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학술저널
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대한바이러스학회 JOURNAL OF BACTERIOLOGY AND VIROLOGY 大韓바이러스學會誌 제1권 제1호
발행연도
1971.6
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5 - 16 (12page)

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The plaque techniques of Dulbecco (1952) was first applied to the titration of arboviruses in culture of chicken fibroblast cells, also Poterfield (1959) described modificatory the plaque technique for titrating the infectivity of animal viruses. Most of arboviruses multiply in mammalian cells giving a variable amount of cytopathic degeneration. Some will produce plaque quite readily a monolayer of these cells under agar and agarose but other fail to do so. At a time when efforts were being made in this laboratory to improve the sensitivity of plaque reduction in mammalian cell by arboviruses, report were published (Miles, 1963 and Suitor, 1965) coneerning plaque production with arbovirus. They showed that several arboviruses produce minute plaques on monolayer under agar and agarose because it was inhibited by sulfate polysaccharide. They further demonstrated that this sulfated polysaccharide, which is a break down product regularly present in agar or agarose, can be adsorbed to and inactivated by diethylaminoethyl (DEAE)-dextran or protamine sulfate. When these substances were added to the agar or agarose in an appropriate dose the susceptible some arbovirus produced plaques quite as large and clear cut as those of the normal strain. This paper describes experimental modification of the plaque technique and reports obtained employing different overlay systems. The cells were used Inoues a clonal line from porcine kidney stable PS (Y-15') throughout this study. This cell cultured in Inoue's or Westaway's medium in 2 oz prescription bottle until monolayer formed. Virus strains, all of the 2 group A(Chikungunya, Western equine encephalitis) and 4 group B(Dengue II and IV, West nile, Yellow fever) arboviruses tested to produce plaques. Protamine sulfate and DEAE- dextran prepared as a various in distilled water. This stock solution were sterilized by autoelave(dextran) or filtration. They were added to the melted noble agar or agarose in appropriate amounts immediately before pouring. In those cases chikungunya virus, the plaque size are nearly same in diameter with each of six overlay systems but number of plaques are slightly difference. WEE virus that plaque size shows a range from small to large. The total number of plaques show a less than two fold difference from bottle to bottle. Dengue II virus with six overlay systems are seen. The plaque size shows a range from small (0.3 - 0.4) to large (l.5 - 2.0) The total number of plaque, both large and small, show a less than two fold difference from bottle to bottle. This virus shows variation in plaque size between agar and agarose overlay. In Dengue IV virus, plaque size shows a range from small to large and number of plaques were less than three fold difference in overlay systems. In West nile and Yellow fever virus, plaque size show small and large also total plaque number were variable under six overlay systems. West nile virus was effected with DEAE dextran in agarose overlay. The addition of these chemical reagents in optimal concentrations do not inhibit plaque sizes and number, those plaques by some viruses bear polymorphic appearances.

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