structural changes in ceramics and ceramic composites, as well as to study thermal residual stress redistribution under applied compressive loads. A broad variety of Raman active ceramics, including the traditional structural ceramics 3mol%Y2O3-ZrO2, B4C, SiC, Si3N4, as well as exotic materials such as LaCoO3 and other perovskites will be studied using this system. Calibration of the systems load cell was performed in the configured state using MTS universal testing machines. To ensure residual stresses from mounting the load cell did not invalidate the original calibration, the in-situ compression stage was tested once attached to the Renishaw Raman spectrometer using LaCoO3 ceramic samples. The Raman shift of certain peaks in LaCoO3 was detected indicative of the effect of the applied compressive stress on the ceramics understudy.
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Date Issued
2019
Identifier
CFE0007824, ucf:52809
Format
Document (PDF)
PURL
http://purl.flvc.org/ucf/fd/CFE0007824