Primary rhesus monkey kidney (MK) cells have long been the cells of choice for isolation and propagation of the human paramyxoviruses (parainfluenza 1, 2, 3, 4A, 4B, and mumps). However, problems with the supply and cost of MK cells and the presence of endogenous viruses, including herpes B virus and SV-5, necessitated a search for an alternative cell line. Continuous cell cultures of human origin (L132, A-549, HuT-292, HEK, G-293, G-401, A-498, A-704, CAKI-1, RD) and simian origin (LLC-MK2, BSC-1, MA-104, Vero) were evaluated for their capacity to support the growth of the human paramyxoviruses, as followed by cytopathic effect, hemadsorption, hemagglutination, and EIA. NCI-H292 (HuT-292) human lung mucoepidermoid carcinoma cells (ATCC # CRL-1848) proved to be the most sensitive line for cultivating all serotypes and strains of the paramyxoviruses. These cells were also shown to be a suitable substitute for MK in primary isolation of paramyxoviruses from clinical specimens. RPMI-1640 with 1.5 micrograms/ml trypsin was the preferred maintenance medium; alternatively, Eagle's MEM supplemented with 1.5 micrograms/ml trypsin and 0.1% ITS was satisfactory. NCI-H292 cells are a continuous line with excellent growth characteristics, although the genetic polyploidy of the cells may limit the number of passages of usable cells.
NCI-H292 mucoepidermoid carcinoma cells from human lungs were shown in an earlier report to be a fully adequate substitute for primary rhesus monkey kidney (MK) cells for the isolation and propagation of the human paramyxoviruses. Although sensitivity for ortho-and paramyxoviruses was the principal reason for using MK cells, the cells were also sensitive to many other viruses, which constituted another important value of MK cells. That MK cells supported the initial isolation and growth of so many respiratory viruses made it a mandatory cell type for any clinical laboratory. We therefore felt it was imperative to evaluate the virus spectrum of NCI-H292 cells, which are being used as a substitute for MK cells in many laboratories. In the present report, we show that NCI-H292 cells are sensitive for vaccinia virus, herpes simplex virus, adenoviruses, BK polyomavirus, reoviruses, measles virus, respiratory syncytial virus, some strains of influenza virus type A, most enteroviruses, and rhinoviruses, in addition to the parainfluenza and mumps viruses originally reported. Furthermore, these viruses replicate in NCI-H292 cells to the same virus and antigen titers and at the same speed of replication as they do in their usually preferred cells. The NCI-H292 cells are therefore an excellent substitute for MK cells in terms of laboratory safety, ease of availability, paramyxovirus isolation, and broad virus spectrum but cannot substitute for MK cells for the isolation of influenza viruses.
Monoclonal antibodies were prepared to the F and M proteins of parainfluenza 4A and 4B and to mumpsvirus to obtain reagents that could be configured into type-specific yet broadly-reactive IFA, EIA, and TR-FIA tests. Several antibodies to parainfluenza 4A also detected subtype 4B, although to a somewhat lower signal, and thus were well suited to generic parainfluenza type 4 tests that were comparable to similar tests previously described for parainfluenza types 1, 2, and 3. Monoclonals to subtype 4B were less able to detect 4A because of high background problems in one or another test. Monoclonals to mumpsvirus F protein were completely type-specific. These antibodies were screened by IFA and EIA for broad reactivity with diverse strains of each virus and were configured into optimized EIA and TR-FIA tests. The all-monoclonal tests were then compared to polyclonal tests in terms of their ability to detect virus in clinical specimens. The all-monoclonal TR-FIA was uniformly the most sensitive, detecting virus in 80% of culture-positive parainfluenza 4A specimens, 67% of parainfluenza 4B specimens, and 90% of mumps specimens, compared to 40-67% for the monoclonal EIA tests and 33-60% for the polyclonal EIA tests. For parainfluenza 4 TR-FIA, mean P/N values were 379 for subtype 4A cell culture fluids (228 for subtype 4B cultures) and 57 for 4A clinical specimens (43 for 4B specimens). For mumpsvirus TR-FIA, mean P/N values were 27 for culture fluids and 32 for clinical specimens. The sensitivities of the TR-FIA were determined with purified virus to be 0.28 ng virus per well for parainfluenza 4A and 0.70 ng virus per well for mumpsvirus.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.