An attempt to resolve all the various proteins in a single human cell type by two-dimensional electrophoresis: I. Extraction of all cell proteins.

A concept is presented for estimation of the total number of different proteins in a single human cell type (exemplified here by Hep cells) by use of two-dimensional electrophoresis (2DE). This concept includes three problems, the first, investigated in this study, being the transfer of all protein species of the cells into a sample useful for separation by 2DE. Five different extraction media containing--in various combinations--urea, Nonidet P-40, Zwittergent, mercaptoethanol, dithiothreitol, and sodium dodecyl sulfate were used step by step in three different extraction procedures to extract the cell proteins.

Separation of polychlorinated biphenyls (Aroclor 1254) by high-performance liquid chromatography.

The separation of Aroclor 1254 on various reversed-phase columns was investigated. The results show that the cyano- and phenyl-bonded columns performed poorly, but C18 bonded columns gave better results. It was also found that the absorption wavelength at which the effluent is monitored has great effect on the detection of these isomers.

Analysis of two-dimensional protein patterns from mouse embryos with different trisomies.

Two-dimensional protein patterns from whole mouse embryos with different trisomies (Ts) (Ts1, -12, -14, -19) and from different organs (normal or malformed) and developmental stages of Ts12 embryos were analyzed by comparison with control patterns. Quantitatively altered proteins were found, and a portion of these (21/approximately equal to 1,000, average) was attributable to the Ts conditions. Most of these variants were found always (regularly) in Ts embryos.

Two-dimensional electrophoresis of soluble and structure-bound proteins from cultured human fibroblasts and hair root cells: qualitative and quantitative variation.

Proteins from cultured human fibroblasts and native human hair root cells were investigated using the two-dimensional electrophoresis (2DE) technique. Cell material from 35 different healthy persons was examined. Proteins of different sources were separated: total proteins of fibroblasts (12 cell lines), soluble proteins of fibroblasts (12 cell lines), structure-bound proteins of fibroblasts (eight cell lines) and soluble proteins of hair root cells (12 subjects).

Teratogenic effects of cyproterone acetate and medroxyprogesterone treatment during the pre- and postimplantation period of mouse embryos. II. Cyproterone acetate and medroxyprogesterone acetate treatment before implantation in vivo and in vitro.

The study was performed to investigate direct embryotoxic effects of maternal progestin treatment during the preimplantation period. In the first experiment pregnant mice received a single subcutaneous injection of either cyproterone acetate (CA) or medroxyprogesterone acetate (MPA) on day 2 of pregnancy (5-600 mg/kg). In a second experiment four-cell embryos were exposed to CA or MPA in vitro (3 or 30 micrograms/ml medium).

Analysis of protein patterns in two-dimensional gels of cultured human cells with trisomy 21.

The occurrence of one chromosome of the cell in triplicate (trisomy, Ts) should increase the amount of all cell proteins coded by genes located on this chromosome. Many other proteins should be altered in their quantity by regulator genes, present in a threefold dosage, and by several indirect effects of the trisomy. We used two-dimensional electrophoresis to investigate the effect of the human Ts 21 on the proteins.

Mutagenicity testing on non-selectively cloned Chinese hamster ovary cells with the protein-mapping method.

Chemical mutagenesis was studied on Chinese hamster ovary cells by protein mapping. Cell cultures were treated with methylnitrosourea and the cells were cloned in non-selective media. The proteins of single clones were separated by 2-dimensional electrophoresis and analysed for qualitative (electrophoretic mobility) and quantitative (staining intensity; presence/absence) changes in the protein patterns.

High-resolution protein mapping of human fibroblasts and hair root cells: a standardized reproducible procedure considering the effect of cell culture parameters.

The effect of different cell culture parameters on two-dimensional polypeptide maps of human fibroblasts is considered. Improved culture methods are introduced to get better resolution and reproducibility. Based on these investigations, highly standardized techniques for the protein mapping of this tissue and also of hair root cell lysates are presented.

Genetic variability of proteins from plasma membranes and cytosols of mouse organs.

The genetic variability of membrane proteins (structure-bound proteins) and cytosol proteins (water-soluble proteins) was investigated in two inbred strains of the mouse, C57BL/6J and DBA/2J. Membrane proteins and cytosol were isolated from the brain and liver of the mouse. The proteins were separated by two-dimensional electrophoresis.

High resolution protein mapping in fibroblast cell lines and hair roots from patients with genetic disease.

Protein patterns of cultured fibroblast and hair root lysates from healthy controls and patients with genetic diseases (Duchenne muscular dystrophy, Friedreich's ataxia, Marie's ataxia, Lesch-Nyhan syndrome, maple syrup urine disease, and trisomy 13, 18 and 21) were obtained with two-dimensional electrophoresis. The analysis of these patterns in 39 gels by visual comparison revealed differences in the presence and absence of 20 specific protein spots. However, this variability, which has been observed in healthy controls as well as in patients, could not provide a diagnosis for a specific genetic disease.

Analysis of inborn errors of metabolism and other genetic defects in human fibroblasts using two-dimensional polypeptide mapping.

A standardized technique for the two-dimensional polypeptide mapping of cultured human fibroblasts has been used for the study of cellular protein variations in healthy controls, patients with inborn errors of metabolism and some other genetic defects. The analysis of about 50 gels has established that this method is very reproducible and enables the examination of about 600 polypeptides in a single gel. The inter-individual variation has been rather low (about 3%).

Vacuum-ultraviolet spectral-irradiance calibrations: method and applications.

A method to determine the spectral irradiance of a radiation source in the vacuum ultraviolet through the use of recently developed spectral-radiance standards is described. The method has been applied between 138 and 310 nm, and the spectral irradiances of several different light sources have been measured on an absolute scale with estimated uncertainties less than 10%.

Seasat low-rate data system.

The Seasat low-rate data system is a distributed, nonreal-time, magnetic-tape system for information processing. Its function is to apply the necessary calibrations, corrections, and conversions to yield geophysically meaningful products from raw spacecraft telemetry data. It also provides a remotely accessible catalog of satellite data.

Isoelectric focusing and electrophoresis combined as a method for defining new point mutations in the mouse.

Isoelectric focusing followed by electrophoresis is a method that allows the survey of the expression of an enormous number of genes in one organism when this method is used for separation of complex protein solutions from the tissue of the organism (protein-mapping method). This makes it possible to use this method for developing a test system aiming at the detection of newly induced or spontaneous point mutations in mammals. A large number of loci could be tested on a relatively small number of individuals.

Effects of various cephalosporins on the proximal tubule of the human kidney.

Cephamandol 6.0 g, cephazolin 6.0 g or cephacetrile or cephalothin 8.

The protein-mapping-method employed to test for chemically induced point mutations in mice.

A test system for detecting point mutations chemically induced in the germ cells of mice has been proposed in the past. In the present investigation the effect of the mutagenic substance methylnitrosourea (MNU) in such a test system was studied. Male mice 10-12 weeks of age of the inbred strain BALB/c Han.

Gel isoelectric focusing of mouse lactate dehydrogenase: heterogeneity of the isoenzymes A4 and X4.

LDH of mouse organs (including testis) was investigated by isoelectric focusing in polyacrylamide gels. The number of LDH bands in this pattern considerably exceeds the five (six in testis) of the standard electrophoretic pattern. An attempt was made to identify these bands as tetrameric isoenzymes formed by random association of different subunits.

Protein mapping by combined isoelectric focusing and electrophoresis of mouse tissues. A novel approach to testing for induced point mutations in mammals.

The protein-mapping method which combines isoelectric focusing in acrylamide gel and gel electrophoresis was previously used mainly for the separation of plant proteins and human serum proteins. We investigated with this technique soluble proteins of mouse tissues (whole embryos, the liver of fetal and adult mice, kidneys) and the proteins of mouse serum. The technique was tested under a number of different conditions to find those best for our purpose; they may represent some general improvements in the method.