Toluylene red staining solution

Example 2

The H-1PV virus (stored by −20° C. or ≤−60° C. in 48% Iodixanol/Ringer solution obtained by mixing 73.62% (w/v) VISIPAQUE™ (iodixanol) 320 (GE Healthcare) with 26.30% (w/v) Ringer solution (Delta Select GmbH, Dreieich, Germany)) is stable in activity measured with plaque forming units (PFU) for more than 2 years. Plaque assays were done essentially as described by Tattersall and Bratton (J. Virol. 46 (1983), 944-55). NB-324K cells were grown in monolayer cultures in MEM medium containing 5% FBS, 100 μg/ml penicillin, 100 μg/ml streptomycin, and 2 mM L-glutamine. They were infected at 60% confluence with serial dilutions of H-1PV and incubated for 1 h at 37° C. Then the inoculum was replaced with a bacto-agar overlay (1.7% in MEM containing 5% FBS). On day four post-infection, living cells were stained for 18-24 h by addition of 0.02% toluylene red staining solution (Sigma, Germany) containing bacto-agar (Becton Dickinson, Germany). The dishes were incubated at 37° C. under 5% CO₂. Plaque-forming units were counted 5 days post-infection on a light box and their concentration expressed in PFU/ml.

The results are shown in FIG. 5. There it can be seen that a virus concentration less than 1×10⁸ pfu/ml is more instable in 48% Iodixanol/Ringer solution than a higher virus concentration of about 1×10⁹ or 1×10¹⁰ pfu/ml.

Example 2

The H-1PV virus (stored by −20° C. or ≤−60° C. in 48% Iodixanol/Ringer solution obtained by mixing 73.62% (w/v) VISIPAQUE™ (iodixanol) 320 (GE Healthcare) with 26.30% (w/v) Ringer solution (Delta Select GmbH, Dreieich, Germany)) is stable in activity measured with plaque forming units (PFU) for more than 2 years. Plaque assays were done essentially as described by Tattersall and Bratton (J. Virol. 46 (1983), 944-55). NB-324K cells were grown in monolayer cultures in MEM medium containing 5% FBS, 100 μg/ml penicillin, 100 μg/ml streptomycin, and 2 mM L-glutamine. They were infected at 60% confluence with serial dilutions of H-1PV and incubated for 1 h at 37° C. Then the inoculum was replaced with a bacto-agar overlay (1.7% in MEM containing 5% FBS). On day four post-infection, living cells were stained for 18-24 h by addition of 0.02% toluylene red staining solution (Sigma, Germany) containing bacto-agar (Becton Dickinson, Germany). The dishes were incubated at 37° C. under 5% CO₂. Plaque-forming units were counted 5 days post-infection on a light box and their concentration expressed in PFU/ml.

The results are shown in FIG. 5. There it can be seen that a virus concentration less than 1×10⁸ pfu/ml is more instable in 48% Iodixanol/Ringer solution than a higher virus concentration of about 1×10⁹ or 1×10¹⁰ pfu/ml.