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At least 19 recordsLinked to original sources
PubMed1977

Differentiation of rennet from other milk-clotting enzymes by polyacrylamide gel electrophoresis.

Polyacrylamide gel electrophoresis was used to differentiate animal rennet and other milk-clotting enzymes. After electrophoresis, the separated components were visualized by staining with aniline blue-black. Two prominent proteins were found in calf and bovine rennet, while only 1 major protein was observed in pepsin and enzymes of microbial origin. These patterns provided a basis for distinguishing animal rennet and the other enzymes as well as a means of identifying each type of enzyme by the characteristic pattern shown.

Animals↗
PubMed1976

The separation of plasma lipoproteins using gel electrofocusing and polyacrylamide gradient gel electrophoresis.

Polyacrylamide gel electrofocusing and gradient electrophoresis have been used to separate the lipoproteins in whole plasma and in fractions prepared by sequential flotation in the ultracentrifuge and by precipitation with dextran sulphate and manganous chloride. After the two-dimensional separation, high density lipoproteins appear as a zone showing noticeable heterogeneity with respect to both isoelectric point and molecular weight. Low density lipoproteins are resolved as a compact spot while very low density lipoproteins are visible as a long horizontal streak across the top of the electrophoresis gel. The implications of the technique for the analysis of lipoprotein patterns in pathological plasmas are discussed.

Electrophoresis, Agar Gel↗
PubMed1979

Glutamine synthetase regulation, adenylylation state, and strain specificity analyzed by polyacrylamide gel electrophoresis.

We used polyacrylamide gel electrophoresis to examine the regulation and adenylylation states of glutamine synthetases (GSs) from Escherichia coli (GS(E)) and Klebsiella aerogenes (GS(K)). In gels containing sodium dodecyl sulfate (SDS), we found that GS(K) had a mobility which differed significantly from that of GS(E). In addition, for both GS(K) and GS(E), adenylylated subunits (GS(K)-adenosine 5'-monophosphate [AMP] and GS(E)-AMP) had lesser mobilities in SDS gels than did the corresponding non-adenylylated subunits. The order of mobilities was GS(K)-AMP < GS(K) < GS(E)-AMP < GS(E). We were able to detect these mobility differences with purified and partially purified preparations of GS, crude cell extracts, and whole cell lysates. SDS gel electrophoresis thus provided a means of estimating the adenylylation state and the quantity of GS present independent of enzymatic activity measurements and of determining the strain origin. Using SDS gels, we showed that: (i) the constitutively produced GS in strains carrying the glnA4 allele was mostly adenylylated, (ii) the GS-like polypeptide produced by strains carrying the glnA51 allele was indistinguishable from wild-type GS(K), and (iii) strains carrying the glnA10 allele contained no polypeptide having the mobility of GS(K) or GS(K)-AMP. Using native polyacrylamide gels, we detected the increased amount of dodecameric GS present in cells grown under nitrogen limitation compared with cells grown under conditions of nitrogen excess. In native gels there was neither a significant difference in the mobilities of adenylylated and non-adenylylated GSs nor a GS-like protein in cells carrying the glnA10 allele.

Adenosine Monophosphate↗
PubMed1979

A viscosity model of polyacrylamide gel electrophoresis.

In current theories of polyacrylamide gel electrophoresis, the idea prevails that molecular sieving relies on different accessibility of volume fractions and of cross-sectional area fractions (denoted "pores") to different-sized ions due to the effect of "geometric exclusion". This correlates with the assumption that all elements of a polyacrylamide network occupy fixed and unchangeable positions thus forcing colliding macro-ions to diffuse laterally in order to find an "accessible pore" and to resume motion in direction of the electrical field. However, the alternative conception would be equally well justified, i.e. the assumption that polyacrylamide chains represent smooth obstacles cleared aside under the electrokinetic pressure of a macro-ion. This explanation would even be preferable with respect to the molecular sieving effects occurring in solutions of "liquid polyacrylamide". Yet no theory exists as to describe such effects in quantitative terms. In the present article, a parameter is defined and discussed, which can be estimated by experiment, and which seems to be apt to characterize local resistivity of polymer structures against dislocation and deformation: the "fractional specific resistance". Definition of this parameter is based on the model of a "viscosity-emulsion" composed of two interpenetrating liquid compartments which are characterized by different levels of hydrodynamic friction and the spatial dimensions of which are inferred from Ogston's theory. This concept of "localized viscosity" may also serve as a link between theories of molecular sieving and of "macroscopic viscosity" of flexible polymers. The data of Morris, formerly taken as verifications of the "rigid-pore" concept, are now interpreted in terms of four factors responsible for sizediscrimination: collision frequency, duration of single contacts, size-dependent frictional force, and the extent of cooperation among fibres, due to crosslinking and to simultanous contacts of several fibres to a single macro-ion. Some functions relevant for problems of molecular weight determination by gel electrophoresis are discussed in relation to the suggested model.

Acrylamides↗
PubMed1979

Identification of initiation factors and ribosome-associated phosphoproteins by two-dimensional polyacrylamide gel electrophoresis.

A two-dimensional polyacrylamide gel electrophoresis procedure has been used to identify initiation factors rapidly in the high-salt-wash fraction from reticulocyte ribosomes. Initiation factors are identified by relative mobility and by co-electrophoresis with purified factors. A creatine phosphate/ATP/GTP/Pi exchange system is described which has been used to maintain [gamma-32P]ATP and [gamma-32P]GTP at constant specific activity in the cell-free protein-synthesizing system. Phosphorylated proteins associated with the protein-synthesizing complex have been identified using a combination of the two procedures. The salt-wash fraction contains eight major phosphorylated proteins and a number of minor ones. Two phosphorylated proteins are observed to comigrate with two of the three subunits of eukaryotic initiation factor 2 (eIF-2), the initiation factor involved in binding Met-tRNAf onto the 40-S subunit and promoting dissociation of 80-S ribosomes. eIF-4B, one of the proteins involved in binding mRNA to 40-S subunits is also phosphorylated. The remainder of phosphorylated proteins in the high-salt-wash fraction are not previously characterized initiation factors and have not been identified further. Two of the six phosphoproteins associated with the salt-washed ribosomes comigrate with ribosomal proteins; one is the major phosphorylated protein in 40-S ribosomal subunits, the other is an acidic protein.

Animals↗
PubMed1979

Rapid serogroup identification of Neisseria meningitidis by using antiserum agar: Prevalence of serotypes in a disease-free military population.

Nasopharyngeal cultures from 414 Marines were plated directly onto antiserum agar containing the antibiotics vancomycin, colistin, and nystatin for meningococcal isolation and serogroup identification. Meningococci were isolated from 267 Marines, giving a carrier prevalence of 64.5%. A total of 58% of the isolates could be placed into serogroups; of these 22.3% were group B, 4.7% were group C, 25.7% were group Y, 24.3% were group W135, and 23.0% were group 29E. No serogroup A organisms were recovered. Serotyping by agar gel double diffusion was performed on 148 strains. More than 70% of these strains were nontypable, and the disease-associated serotype 2 was present only in two group Y isolates. The same 148 isolates were also classified by major outer membrane protein patterns after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Polyacrylamide gel electrophoresis pattern IV was most common among all serogroups. The data demonstrate the effective use of antiserum agar for meningococcal surveillance and document the frequency of specific serotypes and polyacrylamide gel electrophoresis types among carrier isolates obtained from a nonrecruit military population.

Carrier State↗
PubMed1979

An evaluation of a simple polyacrylamide gel electrophoresis system for lipoproteins and its use for the characterisation of hyperlipoproteinaemia.

The use of polyacrylamide gel electrophoresis as a screening procedure for the characterisation of hyperlipoproteinaemia is described. Sixty-one samples were investigated and classified by four different methods: (1) chemical profile derived from visual appearance and estimation of total cholesterol and triglyceride; (2) 'SML' profile by nephelometry; (3) electrophoresis on agarose gel; and (4) electrophoresis on polyacrylamide gel. Polyacrylamide gel electrophoresis showed the closest correlation with the chemical profile when compared with the other two methods. Polyacrylamide gel electrophoresis was found to be a rapid, reliable, and satisfactory procedure for plasma lipoprotein phenotyping.

Blood Protein Electrophoresis↗
PubMed1977

System for simplified discontinuous-gradient polyacrylamide-gel electrophoresis.

We describe a modified polyacrylamide gel electrophoresis method with which plasma proteins are excellently resolved in 4 h. We made a three-layer discontinuous gradient gel with concentrations of 80,60, and 45 g of polyacrylamide per liter of tris(hydroxymethyl)aminomethane/ethylenediaminetetraacetic acid/borate buffer, pH 9.2. Time and material needed in the preparation of the gel was greatly decreased, a well-resolved (22-25 bands) pattern was obtained in 1 h, and the gel slabs were easier to remove from the cell than were gels prepared according to the manufacturer's specifications. We saw a marked shift in the higher polymers of haptoglobin type 2-2 toward the cathode in this system, and also a clearly resolved double band of alpha2-macroglobulin in whichthe more anodal band appears to represent alpha2-macroglobulin/protease complex.

Blood Protein Electrophoresis↗
PubMed1979

Microheterogeneity of rat, mouse and human alpha1-fetoprotein as revealed by polyacrylamide gel electrophoresis and by crossed immuno-affino-electrophoresis with different lectins.

Polyacrylamide gel electrophoresis and crossed immuno-affino-electrophoresis with several free lectins have been used to characterize and to compare the molecular heterogeneity of rat, mouse and human alpha1-fetoproteins. Each alpha1-fetoprotein contains a variable number of electrophoretic variants depending on the gel porosity. In SDS electrophoresis, two molecular size populations are present in rat alpha1-fetoprotein (Mr = 74 000 and 72 000) and in mouse alpha1-fetoprotein (Mr = 73 000 and 72 000) but only one is observed in human alpha1-fetoprotein (Mr = 70 000). The crossed immuno-affino-electrophoresis patterns square with affinity chromatography results and reveal a marked and characteristic heterogeneity for the three alpha1-fetoprotein species with Concanavalin A, Ricinus communis and Lens culinaris lectins. No lectin-alpha-fetoprotein interaction is apparent with Ulex, Lotus and wheat germ lectins. Since similar patterns are obtained whether with purified alpha1-fetoprotein or with unfractionated fresh fetal sera, it is likely that this heterogeneity is not a consequence of artefactual molecular modifications arising during the purification procedure.

Animals↗
PubMed1978

Serum alkaline phosphatase (Al-Pase) isozyme in gastric and colonic cancer (using a simple thin layer polyacrylamide gel electrophoresis).

Using the simple thin layer polyacrylamide gel electrophoresis, serum alkaline phosphatase could be separated 5 isozyme bands in various digestive diseases, consisting of 54 cases of gastric cancer, 11 of colonic cancer, 12 of hepatoma, 4 of cholangioma, 14 of pancreatic cancer, 81 of benign hepatobilliary diseases, 13 of cancers of other organs and 61 of control. The obtained results were as follows: 1) The electrophoretic analysis of serum alkaline phosphatase showed the specific band remaining at the origin, already reported as "alkaline phosphatase O", in primary and metastatic cancer of the liver and cholelithiasis. On the contrary, alkaline phosphatase O was never found in gastric and colonic cancer without cholelithiasis. On the contrary, alkaline phosphatase O was never found in gastric and colonic cancer without cancerous metastasis to the liver, and it was also inclined to be positive with the progress of liver metastasis among them. 2) Intestinal alkaline phosphatase was usually found in higher frequency in blood group B and O than in the others, and it was apt to disappear in gastric or colonic cancer with an exacerbation of its cancerous lesions. 3) Heat-stable alkaline phosphatase was found in 10% of gastric or colonic cancer, all of which were histologically proved to be well differentiated adenocarcinoma.

Adenocarcinoma↗
PubMed1977

Preparative polyacrylamide gel electrophoresis purification of Clostridium perfringens enterotoxin.

Preparative polyacrylamide gel electrophoresis has been used to purify the enterotoxin of Clostridium perfringens from Sephadex G-100 extracts. Purified toxin of high specific activity was eluted in 1 to 3 h, depending upon the length of the acrylamide gel used. Recovery of biological activity with this technique ranged from 80 to 90%. The purity and physical characteristics of the toxin were similar to those previously reported for the protein purified by other methods. Use of preparative electrophoresis will enable the production of larger amounts of high-specific-activity toxin in a shorter time than other currently available procedures. This method was also used to isolate a form of enterotoxin that has a mobility, relative to bromophenol blue tracking dye, of 0.87 to 0.90 in 7% acrylamide gels.

Clostridium perfringens↗
PubMed1977

Evidence for recombination between N- and B-tropic murine leukemia viruses: analysis of three virion proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

We have sodium dodecyl sulfate-polyacrylamide gel electrophoresis to analyze the virion proteins of an N- and a B-tropic C-type virus derived from the BALB/c mouse and 21 putative recombinants, designated XLP-N viruses, obtained from seven crosses between these N- and B-tropic viruses. All the XLP-N viruses are N-tropic but posses the XC plaque morphology of their B-tropic virus parent. Three virion proteins, p15, p30, and gp70, of the parental viruses each differ in electrophoretic mobility. Two recombinants were found that possess a p15 that comigrates with p15 of the B virus; 19 possess a p15 that comigrates with N virus p15. Sixteen recombinants possess a gp70 that migrates like the gp70 of the B virus: four have gp70 with an electrophoretic mobility like that of the N virus gp70. All 21 recombinants possess a p30 that comigrates with p30 of their N virus parent. Given the origin and phenotype of XLP-N viruses, these results would seem to provide good evidence that these viruses are recombinants.

Electrophoresis, Polyacrylamide Gel↗
PubMed1978

Polyacrylamide gel electrophoresis of visna virus polypeptides isolated by agarose gel chromatography.

The proteins of visna are separated into nine major peaks by agarose gel chromatography in 6 M guanidine hydrochloride (GuHCl). The polypeptides in eack peak were isolated by acid precipitation and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The patterns of SDS-PAGE show that the excluded material from the GuHCl column contains an aggregate of 10 non-glycosylated polypeptides. It is shown that this aggregate represents virus substructures that are not completely solubilized by GuHCl. Two glycoproteins, gp175 and gp115, were isolated from the column eluate. The major glycoprotein gp115 was coeluted with P90, P68, and P61 in GuHCl 4. Each of the four major peaks (GuHCl 5 to 8) contains more than one nonglycosylated polypeptide. However, a small polypeptide, P12, can be isolated in a homogeneous form in the last peak, GuHCl 9. Analysis of the virus proteins (100 microgram) by SDS-PAGE shows that 20 radioactive bands can be recognized. During fractionation of the protein on agarose gel columns followed by analysis with SDS-PAGE, a number of minor polypeptides that were not detected before became clearly recognizable. Thus, the combined use of column chromatography and SDS-PAGE shows that visna virus is composed of 25 proteins.

Chromatography, Agarose↗
PubMed1978

[Comparative studies of avian mycoplasmas by flat gel polyacrylamide electrophoresis (author's transl)].

The phenol-acetic-acid extraced cell proteins of Mycoplasma (M.) and Acholeplasma (A.) reference strains (PG31 (M. gallisepticum), PG 16 (M. gallinarum), PG30 (M iners), 17529 (M. meleagridis), WVU 1853 (M. synoviae), 1340 (M. anatis), PG8 and PG9 (A. laidlawii), CKK (Serovar C), DD (Serovar D), WR1 (Serogroup F), 695 (Serogroup I) and 694 (Serogroup L) were anlysed by the flat gel polyacrylamide electrophoresis. With exception of PG8 and PG9 the Coomassie Blue-stained protein patterns show that each of the strains produced reproducible characteristic electrophoretic pattern by which the reference strains could be differentiated. However, before the question could be answered whether the procedure described is suitable to replace the serological species differentiation of avian mycoplasmas, serological and electrophoretic studies of a relevant number of field strain are necessary.

Animals↗