254 IMMUNOLOGY 



While active immunization against diphtheria in the experi- 

 mental animal began with the production of antitoxin by Behring, 

 it was not possible to apply it to man until a safe and dependable 

 antigen was produced and some method devised to determine on 

 a large scale the susceptibility of children and adults to diphtheria. 



The history of the research which led ultimately to the develop- 

 ment of a satisfactory susceptibility test in man is of considerable 

 interest and importance. 



In 1909, Homer and Sames introduced the intracutaneous method 

 of titrating toxin and antitoxin. At the present time it is uni- 

 versally employed in determining the virulence of C. diphtheriae 

 and is used frequently in the titration of toxin. According to 

 Glenny (1921) it is a convenient guide in following the trans- 

 formation of toxin to toxoid. 



M.R.D. — Romer adopted two units of toxin, the M.R.D. (min- 

 imum reacting dose) and the Lr (limit of reaction) dose, respec- 

 tively. Glenny (1931) has defined the M.R.D. as the least amount 

 of toxin which when injected intracutaneously into a guinea pig 

 will produce, within 36 hours, an area of hyperemia that is at 

 least 5 mm, in diameter. He defines the Lr dose of toxin as the 

 least amount of toxin which when added to one unit of antitoxin 

 will yield a mixture of such toxicity that 0.2 c.c. will produce a 

 minimal skin reaction when injected intracutaneously into a 

 guinea pig. Other doses of toxin such as Ly/lOO or Lr/500 refer 

 to the least amount of toxin which when mixed with 1/100 or 

 1/500 unit of antitoxin, respectively, will give a minimal skin 

 reaction in a guinea pig. Whether one employs an end point 

 represented by a faint area of hyperemia, or by an area of 

 hyperemia with resulting necrosis, is of importance since they 

 represent different amounts of toxin. Glenny and his associates 

 prefer the former wliile Romer made use of both end points. 

 Romer (1909) found that 1/500 of an M.L.D. will produce hyper- 

 emia only, while 1/250 of an M.L.D. will cause hyperemia fol- 

 lowed by necrosis within 72 hours. Glenny, Pope and Wadding- 

 ton (1925), using an improved intracutaneous technique, conclude 

 that 1 M.L.D. is equal to 1,000 M.R.D. 's. Glenny (1931) says that 

 more recent work in which a faint area of hyperemia 4 to 5 cm. 

 in diameter is emploj'ed indicates that 1 M.L.D. is equal to 2,000 

 M.R.D. 's. 



