In 1915 Clough1 reported intracutaneous reactions in lobar pneumonia patients. He prepared his antigen as follows: Cultures used were made from the lungs in cases of fatal pneumonia, with no statement as to type. From twenty-four to thirty-six hour old cultures in 5 per cent. glucose broth plus calcium carbonate were decanted off the carbonate, centrifuged, washed with salt, recentrifuged, the sediment taken up in a few drops of distilled water, dried, weighed, ground for three hours with sterile sand, extracted with 10 c. c. saline for each gram of dried material, incubated for eighteen hours; centrifuged at high speed for several hours until no more sediment came down; used in this form or precipitated with absolute alcohol. One gram of the dried culture residue yielded about 0.15 gm. dried alcoholic precipitate.
He used two methods of inoculation: first, allowing a drop of 1 per cent. solution of his antigen
BIGELOW GH. INTRACUTANEOUS REACTIONS IN LOBAR PNEUMONIA. Arch Intern Med (Chic). 1922;29(2):221-235. doi:10.1001/archinte.1922.00110020080003