A study has been made of the pH stability of centrifugally purified strains of influenza virus with respect to the biological properties of mouse infectivity and chicken red blood cell agglutinating activity. Observations also were made on the importance of composition of buffer, temperature of storage, and concentration of virus protein to the stability of the virus.
When tested for stability at a protein concentration of 0.1 mg. per cc. in phosphate buffer, the infectivity of PR8 virus was found to be most stable at pH 6.5–7; the swine virus, at pH 7–7.9; and the Lee strain, at a pH of 7.9 or higher. The CCA activity of the PR8 virus in phosphate buffer was most stable at pH 7, that of the swine virus at pH 7–8, and that of the Lee virus at a pH greater than 9. Furthermore, the Lee virus was much less stable in dilute solution in phosphate buffer, even under optimum conditions of pH, than either the PR8 or swine strains.
The different strains of influenza virus were found to possess certain characteristics in common. They lost infectivity and CCA activity on the acid side of optimum pH conditions much more rapidly than on the alkaline side. Under suitable conditions of buffer and pH, the infectivity decreased while the CCA activity remained unchanged. In general, the rate of loss in infectivity was greater than the rate of loss in CCA activity.
When tests of stability were carried out at a protein concentration of 0.1 mg. per cc. in a composite phosphate-glycine-NaCl buffer, the virus strains showed less marked differences and possessed much higher stabilities of CCA activity and infectivity than when stored at the same concentration in phosphate buffer alone. Under the modified conditions, all three viruses possessed maximum stabilities of CCA activity and infectivity at pH 7–8 with the exception of the PR8 virus whose infectivity appeared more stable at pH 7 than at pH 8. In detailed experiments with the Lee virus, it was found that the infectivity and CCA activity of this strain at pH 7 and at a protein concentration of 0.1 mg. per cc. were maintained best in the composite phosphate-glycine-NaCl buffer, less well in a buffer containing glycine and NaCl, and least well in phosphate buffer alone.
In tests with PR8 virus, the activity was found to be much more stable at 4° C. than at 23° C.
When stored at a concentration of 2 mg. per cc. at 4° C. in phosphate buffer at pH 7, the PR8 and Lee strains were found to be much more stable than when stored at the concentration of 0.1 mg. per cc. At the higher concentration, no significant losses in either infectivity or CCA activity were observed over a period of 2 months.