| Abstrakt: |
The neutralization test remains the standard tool for determining titer of specific antibodies for all rhinovirus types (1). The method requires use of a small dose of virus to achieve needed sensitivity. Tests are generally performed in tube cultures of a cell system such as WI-38 in which cytopathic effect can easily be detected (2). The technique is highly sensitive and reproducible but is expensive in terms of cell cultures required and media and sera used.A simple micromodification of the procedure would clearly be of importance if it retains the sensitivity of the original technique. The present report describes the development of such a method, which can be performed with standard microtiter equipment. An added advantage is that endpoints can be read macroscopically and the plates preserved for later reference.Materials and Methods. Sera and viruses.A total of 147 sera collected during the study of respiratory infections in Tecumseh, Michigan, were tested for antibody content. In general, three specimens had been collected from each individual during the course of 1 year's surveillance (3). The sera were inactivated at 56° for 0.5 hr and were diluted 1:2 in Hanks' Balanced Salt Solution (BSS). Rhinoviruses used had originally been isolated from individuals in the Tecumseh community in WI-38 cells and had been identified as types 12 and 15 (4).Microprocedure.HeLa (Rhino) cells, originally obtained from Grand Island Biological Co., were propagated serially in our laboratory; for growth, Eagle's Minimum Essential Medium (MEM) in Hanks' salts was used, supplemented with 10% calf serum and with 100 U of penicillin and 100 μg of streptomycin per ml. Bottle cultures were trypsinized after a full cell sheet had formed, usually following 3–4 days of growth. These cells were used to make either microplates or tube cultures. The tube cultures were employed for adaptation of rhinoviruses to the HeLa (Rhino) cell system for later use in the microprocedure (5). |
