Macromolecular Components of Chick Embryo Tissue Diseased With the Virus of Equine Encephalomyelitis. ∗

Chick embryo tissue diseased with virus of equine encephalomyelitis (Eastern strain) has been studied in relation to the homogeneous macromolecular substances recently obtained from normal tissue 1 and the results observed are described here.

Eleven-day chick embryos were inoculated with virus and harvested when moribund. The heads were removed and the body tissue was ground without diluent in chilled Ten Broeck grinders. Extraction, ultracentrifugal fractionation and analysis were similar to those of the previous studies. 1

When the diseased tissue mince was extracted in 20% suspension with a mixture of equal parts 0.9% sodium chloride solution and glycerin at pH 8.5 and the pellets were dissolved in 0.9% saline (the sequence employed by Wyckoff 2 for isolation of the equine encephalomyelitis virus protein 3 ) the component with s₂₀° = ca 250 × 10⁻¹³ cm sec⁻¹ dynes ⁻¹ was observed in the second or third ultracentrifugal cycle (Fig. 1 and 2). This product was infectious to the order of 10¹⁴ mouse units per gram, and the result corroborated previous findings. 3 .

On the other hand, when extraction was made with water, no evidence of this material was seen; instead, only boundaries with s₂₀° = ca 70 × 10⁻¹³ cm sec⁻¹ dynes ⁻¹ were obtained (Fig. 3) as from non-diseased tissue. 1 Infectivity to the order 1014 was likewise associated with this product.

Fractionation of 0.9% saline extracts of virus-diseased tissue formolized to 0.4% yielded pellets which, on solution in saline, gave diffuse boundaries indicative of s₂₀° = ca 70 × 10⁻¹³. Formolization and extraction with water at pH 7.0 gave a product with relatively sharp boundaries (Fig. 3). These derivatives of 0.9% saline and of water extraction were both highly antigenic, protecting guinea pigs against 1000 mouse infectious units of virus given intracerebrally.