CHAP. II.] THE BLOOD. 85 



Methods of It must be stated in limine that great difference 



preparation exists in the ease with which this body can be obtained 



mogiobm 6 " in an odiously P ur e condition from the blood of 



different animals. By obviously pure condition we 



mean to indicate in the form of well-defined crystals. 



The principle upon which nearly all methods of separating oxy- 

 haemoglobin is based is the following : to effect the solution of the 

 haemoglobin of the red corpuscles either in the serum or in water 

 added to the separated corpuscles, and then either by the addition of 

 alcohol or by the agency of cold, or of both conjointly, to cause the 

 oxy-haemoglobin (which is sparingly soluble in dilute alcohol and at 

 low temperatures) to crystallize out. 



From the blood of some animals, and especially of the rat, oxy- 

 haemoglobin can be obtained for microscopic examination in two or 

 three minutes by receiving a drop of blood on a glass slide, adding to 

 it a drop of distilled water, and after mixing the two together cover- 

 ing with a microscopic covering-glass. Needle-shaped crystals form 

 almost at once. In order to separate considerable quantities of 

 oxy-haemoglobin or even to obtain large crystals for microscopic 

 observation it is advisable to follow one or other of the following 

 methods, of which the fifth and seventh are those which are most 

 easily carried out and most uniformly successful 1 . 



I. Blood is allowed to coagulate and the clot is allowed to contract so 

 as to separate the serum as completely as possible. (This end is naturally 

 most readily attained by employing a centrifugal apparatus.) The clot is 

 finely divided and then squeezed in a cloth; in this way the corpuscles 

 are separated from the fibrin of the clot. 



Water is added to the expressed grumous liquid (cruor) in quantity 

 equal to one or one and a half times its volume. A stream of oxygen gas is 

 now passed through the liquid for half an hour, and then a stream of C0 2 

 for ten minutes. After about five minutes a turbidity appears, crystals 

 commence to form, a large quantity separating out in the course of two 

 hours. By this method crystals are obtained only from the blood of the 

 guinea-pig, the rat, and the mouse. In order to obtain them from 

 the blood of the dog and other animals, before and during the passage of the 

 gases, dilute alcohol is added in small quantities to the fluid, which then 

 often yields a magma of crystals. Crystals thus obtained are, however, 

 not pure, and in order to separate them from adhering impurities they 

 must be washed with distilled water, or water holding a little alcohol in 

 solution, until the nitrate is no longer precipitated by solutions of silver 

 nitrate or of mercuric chloride 2 . 



Preyer has found that by merely passing air free from carbonic acid 

 through the defibrinated blood of the dog crystallization ensues, even 

 though the temperature of the blood be as high as 35 38 C. 



1 The description of the first six methods of preparing oxy-haemoglobin is based 

 upon that given by Preyer (in his admirable work entitled Die Blutkrystalle, Jena, 1871) 

 as abridged in Maly's Jahresbericht, Vol. i. (1873) p. 57; the seventh the Author 

 learned from Professor Kiihne ; he can highly recommend it. 



2 Lehmann, Ber. d. konigl. sachs. Ges. d. Wiss. zu Leipzig, 1853. Also Physio- 

 logical Chemistry. Translation by Day. Cavendish Society, 1854. Vol. in., p. 489 

 et seq. 



