Bovine Virus Diarrhea Virus (BVDV) is not as easily detected by FA in chronic cell culture infections as it is in acute infections. One example is observed during 9CFR 113.51-2 testing, which requires an FA test after 3-4 weeks and 1-2 passages of indicator cells. A BVDV infection that might have been a 3-4+ by direct FA in nearly every cell as an acute infection before the first passage may only show a 2+ fluorescence in one out of 20 cells at the end of the required 21-28 days. For maximum sensitivity when testing biological fluids for infection by BVDV, we recommend that FA test be performed at each cell culture passage. Furthermore, indicator cells must be grown on glass, either in chamber slides or on cover slips, and fixed in pure acetone only.