Jul 30, 1981 - The immunological relationship of canine distemper and measles ... 1980a, b) and herpesvirus strains '(Pereira et al., 1980). We have recently ...
J. gen. Virol. (1981), 5"/, 179-183. Printed in Great Britain
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Key words: measles/SSPE/monoclonaUeanine distemper
Differentiation of Measles Virus Strains and a Strain of Canine Distemper Virus by Monocional Antibodies (Accepted 30 July 1981) SUMMARY
Monoclonal antibodies raised against the Hall~ strain of measles virus were tested for their strain specificity using other measles virus isolates (Edmonston, Leningrad, Lec and two fresh isolates JT and Fasquelle). Monoclonal antibodies to L, HA and NP polypeptides, with one exception, reacted with all the measles virus strains tested; antibody from hybrid line 25 failed to react with the NP polypeptide of JT virus by immunofluorescence or radioimmunoprecipitation. Three other monoclonal antibodies, two reacting with NP and one with HA, although giving a positive immunofluorescence reaction, did not immunoprecipitate labelled antigen from JT virus-infected cells. The immunological relationship of canine distemper and measles viruses was also investigated with monoclonal antibodies. Two of the three anti-NP and three of the four anti-HA monoclonal antibodies reacted with canine distemper virus in an immunofluorescence reaction but only one (anti-NP) reacted in a radioimmunoprecipitation test. The hybridoma technique pioneered by Kfhler & Milstein (1975) for the production of monoclonal antibodies, has been applied to a variety of antigens. In virology, they have been used for numerous problems including the differentiation of rabies virus (Flamand et aL, 1980a, b) and herpesvirus strains '(Pereira et al., 1980). We have recently prepared monoclonal antibody against measles virus (Giraudon & Wild, 1981), which we wish to use as immunological probes in measles virus and measles virus associated diseases. Measles is a member of the morbillivirus subgroup of the paramyxoviruses and contains six structural polypeptides: L, associated with the transcriptase; HA, haemagglutinin; P, phosphoprotein; NP, nucleoprotein; F, fusion antigen and M, matrix. Our monoclonal antibodies were prepared against the Hall6 measles virus isolated from a subacute sclerosing encephalitis (SSPE) patient. In our present study, we have determined the specificity of our monoclonal antibodies with a number of measles virus isolates and also canine distemper virus (CDV). The latter virus was included as it has been shown to be closely related to measles virus (Orvell & Norrby, 1974; Hall et al., 1980) and together with measles virus has been implicated in multiple sclerosis (Cook et al., 1979). In the present study, we used six strains of measles virus; two vaccine strains (Edmonston and Leningrad), two SSPE-derived strains (Hall+ and Lee) and two recently isolated wild strains kindly provided by Dr Lebon, Paris (JT and Fasquelle). The Onderstepoort strain of CDV was also used. Veto cells were grown in tissue culture chamber slides (Lab-Teck) and infected at 0.1 p.f.u./cell. After a further 24 to 48 h depending on the virus strain used, cells were washed in phosphate-buffered saline and fixed with acetone. The infected cells were incubated with the various monoclonal antibodies and positively reacting cultures revealed with a fluoresceinconjugated anti-mouse y-globulin as previously described (Giraudon & Wild, 1981). With the exception of antibody from hybrid line 25 (anti-NP) all the monoclonal antibodies react with 0022-1317/81/0000-4709 $02.00 © 1981 SGM
RIP*
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Lec
Hall~
JT Fasquelle
CDV
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48
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55
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Specificity of measles (Halle') virus monoclonal antibodies
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Anti-L
* Immunofluorescence (IMF) and radioimmunoprecipitation (RIP) were performed as previously described (Giraudon & Wild, 1981); ND, not determined.
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Short communications
Fig. 1. Immunofluorescence of cells infected with Hall6 (SSPE), JT (fresh isolate) strains of measles virus and a CDV strain (Onderstepoort). (a, b) Hall~ strain; (c, d) JT strain; (e,f) CDV strain; (a, c, e) monoclonal antibody from hybrid line 25 (anti-NP); (b, d , f ) monoclonal antibody from hybrid line 22 (anti-NP).
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Short communications
the various measles virus strains (Table 1, Fig. 1). The latter antibody failed to react with the JT isolate. Two of the three anti-NP (no. 22 and 34) and three of the four anti-HA (no. 18, 48 and 55) monoclonal antibodies reacted with CDV. No cross-reaction was observed with the anti-L antibody and CDV. To further investigate the specificity of the immunological reaction, BGM cells were infected with either the Hall6, Fasquelle, JT or CDV strains (0-1 to 3 p.f.u./cell). When cytopathic effect involved approx. 30 to 50% of the cell sheet, the cells were incubated in methionine-free Eagle's medium for 1 h and then radiolabelled with 50 flCi/ml of [35S]methionine (1.010 mCi/mM, Amersham International) for 2 h. Proteins were extracted, immunoprecipitated and analysed by S D S - P A G E as previously described (Giraudon & Wild, 1981) (Table i). The lack of reactivity of antibody from hybrid line 25 with the JT strain was confirmed by the radioimmunoprecipitation (RIP) test, but additionally three other monoclonal antibodies, no. 18, 22 and 48 (all IgM) also failed to precipitate their corresponding antigens. The RIP test confirmed the immunoftuorescence results of the non-reactivity of antibody from lines 15 and 25 with CDV, but showed additionally the lack of reactivity of antibody from lines 22 and 55. The results show that immunofluorescence is the less discriminatory of the two techniques employed. The reason for these differences is not understood, but it may be related to the more restrictive conditions imposed in the RIP test as several detergents are included to reduce non-specific binding. Although the monoclonal antibodies to Hall6 HA produced by lines no. 18, 48 and 55 reacted positively in immunofluorescence with the corresponding antigen of CDV (Onderstepoort strain) they did not neutralize this virus. This suggests that the common epitope(s) recognized by these antibodies are presented in a different fashion on these two viruses. Our previous observation that polyclonal sera against the two strains do not cross-neutralize supports this hypothesis (Wild & Huppert, 1980). The present study and also that of Trudgett et al. (1981), have shown that naturally occurring variants of measles virus exist. At present the number of hybrid clones available is limited and insufficient to fully differentiate virus from different origins. We are at present enlarging the library of hybrid clones in an attempt to include a wider spectrum of antibodies. We wish to thank Dr J. Huppert for his fruitful discussion, Mme M. F. Jacquier for photography and Mlle A. Mary for typing the manuscript. This study was supported by grant A.T.P.I.N.S.E.R.M. no. 71 78 103. Unite de Virologic Fondamentale et Appliquee I.N.S.E.R.M. ( U.51) Groupe de Recherche C.N.R.S. 33 1, place Professeur Joseph Renaut 69371 Lyon Cedex 2, France
P. GIRAUDON
T. F. WILD*
REFERENCES
COOK, S. D., DOWLING, P. C. & RUSSELL, W. C. (1979). Neutralizing antibodies to canine distemper and measles virus in multiple sclerosis. Journal of Neurological Science 4 l, 61-70. ELAMAND, A., WIKTOR,T. J. & KOPROWSK1,H. (1980 a). Use of hybridoma monoclonal antibodies in the detection of antigenic differences between rabies and rabies-related virus proteins. I. The nucleocapsid protein. Journal of General Virology 48, 97-104. ELAMAND, A., WIKTOR, T. J. & KOPROWSKI, n. (1980 b). Use of hybridoma monoclonal antibodies in the detection of antigenic differences between rabies and rabies-related virus proteins. II. The glycoprotein. Journal of General Virology 48, 105-109. GIRAUDON, P. & WILD, T. F. (1981). Monoclonal antibodies against measles virus. Journal of General Virology 54, 325-332.
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HALL, W., LAMB, R. A. & CHOPPIN, P. W. (1980). The polypeptides of canine distemper virus: synthesis in infected cells and relatedness to the polypeptides of other morbilliviruses. Virology 100, 433-449. KOHLER, G. & MILSTEIN, C. (1975). Continuous cultures of fused cells secreting of predefined specificity. Nature, London 256, 495-497. ORVELL, C. & NORRBY, E, (1974). Further studies on the immunologic relationships among measles, distemper, and rinderpest viruses. Journal of Immunology 113, 1850-1858. PEREIRA, L., KLASSEN, T. & BARINGER, J. R. (1980). Type-common and type-specific monoclonal antibody to herpes simplex virus type I. Infection and Immunity 29, 724-732. TRUDGETT, A., GOULD, E. A., ARMSTRONG, M., MINGIOLI, E. S. & MAcFARL1N,D. E. (1981). Antigenic differences in the hemagglutinin of measles and related viruses. Virology 109, 180-182. WILD, T. F. & HUPPERT, J. (1980). Specificity of measles and canine distemper virus antibodies. Annales de l'Institut Pasteur 131E, 73-84.
(Received 5 May 1981)
