242 MEDICAL BACTERIOLOGY 



The third method is preferable because it is least dangerous to the patient 

 and least painful. It does not require as large a needle as the second method 

 and precludes soiling the clothes with blood and protects the withdrawn blood 

 from contamination. 



The first method is that of necessity and should never be resorted to when it 

 is possible to obtain the blood by vein puncture. 



However the blood is obtained the procedure must be made under aseptic 

 conditions and guarded with the usual surgical precautions. 



The blood is collected in a sterile test-tube at least ^ inch in diameter. If 

 this tube is slanted as soon as the blood enters it, and allowed to remain at rest 

 until the blood coagulates, then placed upright, clear serum will separate in 

 several hours and can then be poured into another tube, inactivated and tested. 

 If the tube must be kept upright from the time the blood enters it the serum is 

 apt to be cloudy and should the interior of the tube be rough, the diameter less 

 than J^ inch, serum may not separate from the clot until it is stirred with a glass 

 rod, then it is always cloudy. When the serum is cloudy, has suspended matter 

 in it, it must be centrifugalized until perfectly clear and free of suspended matter 

 before testing. As a result of forceful expulsion of blood from a syringe into 

 a tube, agitation during transportation or stirring with a glass rod, blood cells 

 are ruptured and their hemoglobin tints the serum red, this discoloration of the 

 serum cannot be removed, nor is there any need to; it does not affect the test. 

 Serum that is sterile may be kept for weeks or months and still give the same 

 reaction as when recently removed, but, for reasons that will be later explained, 

 it is best to make the test within 48 hours after removal of the serum from the 

 patient. 



Occasionally, it is desirable to make a test as soon as possible, then the blood 

 is collected in a centrifuge tube, gently stirred with a glass rod and centrifugal- 

 ized at high speed for 5 minutes ; in that time clear serum will collect at the top 

 of the tube and can be at once removed, inactivated and tested. 



The experience of the vast majority of serologists shows it is best to inacti- 

 vate the patient's serum. This deprives it of complement. The serum should 

 be inactivated immediately before testing and control sera should be inactivated 

 every time they are used, regardless of how often that may be. 



Sera are inactivated by placing the tubes containing them in a water bath 

 at 55 or 56C. for J^ hour. It will not matter if they are heated a few minutes 

 more, but it is imperative that the temperature is maintained at 55C. or 56C. 



Just as different rabbits injected with red blood cells have different amounts 

 of amboceptor in their sera, different individuals infected with syphilis have 

 different amounts of amboceptor in their sera. Wassermann, Neisser, Buchner, 

 Morgenroth, Ehrlich and others discovered by experimentation that from 0.20 

 to 0.02 cc. of patient's serum might be used to make a complement fixation test 

 for syphilis. 



For reasons that will be discussed later, o.i cc. of patient's serum seems the 

 ideal quantity to use, and in the Wassermann test as described in this book o.i 

 cc. of the patient's serum is the amount always employed. 



