TECHNIC OF PRECIPITIN REACTIONS 331 



clear immune serum, The tubes must not be shaken. Within from one 

 to five minutes a faint, misty cloud appears at the bottom of the tubes 

 reacting positively, and this becomes a distinct precipitate within one- 

 half to one hour (Fig. 93). 



Before performing the actual test with the unknown blood-stain it 

 is advisable to test the entire reaction with a similar known blood-stain 

 in order to make sure that all ingredients are in good working order. 

 In laboratories equipped for medicolegal examinations stains upon linen, 

 as by the blood of man, dog, cat, ox, horse, etc., and their respective 

 antiserums are always kept in readiness for making the preliminary and 

 actual tests. 



Technic of the Test. The following mixtures are set up in a series of 



test-tubes. Fresh sterile pipets should be used in handling the various 



solutions. The immune serum should be added slowly, and in such a 



way that it will flow down the side of the tube and collect at the bottom. 



Tube 1: 2 c.c. of extract of unknown substance in dilution of 



1:1000+0.1 c.c. of immune serum. 

 Tube 2: 2 c.c. of the unknown extract in dilution of 1 : 5000+0.1 



c.c. of immune serum. 

 Tube 3: 2 c.c. of the unknown extract in dilution of 1 : 10,000+0.1 



c.c. of immune serum. 

 Tube 4: 2 c.c. of the unknown extract in dilution of 1 : 100+0.1 c.c. 



of normal rabbit serum (control). 



Tube 5: 2 c.c. of a 1:1000 dilution of the serum of that species of 

 animal whose blood is suspected to be present in the unknown 

 solution +0.1 c.c. of immune serum (control). 



Tube 6: 2 c.c. of the extract of unknown substance alone (control). 

 Tube 7: 0.1 c.c. of the immune serum +2 c.c. of normal salt solu- 

 tion (control). 

 Tube 8: 2 c.c. of the extract of the unstained portion of clothing+ 



0.1 c.c. of immune serum (control). 



The tubes are not shaken, are kept at room temperature, and the 

 results are read after from ten to twenty minutes. Exposure to incu- 

 bator temperature facilitates the reaction. With proper immune serums, 

 and especially in medicolegal work, a positive reaction should appear 

 within two to five minutes as a faint, misty cloud at the bottom of the 

 test-tube. Within five minutes this becomes more definite, and in from 

 ten to twenty minutes the precipitate is seen (Fig. 94). Any cloudi- 

 ness that develops later than twenty minutes after the beginning of the 

 reaction has no significance. 



