Silver hnpregnalion 177 



Ramon y Cajal type uses a single ammoniated silver impregnation fol- 

 lowed by reduction in pyrogallol, hydroquinone, or one of the amino 

 phenols. The Bielschowsky type uses double impregnation (silver nitrate 

 followed by ammoniated silver) and reduction in formalin. In (3), the 

 del Rio-Hortega method, sodium carl3onate (sometimes lithium car- 

 bonate) is used to form the precipitate and is followed by ammonia, the 

 chief product being silver diammino carbonate. (Kubie and Davidson, 

 1928) 



The reactions of these solutions to various conditions should be 

 luiderstood: 



1. The ammoniacal silver nitrate is most stable, least sensitive to 

 light, least readily reduced, and combines least easily with tissues. 

 Dining formalin reduction, a cloud of finely divided gray dust s!(^wly 

 develops and the staining is slow. This method is rarely used in prefer- 

 ence to other solutions. 



2. Ammoniacal silver hydroxide lies to the other extreines; it is least 

 stable, most sensitive to light, most readily reduced, and combines most 

 easily ^vith tissues. Almost instantly a heavy black cloud appears. It com- 

 bines almost at once with the tissue, the solution darkens quickly, and 

 a precipitate begins to form. Since silver nitrate is not reduced in an 

 acid solution, but reduces readily in an alkaline solution, it stands to 

 reason that the ammoniacal silver hydroxide solution is the most 

 sensitive of the three. 



3. The ammoniacal silver carbonate solution has properties lying be- 

 tween the abo\'e two. Its precipitate forms more proinptly than that of 

 ,(1) ammoniacal silver nitrate, but not as fast as that of (2) ammoniacal 

 silver hydroxide. It is darker than that of (1) but not to the degree of 

 (2). Its reaction begins within 5-10 minutes and reaches optimum 

 color before the solution begins to darken. Although solutions niniiber 

 2 and 3 are used interchangeably, the carbonate solution has the ad- 

 vantage that its hydroxide (OH)- ion concentration is not high enough 

 to render it as luistable and oversensitive as the hydroxide solution, and 

 the presence of buffer salts makes the reduction proceed steadily and 

 evenly. As the acid, HNO3, is formed during the reduction the buffer 

 absorbs it and blocks its effect; thus the reduction is not lessened. In 

 addition, the presence of CO3 ion buffers the formalin and prevents for- 

 mation of formic acid, also an effecti\ e stop to further reduction. Foot 

 (1929) buffered his formalin and prolonged the reducing action, making 

 his results darker and more intense. He ^varned, however, to keep the 

 buffer to a minimimi so the reaction will not become too intense. 

 Equimolar silver solutions produce the most uniform results. In most 



