A new technique for autoradiography of whole-body sections mounted on glass slides.

Abstract

A simple and new technique was developed to ensure intimate contact of film with whole-body sections mounted on glass slides. Mice were injected with 125I-labeled insulin intravenousely.Whole-body frozen sections were prepared using adhesive tape along with a piece of dry paper and transferred to cold glass slides coated with egg albumin and glycerin. After freeze drying, a slide-mounted section was placed in a plastic bag, and brought into contact with the film. The bag was subsequently evacuated and sealed by a vacuum sealer. By this method, film maximum contact could be maintained with the section during exposure. Following development of the film, the section was stained and used for histological identification of tissues in the autoradiograph.A method for preparing whole-body sections for autoradiography and staining was previously reported (6), and is essentially the same as that developed by Kawamoto and Shimizu (3) who used adhesive tape with dry paper in place of cellulose backed (7) and cotton backed (10, 11) adhesive tape. These methods make possible the mounting of whole-body cryo-sections on the glass slides. The sections are pressed against X-ray films and stored for several days or weeks. To ensure that contact is even and uniform, the section and film are exposed in a press at slight pressure, generally using standard X-ray cassettes. However, in some cases, there is uneven contact between the section and film.In this paper, a simple and new method for autoradiography of whole-body sections mounted on glass slides is described. MATERIALS AND METHODSAnimals: Adult male mice of the ICR strain, weighing approximately 25 g, were fasted overnight with free access to water. Labeled compound: Porcine, 125I-TyrA24insulin (specific activity 81.4 TBq/mmol) purchased from New England Nuclear, Boston, MA, USA was used as the tracer for whole-body autoradiography and dissolved in Ringer's solution (462.5 kBq/ml). Sectioning: Radioactive insulin (185 kBq) was injected into the tail vein followed by immediately anaesthetizing the animals by intraperitoneal injection of sgdium pentobarbital. Three min later, the mice were purfused with Ringer's solution for 1 min through the left ventricle. The incised region for perfusion was then covered with 3% carboxymethyl cellulose (CMC) frozen by dry-ice powder, and the entire body of an animal was frozen in a mixture of dry-ice and acetone. The frozen animal was wiped with tissue paper, painted with 3% CMC, mounted on microtome stage with 6% CMC, and frozen by dry-ice snow. It was then placed on a microtome in a cryostat (LKB-2250, Sweden) adjusted at -20°C. Frozen whole-body sagittal sections (20 pm thick) of the mouse were prepared and mounted on cold glass slides previously coated with egg albumin and a glycerine mixture according to the methods of Shimono et al. (6). The sections were freeze-dried in the cryostat and brought to room temperature in a desiccator containing silica gel. Whole-body autoradiography: Each slide-mounted dry section to be brought into contac...

Histologic distribution of insulin and glucagon receptors

“…D19 (Kodak, Rochester, NY) is a good emulsion developer because it shows a large range of gray levels and results in good proportionality between absorbance and radioactivity (54). There is an alternative method of whole body radioautography in which whole body cryosections on glass slides are used rather than cryosections adhering to adhesive tape (55).…”