Fractionation of Pollen Proteins and Their Skin Reactions on Sensitive Individuals.

Dunbar 1 showed that hayfever was caused by sensitiveness to the proteins of pollen. We are attacking the problem on the following assumption: (1) That hayfever is due to sensitivity of an individual to one or more proteins of pollen. (2) That this sensitivity is due only to substances of protein nature (we are aware that evidence has been presented that sensitivity may be due to non-protein substances. (3) That the isolation of the offending protein or proteins in pure form should be the ideal therapeutic agent. Our present paper is concerned only with the separation of the proteins of giant (Ambrosia trifida) and common (Ambrosia elatior) ragweed pollen, and the results of skin reactions of the proteins obtained on sensitive individuals.

Twenty-eight gm. of pollen was exhaustively extracted in a ball-mill with the following solvents in the order indicated: Distilled water, 5% NaCl, 1% NaF, 0.25% Na₂CO₃, and 0.1% HCl. It was determined clinically that water extracted practically all of the active substances. Hence, our interest is mainly in the water soluble proteins.

To a solution of the water soluble proteins an equal volume of a saturated solution of ammonium sulphate was added. The precipitated proteins were filtered off, dissolved in water, and reprecipitated from the water solution 3 times by adding an equal volume of ammonium sulphate as indicated, finally the proteins were dissolved in water and dialyzed free from salt. The proteins which precipitated between these concentration limits with ammonium sulphate are the globulins. The filtrate from the above fraction was raised to 64% by adding saturated ammonium sulphate. The proteins which were precipitated between these limits were reprecipitated 3 times, finally dissolved in water and dialyzed the same as the previous fraction.