Offprint in back pocket / 248 leaves : / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (M.D. 1973) from the Dept. of Medicine, University of Adelaide

Thesis (M.S.)--University of Wisconsin--Madison, 1984. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 78-87).

Thesis (Ph. D.)--University of Wisconsin--Madison, 1984. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Platelets play an integral part in the blood clotting process. The normal platelet count ranges from 150 to 400 x 103/μl. To date, twenty-four platelet-specific antigens have been defined. Human Platelet Antigens (HPA) 1-5 and –15 belong to diallelic systems and antibodies formed in response to immunization against these antigens, following transfusion or transplacental haemorrhage, have been responsible for life-threatening conditions in which the platelet counts of patients are markedly decreased. This is due to the destruction of platelets by platelet antigen - specific antibodies in Neonatal Alloimmune Thrombocytopenia and Post-Transfusion Purpura and in some instances, Platelet Refractoriness. In these clinical situations, transfusions of platelets from random donors do not result in post-transfusion platelet increments. Ideally, a database of blood donors who have been HPA typed should be established. This database can then be used to search for HPA – compatible donors and matched platelet donations can be made available to these patients. The aims of this study were primarily to determine the HPA frequencies in the South African blood donor population and establish a register of HPA-typed donors. Secondly, the established frequencies would be compared to those of other world populations. Anticoagulated blood samples from one hundred and fifty donors from each of the four main South African population groups were obtained for typing of HPA 1-5 and –15 by the PCR-SSP method in order to determine their gene and genotype frequencies and establish a platelet donor database. Two methods of analyses were used to determine statistical similarities and differences between the South African population and a number of world populations (X2) and the degree of genetic distance between them (FST). Phylogenetic trees and Principal Components plots were constructed to illustrate relationships between populations. The HPA gene and genotype frequencies of the South African blood donor population have been determined. An accurate, rapid method of HPA typing has been validated and effective HPA matched platelet transfusions are now possible for the first time in South Africa. The gene frequencies of the White population have been found to be similar to those from European populations and the Bantu-speaking population frequencies compared favourably with those of other African populations. The population group of mixed ethnicity (Coloured) was shown to have unique frequencies and were not similar to any of those populations chosen for comparison except with the other populations in South Africa. / Dr. R. L. Crookes Prof. T.L. Coetzer Prof. M. Dutton

The purpose of this project is to investigate the surface properties of platelet discocytes, echinocytes and spherocytes. Normal "non-sticky" discoid shaped platelets (discocytes) can be transformed by ADP into irregularly shaped echinocytes which are "sticky" and aggregate easily in media containing Ca⁺⁺ and fibrinogen. A model is examined here in which an echinocyte attains its "sticky" properties by evagination of a surface-connected canalicular system. Platelets also evaginate this canalicular system upon hypotonic shock, in which case the platelets swell up to form spherocytes. By comparing the properties of the different geometric forms of platelets insight into the nature of "stickiness" was sought. The surface areas of the discocyte and spherocyte measured microscopically were found to be 16.4 and 36.7x10⁻⁸ cm² respectively while that of the echinocyte was estimated to be 23.7x10⁻⁸ cm² using surface chemical analysis. Electron microscopic examination showed that the canalicular system may not be totally evaginated in the echinocyte. Although it was found that the spherocyte could still be agglutinated passively by ristocetin it had completely lost its ability to aggregate. Microelectrophoretic studies revealed 8 and 6 fold increases in the density of Ca⁺⁺ and Mg⁺⁺ binding sites respectively on the echinocyte surface relative to the discocyte. The spherocyte on the other hand had lost most of its Ca binding sites. Electrokinetic analysis of live, fixed and neuraminidase or alkaline phosphatase treated platelets showed major differences in charge as well as amino, sialic acid and phosphate group densities among the discocyte, echinocyte and spherocyte. The evaginated canalicular membrane surfaces of the latter two were also different. SDS-PAGE of platelets radiolabelled via lactoperoxidase iodination, periodate-borohydride tritiation or neuraminidase/galactose oxidase-borohydride tritiation failed to show any difference in the gel patterns between the three platelet forms. No new glycoprotein species appeared during the transformations. The presence of fibrinogen interferes in a concentration related manner with lactoperoxidase iodination of GP-III on the echinocyte surface. An overall picture is presented here showing differences between the surface properties of platelet discocytes, echinocytes and spherocytes. The accumulated evidence suggests that changes in the whole platelet surface occur during activation and the model of a cloistered "sticky" membrane may be an oversimplification. / Medicine, Faculty of / Pathology and Laboratory Medicine, Department of / Graduate

No description available.

published_or_final_version / Anaesthesiology / Master / Master of Research in Medicine

No description available.

No description available.

Previously it has been shown that sequences containing both Trp²⁵ and Leu³³ are the most effective at inducing Th cell proliferation in HLA-DRB3*0101 positive women, alloimmunised with anti-HPA-1a.  The Leu³³/Pro³³ polymorphism is embedded in the N-terminal plexin/semaphorin/integrin (PSI) domain of GPIIIa.  In the present study, amino acids 1-62 of the GPIIIa (Leu³³ or Pro³³) PSI domain were cloned into the vector pGEX-6p-1.  The recombinant proteins were expressed and tested by ELISA, Luminex and Absorption Assays.  The presence of the HPA-1a/-1b epitope was confirmed by the ability of PSI-Leu³³/-Pro³³ recombinant fragments to specifically capture its corresponding HPA-1 antibody.  Cells from a human B cell line (HHKB), homozygous for HLA-DRB3*0101, were pulsed with the recombinant PSI domain fragment of GPIIIa expressing the HPA-1a antigen.  MHC class II/peptide complexes were isolated from the pulsed cells using an immunoaffinity column.  A nested set of naturally processed and presented HPA-1a derived peptides, each containing the residues Trp²⁵ – Leu³³ core epitope was identified.  For the first time a naturally processed and presented HPA-1a peptide that spans the HPA-1a polymorphism has been identified, bound to the class II molecule encoded by HLA-DRB3*0101.  The efficient processing and presentation of this peptide, which includes the putative dominant Th epitope, is likely to be an important contributory factor in the immunogenicity of HPA-1a.  Such peptides may also provide the basis for novel treatments to tolerise the corresponding Th response in HPA-1b1b women at risk of NAIT with an HPA-1a-positive fetus.