ABSTRACT
DNA fragments from apoptotic cells crossing the renal barrier retain their matrix functions, which allows PCR identification of mutant sequences in excreted DNA. We investigated the possibility of detecting k-ras mutations in urinary DNA of tumor patients (colon cancer). In some patients with k-ras codon 12 mutations in tumor cell DNA the same changes were detected in the urinary DNA. The possibility of using this approach for early diagnosis and monitoring of tumors is discussed.
Subject(s)
Colonic Neoplasms/genetics , Colonic Neoplasms/urine , Genes, ras , Mutation , Codon , Colonic Neoplasms/diagnosis , Humans , Polymerase Chain ReactionABSTRACT
Rabbit antibodies against rat thymus and liver chromatin are obtained. Antithymus IgG are found to interact only with homologous chromatin, while antiliver IgG interact with both liver and thymus chromatin. After preincubation of antiliver IgG with thymus chromatin the antibodies interact with homologous chromatin only. Thus chromatin of both organs contains tissue-specific proteins. Antiliver and antithymus IgG are used to investigate a distribution of tissue-specific and tissue-non-specific immunogenic proteins in thymocyte nuclei. It is shown that these proteins are practically lacking in nuclear membranes, matrix, nuclear sap, nucleous chromatin and do not participate in attaching DNA to the nuclear matrix.
Subject(s)
Cell Nucleus/ultrastructure , Chromosomal Proteins, Non-Histone/immunology , Animals , Cell Fractionation , Cell Nucleus/immunology , Chromatin/immunology , Epitopes , Liver/immunology , Male , Nuclear Envelope/immunology , Rats , Thymus Gland/immunologyABSTRACT
Localization of immunogenic tissue-specific proteins in chromatin regions, hypersensitive to endogenous nucleases, has been studied using rabbit antibodies against rat thymus chromatin. It is shown that the first 1-2,5% of the chromatin (calculating on DNA), released by Mg2+-, Mn2+- and Ca2+/Mg2+-dependent nuclear endonucleases are drastically enriched in tissue-specific antigenic determinants. The released chromatin fractions are found to contain a heterogeneous set of nonhistone proteins and are deficient in histones. The cleavage of nuclear DNA by endogenous acidic nuclease, independent on bivalent ions, resulted in a significantly less enrichment of the released fractions with immunogenic proteins.