BLOOD AND LYMPH 



267 



ester of hematin" is formed. Filter and dissolve on the filter paper by sodium 

 carbonate. Save this alkaline solution of hematin and make a spectroscopic ex- 

 amination later after becoming familiar with the use of the spectroscope. How 

 does the spectrum of oxyhemoglobin differ from that of the derived alkali hematin? 

 19. Preparation of Thrombin (Howell). 1 Prepare fibrin from pig's blood 

 according to directions given on page 268. Wash the fibrin thoroughly in water 

 to remove hemoglobin. Squeeze out the water, mince the fibrin and cover with 

 an 8 per cent sodium chloride solution and allow to stand in the cold for 48 hours. 

 Filter. Precipitate the thrombin (and other proteins) from the nitrate by adding 

 an equal volume of acetone. Filter the mixture rapidly through a number of 

 small (25-50 c.c.) filters. Spread out filter papers and precipitate and dry rapidly 

 in]a current of cold air. Cut the dried papers into small pieces and treat with a 

 volume of water equivalent to 66 per cent of the 8 per cent NaCl previously used. 

 Allow to stand one -half hour and filter. Shake the filtrate with chloroform 

 (10-15 c - c - P er I0 c - c - filtrate) until on settling no opalescence is developed by 



FIG. 80. SODIUM CHLORIDE. 



heating a portion of the supernatant fluid. Decant the liquid and evaporate 

 on watch glasses (2 c.c. to a watch glass) in a current of air. Thrombin so pre- 

 pared may be kept indefinitely in a desiccator. 



20. Variation in Size of Erythrocytes. Prepare two small funnels with filter 

 papers such as are used in quantitative analysis. Moisten each paper with physio- 

 logical (isotonic) salt solution. Into one funnel introduce a small amount of 

 defibrinated ox blood and into the other funnel allow blood to drop directly from a 

 decapitated frog. Note that the nitrate from the ox blood is colored, whereas that 

 from the frog blood is colorless. What deduction do you make regarding the 

 relative size of the erythrocytes in ox and frog blood? Does either nitrate clot? 

 Why? 



II. Blood Serum 2 



i. Coagulation Temperature. Place 5 c.c. of undiluted serum in a test-tube 

 and determine its temperature of coagulation according to the method described 



1 Howell: Am. Jour. PhysioL, 32, 264, 1913. 



2 For directions as to preparation of serum, see "Reagents and Solutions." (Page 593.) 



