Profiling antibodies to class II HLA in transplant patient sera.

Immunizing events including pregnancy, transfusions, and transplantation promote strong alloantibody responses to HLA. Such alloantibodies to HLA preclude organ transplantation, foster hyperacute rejection, and contribute to chronic transplant failure. Diagnostic antibody-screening assays detect alloreactive antibodies, yet key attributes including antibody concentration and isotype remain largely unexplored.

Natural epitope variants of the hepatitis C virus impair cytotoxic T lymphocyte activity.

Aim: To understand how interactions between hepatitis C virus (HCV) and the host's immune system might lead to viral persistence or effective elimination of HCV.

Methods: Nucleotides 3519-3935 of the non-structural 3 (NS3) region were amplified by using reverse transcription polymerase chain reaction (PCR). PCR products of the HCV NS3 regions were integrated into a PCR((R)) T7TOPO((R)) TA vector and then sequenced in both directions using an automated DNA sequencer.

Identification of breast cancer peptide epitopes presented by HLA-A*0201.

Cellular immune mechanisms detect and destroy cancerous and infected cells via the human leukocyte antigen (HLA) class I molecules that present peptides of intracellular origin on the surface of all nucleated cells. The identification of novel, tumor-specific epitopes is a critical step in the development of immunotherapeutics for breast cancer. To directly identify peptide epitopes unique to cancerous cells, secreted human class I HLA molecules (sHLA) were constructed by deletion of the transmembrane and cytoplasmic domain of HLA A*0201.

Critical factors in the development of fluorescence polarization-based peptide binding assays: an equilibrium study monitoring specific peptide binding to soluble HLA-A*0201.

There is currently a significant interest in the identification and validation of HLA-restricted CTL epitopes, which are thought to have important implications for the development of preventive and/or therapeutic applications in bacterial or viral infections, autoimmune diseases, and cancer. To better facilitate epitope discovery and validation, we present a cell- and radioisotope-free HLA-A*0201 assay system which relies upon fluorescence polarization. The assay has the advantage of allowing real-time measurements in solution without separation steps.

MHC class I-associated peptides identified from normal platelets and from individuals with idiopathic thrombocytopenic purpura.

Major histocompatibility complex (MHC) class I molecules bind and display peptide antigens on the cell surface. CD8(+) T lymphocytes recognize peptides in association with class I proteins to initiate a cytotoxic immune response. To understand the specificity of such immune responses and to facilitate the development of therapies for disease, it is important to identify MHC-presented peptides.

Development and validation of a fluorescence polarization-based competitive peptide-binding assay for HLA-A*0201--a new tool for epitope discovery.

Various approaches are currently proposed to successfully develop therapies for the prevention and treatment of infectious diseases and cancer. One of the most promising approaches is the development of vaccines that elicit cytotoxic T lymphocyte (CTL) responses. Consequently, identification and exact definition of molecular parameters involved in peptide-MHC class-I interactions of putative CTL epitopes are of prime importance for the development of immunomodulating compounds.

Real-time measurement of in vitro peptide binding to soluble HLA-A*0201 by fluorescence polarization.

Measuring the interaction of class I human leukocyte antigens (HLA) and their peptide epitopes acts as a guide for the development of vaccines, diagnostics, and immune-based therapies. Here, we report the development of a sensitive biochemical assay that relies upon fluorescence polarization to indicate peptide interactions with recombinant soluble HLA proteins. It is a cell- and radioisotope-free assay that has the advantage of allowing the direct, real-time measurement of the ratio between free and bound peptide ligand in solution without separation steps.

Toward a definition of self: proteomic evaluation of the class I peptide repertoire.

MHC class I molecules present host- and pathogen-derived peptides for immune surveillance. Much attention is given to the search for viral and tumor nonself peptide epitopes, yet the question remains, "What is self?" Analyses of Edman motifs and of small sets of individual peptides suggest that the class I self repertoire consists of thousands of different peptides. However, there exists no systematic characterization of this self-peptide backdrop, causing the definition of class I-presented self to remain largely hypothetical.