312 AGGLUTININS 



Tube 4: Control: 4 drops of recipient's serum + 1 drop of re- 

 cipient's red-cell emulsion. Should show no agglutination or 

 hemolysis. 



Tube 5: Control: 1 drop of donor's red-cell emulsion -J- 4 drops 

 of normal salt solution. This serves as a control on the toxicity 

 of the corpuscles and isotonicity of the salt solution. 

 Tube 6: Control: 1 drop of recipient's red-cell emulsion + 4 



drops of saline solution. 



One cubic centimeter of salt solution is added to each tube and the 

 tubes are gently shaken and placed in the incubator for two hours. 

 They are to be inspected every half hour. Agglutination is recognized 

 macroscopically by the clumping of the red blood-cells into small masses 

 that later sink to the bottom of the tube as a small clot. Hemolysis 

 is likewise easily detected, as corpuscles tend to become precipitated 

 within two hours. If doubt exists, the finer grades of hemolysis may be 

 detected after the tubes have been allowed to stand over night in an ice- 

 chest, or at once by thorough centrifugalization. 



Pipet Method. In cases where, for any reason, the quantities of 

 blood previously named cannot be secured, the whole operation may be 

 conducted with smaller amounts, using Wright's capillary pipets 

 (Epstein and Ottenburg). This method is as follows: 



Blood is secured from both recipient and donor by pricking the finger 

 of each and allowing five or six drops of blood to flow into 5 c.c. of sodium 

 citrate solution, and collecting a good-sized Wright capsule full for se- 

 curing the serum. The corpuscles are washed twice and enough normal 

 salt solution added to make up approximately a 10 per cent, suspension. 

 The capsules are centrifuged if necessary, filed, opened, and the serum 

 pipeted off. 



The mixtures are prepared in Wright's capillary pipets (see Fig. 48) 

 fitted with rubber nipples. The unit volume is marked off by a blue 

 pencil on the stem about an inch from the tip. Four volumes of serum, 

 one volume of cell emulsion, and four or five volumes of salt solution 

 are drawn up into the pipet and then mixed gently, running them out on 

 a watch-glass. The entire mixture is then drawn into the barrel of the 

 pipet and the tip sealed. The pipets are carefully labeled, incubated for 

 two hours, and examined for agglutination and hemolysis. 



The results are somewhat more difficult to read, but the method is 

 of value when many donors are to be examined or when the supplies of 

 serum and corpuscles are limited. 



Microscopic Hemagglutination Method of Brem. 1 The following 

 1 Jour. Amer. Med. Assoc., 1916, 67, 191. 



