I04 PATTEN AND REDENBAUGH. [Vol. XVI. 



of the caudal spine. These crabs had been imbedded in cel- 

 loidin, sectioned, and stained in borax carmine, or in haema- 

 toxylin and picro-acid-fuchsin. 



The histology of the heart, pericardium, alary muscles, etc., 

 was studied by means of sections and pieces, excised, stained, 

 and mounted. The heart with the neighboring tissues was cut 

 out and hardened in Flemming's strong solution. Heidenhain's 

 iron haematoxylin and eosin gave in most cases the best stain 

 for sections ; Kleinenberg's haematoxylin and eosin were, also, 

 found very satisfactory. 



The nerves upon the heart were made out by means of 

 Lowit's gold chloride method, and the results verified by the 

 methylen blue method. 



Gold Chloride Method. — The hearts of young Limuli, 

 about 5 inches in length, were dissected out, slit open along 

 the ventral side, spread out, and treated with formic acid (one 

 volume of water to one of the acid) until the tissues became 

 transparent. They were then put for fifteen minutes or half an 

 hour in i per cent gold chloride solution, and afterwards left 

 in the dark for twenty-four hours in dilute formic acid (one 

 part acid to three of water). Finally, they were put into strong 

 formic acid, and left for three or four days in the dark, until the 

 muscles on the inside of the heart had macerated to such an extent 

 that they could be washed away by careful manipulations of 

 the pipette, leaving the nerve plexus intact. The hearts 

 were then spread out on slides and mounted in glycerine, acidu- 

 lated with formic acid. 



Methylen Blue Method. — Methylen blue was used in various 

 ways. Some very good stains of the nerves in the appendages 

 and upon the heart were obtained by injecting small crabs with 

 a I per cent aqueous solution. 



The injection method proved successful in only a small per- 

 centage of cases. More uniform results were obtained by 

 immersing portions of the crabs in a solution of methylen blue 

 in serum, a method first used by Dr. Patten. The serum was 

 obtained from the clotted blood of large Limuli. Enough of the 

 stain was dissolved in the serum to give it a clear blue color, 

 and the solution was kept well oxygenated by forcing air into 



