TABLE III
Advantages and Limitations of Conventional Assays for Virus Detection
 In vitro cell culture tests |
| • ADVANTAGES |
| – Broad range of viruses that are pathogenic for humans may be detected in one assay |
| – Allows amplification of low level virus contaminant |
| • LIMITATIONS |
| – If adventitious viruses do not replicate in the indicator cells to produce CPE, hemagglutination, or hemadsorption, they will not be detected. |
| – Even if adventitious viruses replicate, the CPE may not be obvious and the RBCs used in the testing may not be able to HAg or HAd the virus. |
| – In some cases, vaccine virus may need to be neutralized and high titer serum may not be available or sufficiently effective (giving false-positive results). |
| – Test material may interfere with virus detection (reducing assay sensitivity). |
| – Most retroviruses cannot be detected. |
| – Sensitivity of the tests is not known for naturally occurring viruses. |
 In vivo assays |
| • ADVANTAGES |
| – Broad range of viruses pathogenic for humans may be detected in one assay. |
| – Allows amplification of a low level virus contaminant. |
| • LIMITATIONS |
| – If agents cannot replicate and produce disease or hemagglutinate RBCs in test system, they will not be detected. |
| – For virus seeds, neutralization of vaccine virus might be required. |
| – Components in test article may be toxic, producing non-specific results. |
| – Sensitivity of assays is generally unknown. |
 TEM |
| • ADVANTAGES |
| – Broad detection |
| – Visualization of virus in test article provides information regarding type of viral contaminant. |
| • LIMITATIONS |
| – Low sensitivity |
| – In case of novel virus or low-level contamination, it may need a trained expert to distinguish virus from cellular background material. |
| – Qualitative: it requires further evaluation as to the identity and functionality of virus particles. |
 PCR-based RT assays |
| • ADVANTAGES |
| – Highly sensitive |
| – Broad detection of all retroviruses (infectious and non-infectious) |
| – Quantitative assays available |
| • LIMITATIONS |
| – Need follow-up of positive results to determine identity and infectivity |
| – Caution in interpretation of results |
| ○ Some cellular DNA polymerases can be detected in the assay and therefore source of low-level signal needs to be determined. |
| ○ Some cell substrates may produce a low-level RT activity and therefore origin/nature of signal needs further evaluation. |
 Antibody Production Assays |
| • ADVANTAGES |
| – Broad virus detection using three animal species (15 murine viruses, 5 hamster viruses, 11 rat viruses) |
| • LIMITATIONS |
| – Sensitivity undefined |
| – (Alternative PCR or RT-PCR assay for MAP for 26 or more viruses.) |
 PCR Assays |
| • ADVANTAGES |
| – High sensitivity, rapid, and can reveal virus identity |
| – Assays can be expanded with novel viruses. |
| • LIMITATIONS |
| – Need to test for individual viruses/families |
| – Assays need to be updated based upon virus discovery. |
| – Broad-range PCR primers may need to distinguish viral from cellular origin. |
| – Need follow-up to determine if intact, infectious virus is present |
 Infectivity Assays |
| • ADVANTAGES |
| – High sensitivity, can provide amplification step for detection of low-level virus |
| • LIMITATIONS |
| – Need to use different target cells based upon virus susceptibility |
| – Selection of target cells is based upon known viruses. |
| – Amplification of low-level contaminant requires an extended culture time. |