Use of Formalin-Treated Red Cells for the Study of Influenza A Virus Hemagglutinating Activity

Abstract

Human erythrocytes, treated with formalin, the writer has found, retain their ability to react with influenza A virus in a manner quite similar to that of untreated human erythrocytes. For certain types of experiments the use of such formalized red cells possesses distinct advantages over the use of normal red cells.

To prepare formalized red cells, equal volumes of a 50% (by volume) suspension of washed red cells in saline and formalin containing sodium chloride in a concentration of 0.85%, were mixed and allowed to stand for 2 to 3 days. If the formalin concentration of the mixture was between 10 and 50%, the cells became dark brown gradually over a period of 24 hours or so. The cells also became cohesive and very resistant to hemolytic agents. The stability of the cells thus treated was marked. Samples of formalized cells stored at 4°C have retained their activity undiminished for influenza A virus for almost two years now. Furthermore, the cells have retained the microscopic appearance and shape of fresh, normal, red cells.

Concentrations of formalin of less than 10% (3.6% formaldehyde) in the mixture failed to produce browning of the red cells over 2-3 days at 4°C usually, but did render the cells cohesive. Such cells hemolyzed more readily in hypotonic solutions than did the cells prepared with higher concentrations of formalin.

Before using the formalized red cells in reactions with influenza virus the uncombined formaldehyde was removed by repeated washing with saline or water. The cells were allowed to stand in contact with each wash fluid for a period of at least several hours to permit the establishment of an equilibrium between the free formaldehyde inside and outside the cells.