Analytical separations by high-voltage paper electrophoresis. Amino acids in protein hydrolysates

Atfield, G.; Morris, C. J. O. R.

doi:10.1042/bj0810606

Cited by 283 publications

(69 citation statements)

References 14 publications

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“…This was investigated using the method described by Atfield and Morris (1961). Freeze-dried material (5-6 mg.) was dissolved in 2 ml.…”

Section: Amino Acid Compositionsmentioning

confidence: 99%

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The Perchloric Acid Soluble Basic and Acidic Proteins of the Cytoplasm: Variation in Cancer

Burston¹,

Tombs²,

Apsey³

et al. 1963

Br J Cancer

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A CHARACTERISTIC finding in malignant disease is an increase in the circulating glycoproteins, particularly the serum al-globulin. Such a change has frequently been reported in man (Winzler, 1960; Tombs, James and Maclagan, 1961) and in animals (Darcy, 1960). There has been some speculation as to the source and mechanism of production of the excess ax-glycoprotein. Suggestions have ranged from an increased production by the normal mechanism, or by injured or necrotic tissue, to a preferential utilisation of non-glycoproteins, leaving an apparent preponderance of glycoprotein in the serum (Shetlar, 1961).Good evidence that the liver is the normal source of serum acx-glycoproteins has only recently become available (Spiro, 1959;Hochwalde, Thorbecke and Asofsky, 1961). If, in cancer, the liver is stimulated to produce more ax-glycoprotein, it might be expected that the organ itself would contain more than it normally does. Very little evidence is available at present on the distribution of acidic glycoproteins within normal tissue or tumours. In a previous investigation (Tombs, Burston and Maclagan, 1962) chromatographic analysis of the cytoplasmic proteins of a number of organs showed that a glycoprotein very similar to if not identical with the serum a,-globulin was widely distributed; in particular it was present in the liver. Quantitative analysis of several human livers showed that in cancer the more basic and exclusively cytoplasmic proteins were depressed, and the acidic components, including the oc,-globulin were increased. This was so whether tumours were present in the liver or not. These results suggested that the tumour in some way caused the liver to produce more a,-globulin than it normally does. However, it was felt desirable to obtain more precise estimates of individual proteins than was possible from the somewhat heterogeneous chromatographic fractions. Thus in contrast with the previous experiment, which was a general analysis of all the cytoplasmic proteins, we sought a method of obtaining a,-globulin component in a pure state, and its relatively precise estimation. Perchloric acid was introduced (Winzler, Devor, Mehl and Smyth, 1948) as a reagent for the selective denaturation and precipitation of all the serum proteins except a glycoprotein fraction which remains soluble. While this fraction is by no means homogeneous (Biserte, Havez, Laturaze and Hayem-Levy, 1961) the major component is the a1-glycoprotein. It seemed reasonable to suppose that the use of perchloric acid on tissue extracts would give good preparations of the alglobulin. This was found to be so, but unexpectedly a number of basic cytoplasmic proteins were also found to be soluble in perchloric acid: these substances, one of which exhibits variation in cancer, have not previously been described.

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“…This was investigated using the method described by Atfield and Morris (1961). Freeze-dried material (5-6 mg.) was dissolved in 2 ml.…”

Section: Amino Acid Compositionsmentioning

confidence: 99%

“…of 6 N HCI and heated in a boiling water bath for 24 hours. The hydrolvsates were then prepared as described by Atfield and Morris (1961), and examined in a Shandon high voltage electrophoresis apparatus.…”

Section: Amino Acid Compositionsmentioning

confidence: 99%

The Perchloric Acid Soluble Basic and Acidic Proteins of the Cytoplasm: Variation in Cancer

Burston¹,

Tombs²,

Apsey³

et al. 1963

Br J Cancer

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“…After electrophoresis for 35 min at a voltage gradient of 100 V /cm on Whatman No.1 paper using acetic-formic acid buffer at pH 1·85 (Atfield and Morris 1961), the electrophoretograms were dried and developed by dipping in 0·25 % ninhydrin in acetone, followed by heating for 20 min at 60°C. The resultant bands were eluted in 75 % ethanol containing 12· 5 mg/l00 ml copper sulphate and the optical density measured at 510 nm in a spectrophotometer.…”

Section: (B) Analytical Proceduresmentioning

confidence: 99%

The Uterus of the Ewe II. Chemical Analysis of Uterine Fluid Collected by Cannulation

Rg¹

1973

Aust. Jnl. Of Bio. Sci.

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The chemical composition of uterine fluid collected from ewes cannulated by three different procedures was studied.Sodium and chloride were the major inorganic ions present. Smaller amounts of potassium, magnesium, calcium, ammonium, bicarbonate, and phosphate ions were also found. Orcinol-reactive carbohydrate was present in high concentration but only 1--4 % could be accounted for as the reducing sugar, glucose. Approximately 5 mM lactate was present in the uterine secretions.Protein concentration varied from 1 to 3 g/loo ml and some differences existed between uterine fluid and blood serum both in electrophoretic mobility and relative concentration of the various protein components. Uterine fluid also differed in amino acid composition from blood serum and the results suggest that uterine fluid is, at least in part, a true secretion and not merely a blood transudate.Differences in chemical composition between stages of the oestrous cycle were small and only minor differences were found between fluid collected from the uterine horn and that collected from the body of the uterus. By contrast the composition of fluid collected from ewes with a cannula inserted through the cervical canal differed considerably from that of fluid collected by the other two methods of cannulation and the former method may give a less reliable estimate of the true uterine environment.

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“…Tryptic digests were separated by 2-dimensional paper electrophoresis at high voltage in cooled tanks first at pH 6.5 [16] and secondly at pH 1.9 [17], and the radioactive peptides identified by autoradiography using X-ray films Kodirex from Kodak Co. (35 × 45 cm). For autoradiographic exposures at least 80 000 dpm were loaded and for preparative purposes for thiol group identification 50-100 nmol/peptide were loaded.…”

Section: Methodsmentioning

confidence: 99%

Isolation of cyanogen bromide and tryptic peptides containing the essential thiol groups from isolated myosin heads

Kunz¹,

Walser²,

Watterson³

et al. 1977

FEBS Letters

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Analytical separations by high-voltage paper electrophoresis. Amino acids in protein hydrolysates

Cited by 283 publications

References 14 publications

The Perchloric Acid Soluble Basic and Acidic Proteins of the Cytoplasm: Variation in Cancer

The Perchloric Acid Soluble Basic and Acidic Proteins of the Cytoplasm: Variation in Cancer

The Uterus of the Ewe II. Chemical Analysis of Uterine Fluid Collected by Cannulation

Isolation of cyanogen bromide and tryptic peptides containing the essential thiol groups from isolated myosin heads

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