Glycogen In Sections Research Articles

A modified PAS stain combined with immunofluorescence for quantitative analyses of glycogen in muscle sections.

Simultaneous analyses of glycogen in sections with other subcellular constituents within the same section will provide detailed information on glycogen deposition and the processes involved. To date, staining protocols for quantitative glycogen analyses together with immunofluorescence in the same section are lacking. We aimed to: (1) optimise PAS staining for combination with immunofluorescence, (2) perform quantitative glycogen analyses in tissue sections, (3) evaluate the effect of section thickness on PAS-derived data and (4) examine if semiquantitative glycogen data were convertible to genuine glycogen values. Conventional PAS was successfully modified for combined use with immunofluorescence. Transmitted light microscopic examination of glycogen was successfully followed by semiquantification of glycogen using microdensitometry. Semiquantitative data correlated perfectly with glycogen content measured biochemically in the same sample (r2=0.993, P<0.001). Using a calibration curve (r2=0.945, P<0.001) derived from a custom-made external standard with incremental glycogen content, we converted the semiquantitative data to genuine glycogen values. The converted semiquantitative data were comparable with the glycogen values assessed biochemically (P=0.786). In addition we showed that for valid comparison of glycogen content between sections, thickness should remain constant. In conclusion, the novel protocol permits the combined use of PAS with immunofluorescence and shows valid conversion of data obtained by microdensitometry to genuine glycogen data.

Preservation of glycogen in decalcified hard tissues.

Various fixatives and fixation procedures were tested to evaluate their effects on the preservation of glycogen in sections of decalcified hard tissues. Lower jaws from 1-day-old rats were chosen for the observations. An aqueous solution of glutaraldehyde showed poor preservation of glycogen in the tissues even when employed in the perfusion procedure. Freeze-drying and formaldehyde vapour fixation preserved it much better, but glycogen was still lost to some extent. Freeze-substitution with acetone and various alcoholic fixatives gave a poor result, unless the tissues were fixed with cyanuric chloride. Cyanuric chloride in methanol containing N-methyl morphorine was the best fixative for the preservation of glycogen in the sections. A combination of freeze-substitution with the cyanuric chloride solution, decalcification with the Jenkins's fluid, and subsequent double-embedding in celloidin and paraffin was recommendable for an excellent glycogen preservation.

Histochemical study of the glycogen content of the uterus and placenta of white rats at different stages of gestation

Histochemical studies of the distribution of glycogen in sections of the placenta and the uterus of white rats were made, using the Bauer-Feulgen technique, on material from the 6th, 10th, 15th, 17th, 18th, and 19th days of pregnancy. Only traces of glycogen were found in the non-gravid uterus. During pregnancy there is a correlation between the phosphatase, in particular the acid phosphatase, activities of uterine and placental tissues and their glycogen contents. A gradual rise is seen in the glycogen content of the decidua basalis, the syncytial trophoblast of the labyrinths, and the giant cells of the trophoblast, viz, in those components of the fetal adnexa which play the most important part in the carbohydrate metabolism of the embryo. Only traces of glycogen are to be seen in the vitelline placenta.

Ester Wax as a Medium for Embedding Tissue for the Histological Demonstration of Glycogen

1. The effect of paraffin wax embedding on the lability of glycogen in sections has been tested using as material (a) plerocercoid larvae of Ligula intestinalis; (b) rabbit liver. 2. It was found that glycogen in tissues was very impermeable to wax, and where large masses of glycogen occur prolonged embedding is essential. 3. Improperly embedded material lost glycogen easily; it was concluded that this loss takes place during the processes of cutting and flattening sections. 4. The effect of prolonged embedding was to make tissue hard and refractive. This difficulty was overcome by embedding in Steedman's ester wax. 5. Ester wax blocks allowed thin sections of very hard material to be cut with ease. 6. Ester wax sections brought down to water did not lose glycogen even after standing in water for 7 days. 7. It is emphasized that glycogen exists in two forms: an insoluble desmo- form and a soluble lyo- form. It is suggested that the latter does not dissolve from sections in water because its highly branched molecule is held by the coagulated protein network.