454 BLOOD AND LYMPH. 



matter of fact, solutions of fibrinogen prepared as described some- 

 times clot, although much more slowly, when instead of a throm- 

 bin solution one adds a little calcium chlorid or a solution con- 

 taining calcium chlorid and sodium bicarbonate in about the 

 proportion found in a Ringer's mixture. This latter fact indicates 

 that the fibrinogen solution in such cases contains a trace of some 

 material from which thrombin may be produced . In all probability 

 this material is the antecedent form of thrombin, that is, so-called 

 prothroml^in, which, as we shall see, is converted to thrombin l)y 

 the action of calcium salts. 



Preparation and Properties of Thrombin. — Thrombin, or so- 

 called fibrin ferment, is prepared readily by the method first de- 

 scril^ed by Schmidt. Blood is allowed to clot, and the serum is then 

 precipitated by the addition of a large excess of alcohol (usually 

 twenty volumes). After standing some days or weeks the pre- 

 cipitate is drained off and dried, and is then ground up and ex- 

 tracted with water. The aqueous extract contains proteins, 

 salts, atid other things in addition to the thrombin. A solution 

 made in this way causes a prompt coagulation when added to 

 a solution of pure fibrinogen. That the thrombin thus obtained 

 is not present as such in normal blood, bat is formed after shed- 

 ding, is indicated by the fact that if the animal's blood is allowed 

 to flow directly from the artery into a large bulk of alcohol, due 

 care being taken in the process, the precipitate thus obtained when 

 subsequently dried and extracted with water yields little or no 

 thrombin. 



Another, perhaps simpler, method of obtaining a strong prep- 

 aration of thrombin is to treat fibrin with an 8 per cent, solution of 

 sodium chlorid (Buchanan-Gamgee). Fibrin obtained from a 

 slaughter-house is washed thoroughly in running water until the 

 iiemoglobin is removed, and is then minced and extracted at a low 

 temperature with the strong salt solution for several days. The 

 filtered extract is rich in thrombin, but contains also large amounts 

 of dissolved protein. Starting with such an extract the author* 

 has shown that by repeated shakings with chloroform the coagu- 

 lable proteins present in the extract may be removed completely 

 and the thrombin, in diminished quantities, be left behind in 

 apparently pure condition. 



A simpler method of obtaining relatively pure preparations of thrombin 

 in permanent form for experimental work and demonstrations may be de- 

 scribed briefly as follows: 



1. Fibrin obtained from the slaughter-house is washed thoroughly to re- 

 move all hemoglobin. The mass is squeezed to remove excess of water, is 

 then cut into fine pieces with scissors, and covered with an 8 per cent, solution 



* Howell, "American Journal of Physiology," 26, 453, 1910. 



