"QUAX)" STAIN 



205 



QUARTZ ROD TECHNIQUE 



"Quad" Stain. A recent modification of 

 this excellent orcein-aiizarine-Orange 

 Gphosphotungstic and phosphormolyb- 

 dic acid technique is given in detail 

 by Kornhauser, S. I., Stain Techn., 

 1945, 20, 33-35. 



Quartz Rod Technique for illuminating liv- 

 ing organs. — Written by Dr. M. H. 

 Knisely, Department of Anatomy, Uni- 

 versity of Chicago, September 6, 1946 — 

 The general purpose of this technique is 

 to permit direct microscopic study of 

 living internal organ in situ while main- 

 taining experimental conditions which 

 disturb the structures and processes to 

 be observed as little as possible. Like 

 all techniques it has advantages and 

 limitations; there are specific purposes 

 for which it works well, and purposes 

 for which it has not yet worked at all. 

 The method makes it possible to study 

 at 32 to about 600 diameters magnifica- 

 tion those living structures whose 

 colors and/or indices or refraction differ 

 from those of adjacent structures. 

 With quartz rods we can illuminate for 

 examination under nearly normal condi- 

 tions many living tissues and organs 

 which heretofore have been inacces- 

 sible. The method depends upon two 

 physical principles: 



1. Conducting light from a suitably 

 intense source directly to the structures 

 to be studied by way of a fused quartz 

 rod. Clean, smooth transparent rods 

 conduct light around bends and turns 

 by internal reflection almost like a hose 

 conducts water. With suitably shaped 

 rods brilliant illumination of relatively 

 inaccessible structures is relatively 

 easy. As evidence of intensity, with a 

 750 watt T-12 tungsten filament bulb 

 and a two foot length of 7 millimeter 

 rod, so much light can be sent into a 

 microscope objective that one can 

 scarcely look into the ocular. Lesser 

 degrees of intensity are of course easily 

 obtainable. Substitutes for quartz 

 rods have been suggested and occasion- 

 ally used. (Cole, E. C, Science, 1938, 

 87, 396-398. Williams, R. G., Anat. 

 Rec, 1941, 79, 263-270). We have 

 tested several. No substitute has yet 

 proven as effective for illuminating 

 living tissues as fused quartz itself. 



2. Maintaining the normal tempera- 

 tures of intensely illuminated living 

 structures with a slowly flowing isotonic 

 isothermal wash solution. It is im- 

 possible to illuminate a non-transparent 

 structure without heating it at the same 

 time. The color of an object, even a 

 translucent object, as seen by either 

 transmitted or reflected light is due to 

 the patterns of the wave lengths which 

 reach the eye after parts of the incident 

 light are "absorbed", and the word ab- 



sorbed here means transformed into 

 fieut by and within the substance of the 

 object seen. Light filters as commonly 

 used between light source and illumi- 

 nated object can shelter a specimen 

 from the wave lengths which the filters 

 absorb, but they do not alter the fact 

 that a part of the light energy which 

 passes the filters and falls on the speci- 

 men is always transformed into heat 

 within the specimen by the materials of 

 the specimen itself. Hence, in con- 

 tinuously illuminating a living object 

 heat is sinmltaneously developed in it 

 at a constant rate. If the specimen is 

 small, thin, and very nearly transparent 

 and if its illumination is dim, the small 

 amount of continuously produced heat 

 may be transferred to adjacent objects 

 so rapidly that the temperature of the 

 specimen never rises enough to interfere 

 with its normal functioning. However, 

 in illuminating relatively thick trans- 

 lucent structures such as frog kidney or 

 liver, or mammalian spleens, brightly 

 enough for microscopic study, heat is 

 developed in the illuminated structures 

 faster than it can be removed without 

 assistance. To remove this heat a, flow- 

 ing solution at constant temperature is 

 applied to the illuminated tissue, either 

 through sets of glass tubes, or more 

 recently through hollow tipped quartz 

 rods which deliver both light and flow- 

 ing solution precisely to the selected 

 portions of the specimen. The fluid 

 delivered to the tissue must of course 

 be isothermal and isotonic with the 

 fluid which normally bathes it, i.e. plain 

 water at room temperature is used to 

 carry heat from frog skin or tongue, 

 amphibian Ringer's solution at room 

 temperature to carry heat from frog 

 kidney, and mammalian Ringer's at 

 mammalian bods'" temperature to carry 

 heat from monkey omentum. On ac- 

 count of the high specific heat of water 

 the flowing solution can take up the 

 heat as fast as it is produced with but 

 little change in its own temperature; 

 each small portion of flowing solution 

 is warmed but little as it passes through, 

 then leaves the illuminated field. By 

 these physical mechanisms the heat in- 

 escapably developed by transformation 

 of light energy is removed as fast as it 

 is produced and in consequence the 

 temperature of the illuminated tissue 

 does not rise. 



Thus far in a series of careful tests we 

 have found no visible change in any 

 structure and/or process within any 

 living tissue or organ in response either 

 to a sudden change from dim to intense 

 illumination or to hours of continuous 

 intense illumination, provided the tem- 

 perature of the illuminated specimen 



