Extraction Of Glycogen Research Articles

Radioautographic comparison of the uptake of galactose-H and glucose-H3 in the golgi region of various cells secreting glycoproteins or mucopolysaccharides.

The radioautographic distribution of the label of galactose-H(3) was compared with that of glucose-H(3) in a series of secretory cells of the rat. Whereas the glucose label appeared in all mucous cells, the galactose label was incorporated only into certain mucous cells. Whenever either label was incorporated, however, it was located first in the Golgi region and later in the secretion product, mucus. Several lines of evidence, including extraction of glucose label with peracetic acid-beta glucuronidase, indicated that the material synthesized in the Golgi region was glycoprotein in nature. In chondrocytes, both the galactose and the glucose label appeared first in the Golgi region and later in cartilage matrix; extraction of glucose label with hyaluronidase indicated that much of it consisted of mucopolysaccharide. In all secretory cells, the extraction of glycogen by amylase had no effect on Golgi radioactivity. Such extraction did not eliminate the scattered cytoplasmic label also seen after glucose-H(3) injection, but completely eliminated that seen after galactose-H(3). Consequently, the galactose-H(3) label in the Golgi region stood out more clearly, and was detected in many cells: pancreas, liver, epididymis, and intestinal columnar cells. In the latter, label later appeared in the surface coat. Thus, radioautography after injection of galactose-H(3), as after glucose-H(3), indicates that synthesis of complex carbohydrates takes place in the Golgi region of many secretory cells.

Studies of native glycogen isolated from synchronized Tetrahymena pyriformis (HSM).

Native glycogen was isolated from Tetrahymena pyriformis (HSM) by isopycnic centrifugation in cesium chloride density gradients. A density of 1.62 to 1.65 was isopycnic for glycogen. Most of the banded glycogen existed as 35 to 40 mmicro particles which had a sedimentation coefficient of 214. These particles were composed of aggregates of 2 to 3 mmicro spherical particles. Extraction of glycogen with hot alkali reduced the sedimentation coefficient of native glycogen from 214 to 64.7 and the particle diameter from approximately 40 to 20 mmicro and smaller. Cell division was synchronized by a repetitive 12-hour temperature cycle, and glycogen was measured at several times during the cell cycle. The temperature cycle consisted of 9.5 hours at 12 degrees C and 2.5 hours at 27 degrees C. Approximately 90 per cent of the cells divided during the last 1.5 hours of the warm period. The carbohydrate/protein ratio of cells at the end of the cold period was 0.27 and was reduced slightly during the warm period. Glucose was incorporated into glycogen during both periods, although the rate of incorporation was greater during the warm period. No preferential incorporation on the basis of particle size was noted. Incorporation was measured in both native glycogen and KOH-extracted glycogen. Tetrahymena glycogen is compared with rat liver glycogen previously isolated by similar procedures, and the significance of using combined rate-zonal and isopycnic centrifugation for isolating native glycogen is discussed.

Hydroxypropyl methacrylate, a new water-miscible embedding medium for electron microscopy.

Aldehyde-fixed rat tissues were variously dehydrated and impregnated in water-miscible 2-hydroxypropyl methacrylate (HPMA) containing 3 to 20 per cent water and 0.1 per cent alpha,alpha-azobisisobutyronitrile as catalyst for subsequent polymerization with ultraviolet light. Heat polymerization was also effective. Blocks of embedded tissue readily gave ultrathin sections, which required staining by uranyl acetate and/or lead stains to give adequate contrast for electron microscopy. The ultrastructure of pancreas, kidney, muscle, and intestine was well preserved by aldehyde fixation alone. Use of postfixation in osmium tetroxide or direct osmium tetroxide fixation was unsatisfactory. The fine structure of aldehyde-fixed liver from fasted rats was well preserved, whereas that from normal rats showed considerable disorganization and collapse, apparently because of extraction of glycogen during the embedding procedure. Enzymatic extraction of proteins by pepsin and of ribonucleic acid by ribonuclease after either formaldehyde or glutaraldehyde fixation was rapidly effected by direct treatment of ultrathin sections with solutions of the enzymes. In contrast, no digestion of chromatin by deoxyribonuclease could be detected. In spite of this present limitation, HPMA appears to have several advantages over earlier water-miscible embedding media for electron microscopy and to be particularly suitable for ultrastructural cytochemistry.

A Comparison of Products Obtained by Various Procedures Used for the Extraction of Glycogen

A Comparison of Products Obtained by Various Procedures Used for the Extraction of Glycogen

Extraction of glycogen with dimethyl sulfoxide

Glycogen has been isolated from rat livers by dimethyl sulfoxide extraction in a form which appears to be more representative of “native” glycogen than that isolated by other methods. Evidence has been obtained that the normal type of alkaline degradation might partially be the cause of the lower molecular weight of classical Pflüger preparations.

Formaldehyde extraction of glycogen from liver

Formaldehyde extraction of glycogen from liver