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Leuco‐Adsorption on Cell Cultures Infected with Influenza Virus

Gresser and Chany (1962, unpublished observations) devised a leuco-adsorption test similar to the hemadsorption test, and observed that leukocytes added to Sendai virus-infected cell cultures were rapidly adsorbed to the monolayers (2). Recently, leuco-adsorption has been seen on cells infected with vaccinia (7), mumps, Newcastle disease, and measles viruses (3). These viruses in general acquire a viral envelope from the infected cell membrane containing viral hemagglutinin and they are released from the infected cells by budding. Since influenza virus is a typical enveloped virus with a hemagglutinin, the appearance of leuco-adsorption on infected cells is expected. This paper presents morphological observations on the leuco-adsorption occurring on cells infected with Aor B-type influenza virus and points out a difference between leuco-adsorption and hemadsorption. Four strains of influenza virus, A/Kanagawa/26/74 (H3N2), A/Kanagawa/20/ 75 (H3N2), B/Kagoshima/1/68 and B/Gunma/1/73, were used. Nearly confluent monolayers of MDCK cells grown on a coverslip were infected with stock viruses at multiplicities of infection of 0.04 to 3.4 plaque-forming units (PFU) per cell. After virus adsorption for 30 min at 34 C, the cultures received 1.0 ml of fresh maintenance medium containing fourfold the normal amount of vitamins, twice the normal amount of glucose, 0.2% bovine albumin and 10 units of trypsin per ml. After incubation at 34 C for 20 hr, the maintenance medium was removed, the cell sheets were washed twice with Eagle minimal essential medium (MEM) and 1.0 ml of fresh MEM and 0.1 ml of a chicken leukocyte suspension containing approximately 1 •~ 106 to 107 cells per ml were added. The procedure for separating leukocytes from heparinized chicken blood was described in a previous paper (7). The chickens used had no hemagglutination-inhibition antibody for either influenza A or B virus. After being incubated for 1 hr at 4 C, the cell sheets were washed with MEM to remove the unadsorbed leukocytes, then observed microscopically. We found a large number of leukocytes adhering to the infected cells (Fig. la), but few leukocytes adhered to uninfected cells (Fig. 1c). In parallel experiments, chicken erythrocytes added to the infected cells adhered in the same way (Fig. lb). The number of leukocytes adhering to one infected cell was less than 15, similar to that of erythrocytes. We found no difference in the degree of leuco-adsorption