Comparative genomic hybridisation for the analysis of chromosomal imbalances in solid tumours and haematological malignancies

Comparative genomic hybridisation (CGH) is based on a two-colour, competitive fluorescence in situ hybridisation of differentially labelled tumour and reference DNA to normal metaphase chromosomes. This new technology has made a great impact in molecular tumour pathology due to its possible application to archival specimens and the ability to create copy number karyotypes throughout the whole genome from very small amounts of DNA. If chromosomal imbalances can be correlated with a etiological and clinical features of tumours, CGH could be able to provide new prognostic and diagnostic criteria. CGH findings further provide starting points for the molecular genetic characterisation of altered chromosomal regions harbouring yet unidentified genes involved in tumorigenesis and tumour progression. An overview of the results of published CGH studies on solid tumours and haematological malignancies is presented. Methodological limitations of the CGH technology are reported, as well as future developments which will improve its use in routine analysis.     

An evaluation of evolutionary constraints on microsatellite loci using null alleles

A test to evaluate constraints on the evolution of single microsatellite loci is described. The test assumes that microsatellite alleles that share the same flanking sequence constitute a series of alleles with a common descent that is distinct from alleles with a mutation in the flanking sequence. Thus two or more different series of alleles at a given locus represent the outcomes of different evolutionary processes. The higher rate of mutations within the repeat region (10(-3) or 10(-4)) compared with that of insertion/deletion or point mutations in adjacent flanking regions (10(-9)) or with that of recombination between the repeat and the point mutation (10(-6) for sequences 100 bp long) provides the rationale for this assumption. Using a two-phase, stepwise mutation model we simulated the evolution of a number of independent series of alleles and constructed the distributions of two similarity indices between pairs of these allele series. Applying this approach to empirical data from locus AG2H46 of Anopheles gambiae resulted in a significant excess of similarity between the main and the null series, indicating that constraints affect allele distribution in this locus. Practical considerations of the test are discussed.     

Sample handling for proteome analysis

The main factor limiting the sensitivity range for the identification of proteins isolated by two-dimensional (2-D) electrophoresis is sample handling: protein detection limits on the gel, losses during extraction and digestion, as well as interference of gel contaminants and detergents with the mass spectrometry (MS) detection increasing background noise. At the one hundred picomole level, losses are fairly negligible but when the amounts drop below 1 picomole (and subfemtomole peptide detection limits have been reported recently by MS), the losses become a critical point. In order to extend proteome analysis to include very low copy number proteins, methods must be developed to minimize losses and handling steps, maximize digestion and extraction yields, as well as to lower chemical noise. We present several methods that we have developed in our laboratory to: (i) increase the amount of material available in a sodium dodecyl sulfate (SDS)-free form which does not require staining, (ii) increase protein extraction and digestion yields and lower the contamination by autoproteolytic products, and (iii) allow direct modification of the peptide mixture to generate sequence tags.     

Phosphate und organische Phosphorverbindungen in Lebensmitteln Vl. Mitteilung Auftrennung und Kennzeiehnung von Phosphopeptiden aus enzymatischen Caseinhydrolysaten

Zusammenfassung Aus Abbauprodukten, wie sie bei der enzymatischen Spaltung des Caseins im Verlauf der Käsereifung auftreten, wurden Phosphopeptidgemische abgetrennt, in einzelne Komponenten zerlegt und auf ihren Aufbau hin untersucht. Hierbei erwies sich die kontinuierliche Ablenkungselektrophorese als besonders fruchtbar. Es gelang auf verschiedenen Wegen charakteristische Komponenten zu isolieren, die durch Phosphorgehalt, Aminosäurezusammensetzung, Kettenlänge und Summenformel gekennzeichnet wurden. Neben phosphorreichen Verbindungen wurden phosphorarme Spaltprodukte abgetrennt; unterschiedliches Verhalten beider Gruppen gegen Peptidasen und Phosphatasen und deren enzymatischer Abbauweg wurden diskutiert. Den Untersuchungen zufolge liefern hochmolekulare Phosphopeptide stark saure, das gesamte Phosphoserin und einen Großteil der Glutaminsäure enthaltende Kerne die durch wechselweise Einwirkung von Phosphatasen und Peptidasen gespalten werden können, wobei die relativ geringe Phosphatasenaktivität den limitierenden Faktor darstellt. Die Ergebnisse führen zu Vorstellungen über die Phosphorgruppierung im Caseinmolekül, insbesondere auch über die Lokalisierung des Phosphoserins im Peptidverband des Caseins.     

Digitalis effects on the electrocardiogramm: ergometric studies in healthy subjects and patients with coronary artery disease (author's transl)

Therapeutically effective doses of beta-acetyldigoxin in healthy subjects produced no or only minor S-T depression both at rest and under maximal cycle-ergometry, in individual instances the S-T segment never being depressed by more than 0.1 mV. On the other hand, similar exercise in patients with latent coronary insufficiency produced marked, significantly and dose-dependent S-T segment depressions under increasing digitalisation within the therapeutic range, and in some of the patients there was also decreased exercise tolerance before anginal symptoms. It is concluded that the so-called digitalis effect is definitely of diagnostic significance in the recognition of latent coronary insufficiency, contrary to previously held belief.