B. Hamberger, T. Malmfors, C. Sachs
Feb 1, 1965
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1
Influential Citations
230
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Journal
Journal of Histochemistry and Cytochemistry
Abstract
Adrenergic mechanisms can now be studied directly at cellular and subcellular levels with the help c)f the sensitive fluorescence method c)f Falck and Hillarp for the histochemica! demonstration of certain catecholamines, e.g. the adrenergic transmitter. Briefly this method inve)lves the treatment of freeze-dried or air-dried tissues with formaldehyde gas derived from paraformaldehyde at 80#{176}C(for details, see Dahlstr#{246}m and Fuxe, Ada Physiol. Scand. 62: Suppl. 232, 1964; Norberg and Hamberger, ibid. 63: Suppl. 238, 1964). During this treatment catecholaniines are converted to intensely fluorescent 3, 4-dihydro isoquinolines (Corrodi and Hil!arp, Helv. Chini. Ada 46: 2425, 1963; 47: 911, 1964). This reaction requires the presence of water, but if too much water is present the amines or their fluorescent products can diffuse. Water for the reaction is derived frem three main sources: the tissue, the air initially enclosed in the reaction vessels, and the paraformaldeiiyde used. It is of great importance to standardize the freeze-drying (or air-drying) in such a way that. the pieces will have a low and fairly constant content of water. The dried pieces readily adsorb water from the air and must be handled in a dry atmosphere. Paraformaldehyde under these condi tions becomes the most important source of water. This water comes partly from pyrolysis of the polymer, but of greater importance is adsorbed water, which varies considerably depending for example on the storage of the paraformaldehyde. The present work reports the principles of a simple procedure for standardization of paraformaldehyde based on the finding that this substance takes up or loses water to constant levels when incubated at room temperature (20-22#{176}C)in an atmosphere of constant relative humidity. Incubations were performed in closed vessels containing aqueous solutions of sulfuric acid of varying density. Any relative humidity between 10 amid 90% can be obtained in this way (Handbook of Chemistry and Physics. 44th ed. Chemical * Supported by USPHS Grant (NB 02854-04), National Institute of Neurological Diseases and Blindness, and a grant from the Swedmsh Medical Research Council. Rubber Pumbhishiing Co., Cleveland, 1963). Amounts of adsorbed water were deterniiuied by using the Karl Fischer reagent (Mitchell and South , 4 quametry, Interscience, New York , 1948). A full account of these experiments will be pubhished in a forthcoming paper (Hamnberger, to l)e