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Immunofluorescent antibody response to lactic dehydrogenase virus in different strains of mice
J. Comp.
Path.
1992 Vol.
107, 179-183
Immunofluorescent Dehydrogenase T. Hayashi’, Laboratory
Antibody Response to Lactic Virus in Different Strains of Mice
I. Mori,
Y. Noguchi,
T. Itoh*
and
M. Saitoh*
of Veterinary Pathology, University of Yamaguchi, Yamaguchi and *Central Institute for Experimental Animals, Kawasaki, Japan
Summary
The development of antibody against lactic dehydrogenasevirus in five strains of mice (NZB x NZWF,, BALB/c, C.B-17, ICR and C.B-17 scid or SCID mice) was examined by indirect immunofluorescence (IIF) of infected liver sections. IIF antibody appeared 1 to 3 weeks and rose progressively 2 to 4 weeksafter infection in four strains of mice (NZB x NZWF,,BALB/c, C.B-I 7 and ICR mice). SCID mice did not develop antibody. These results suggest that IIF may be applicable for detecting LDV infection in many other ordinary strains of mice.
Introduction Lactic dehydrogenase virus (LDV) is a member of the unclassified Togaviridae with the mouse as host (Rowson and Mahy, 1985). Infected mice have an inapparent infection and develop persistent elevation of enzyme activity without any clinical signs (Notkins, 1965). The virus contaminates several different types of mouse transplantable tumour (Notkins, 1965). Infected mice exhibit altered humoral and cell-mediated immunity (Notkins, 1965; Rowson and Mahy, 1985). Because contamination with LDV might affect the results and interpretation of experiments through viral action on the host immune system (Notkins, 1965), it is important to be able to detect serum antibody to LDV. However, there are limited reports concerning the development of antibody after LDV infection in mice (Cafruny and Plagemann, 1982; Notkins, Mahar, Scheele and Goffman, 1966; Porter, Porter and Deerhake, 1969). In this brief report, the antibody response to LDV in several strains of mice after infection is described. Materials
and Methods
Animals
Five-week-old male BALB/c, NZB x NZWF, and ICR mice were obtained from SLC Co. (Shizuoka, Japan). C.B-17 scid (designated as SCID) and C.B-17 mice were bred under specific pathogen-free conditions at the Central Institute for Experimental Address for correspondence: University of Yamaguchi, 002 l-9975/92/060179
Dr T. Hayashi, Laboratory 1-1677 Yoshida, Yamaguchi
+ 05 $0&00/O
of Veterinary 753, Japan.
Pathology,
0
Faculty
1992 Academic
of Agriculture.
Press Limited
180
T. Hayashi
et al.
Animals (Kawasaki, Japan) as described previously (Hayashi, Mori, Ozaki, Saito, Itoh and Yamamoto, 1992). Mice were housed at Yamaguchi University in microisolator cages and given sterile commercial food (Oriental MF, Oriental Yeast Co., Tokyo, Japan) and water freely. Each group consisted of five to ten mice. Plasma Plasma was collected by retro-orbital bleeding with heparinized micropipettes erumo Japan) before and 1,3,5,7,14,21 and 28 days, and 2,3 and 4 months after !Zection. Virus A stock preparation of LDV kindly supplied by Dr A. L. Notkins (NIDR, NIH, USA) was used (Hayashi, Saiata, Kingman and Notkins, 1988). Mice were infected with virus at the age of 6 weeks by intraperitoneal (i.p.) injection of 104’5median infectious doses (ID,,). Antibody Titration Antibody titre was determined by a simple modification of the method described previously (Porter et al., 1969). In brief, 24 h after infection, liver tissues from ICR mice were obtained and cryostat sections used for antibody titration. Four-fold diluted samples of plasma were incubated with liver sections for 30 min at 37°C. After washing, tissues were incubated with fluorescein isothiocyanate (FITC)-labelled rabbit anti-mouse immunoglobulins (G, M and A; Cappel, PA, USA) for 30 min at 37°C. The indirect immunofluorescence (IIF) titre was expressed as the reciprocal of the highest dilution of test sample showing positive fluorescence (Fig. 1). The specificity of the reaction was confirmed by the method described previously (Hayashi et al., 1992). Statistics The titre was transferred to log,, and standard errors (S.E.) of the means were determined. Differences between means with P values of less than 0*05 (P < 0.05) were considered to be significant. Results
and Discussion
Detection of Lactic Dehydrogenase Virus (LDV) Mice after Infection
Antibody in Di$erent Strains of
As shown in Fig. 2, antibody was produced in four strains of mice (NZB x NZWF,, BALB/c, C.B-17 and ICR mice) after infection, but there was no antibody
production
in SCID
mice. Antibody
first appeared
1 to 3 weeks
and then rose dramatically 2 to 4 weeks after infection. The titre of nearly 2-3 log,, at 1 month increased slightly up to 4 months (to a titre of 3 log,,) in three strains (BALB/c, C.B-17 and ICR mice). In NZB X NZWF, mice the antibody titre of 3 log,, at 1 month increased further up to 4 months (to a titre of 4.2 log,,). The titre in NZB X NZWF, mice was greater than that in the three other strains
of mice
(PC 0.01, comparison
at 2 weeks
and 1 to 4 months
after
infection). Detection
of LDV
infection
in mice is important
for researchers
using mice
Antibody
Fig.
1.
181
in LDV mice
.~~ Virus antigen-positive cells in the cytoplasm of liver tissue obtained 24 h after lactic dehydrogenase virus (LDV) infection ( 10’51D,). Antibody titre was determined by indirect immunofluorescence (IIF). The antigen-positive cells were scattered throughout the liver. Staining was limited to the cytoplasm of typical Kupffer cells and no fluorescence was observed in hepatocytes or endothelial cells. A sample from NZB x NZWF’, mouse at 3 months after infection showing positive fluorescence. x 600.
A
/
4.0 NZBxNZW
Months Fig. 2.
The development of antibody represents the mean antibody
1
F,
after
infection
to LDV in five strains of mice infected with lO”sID,. Each point titre from 5 to 10 micef SE. Antibody titre was assayed by IIF.
182
T. Hayashi
et al.
because those infected have persistent infection without obvious clinical signs other than elevation of several kinds of enzyme activity (No&ins, 1965; Rowson and Mahy, 1985). Methods for detecting infection are as follows. ( 1) Isolation of virus from infected tissues and/or blood. This is not usually used, since there are no available cell lines (Rowson and Mahy, 1985). Primary macrophages or primary mouse embryo cells are of limited use, since only a few per cent of cells are infected (Oldstone, Yamazaki, Niwa and Notkins, 1973) and there are no cytopathic effects (Brinton-Darnell, Collins and Plagemann, 1975). (2) Detection of neutralizing antibody is also limited, since the titre is very low and the antibody appears 10 weeks after infection (Notkins et al., 1966). (3) Detection of enzyme activity is widely employed but the method lacks specificity. Therefore, the present as well as previous reports (Porter et al., 1969) indicate a simple, easy and rapid method for detecting LDV infection in mice. The IIF method described here is also useful compared with that using infected spleen as target tissue (Porter et al., 1969), since spleen contains many B cells which may react with a second antibody (FITC-labelled anti-mouse immunoglobulins). SCID mice did not develop antibody against LDV though they were persistently infected by the virus (data not shown). SCID mice are used for allogeneic and/or xenogeneic tumour implant experiments, since they lack T and B cells (Bosma, Custer and Bosma, 1983). Therefore SCID mice may acquire virus infection, since many mouse transplantable tumours contain LDV (Notkins, 1965; Stevenson, Rees and Meltzer, 1980; Heremans, Billiau, Coutelier and DeSomer, 1987). In SCID mice, examination of LDH activity in the blood is the best way to detect infection. In the present work, mice were infected with a relatively high dose of infective virus ( 104.51D5,,). A low infective dose of virus may be anticipated as a realistic contamination. It has been reported that mice could be infected by a low dose of virus ( 10”51D50) (Notkins, 1965). It seems likely that a low infective dose of virus contamination in transplantable tumours may cause infection in mice. Further study is needed to clarify this point. Initially, antibody development was very slow but a marked increase of titre was seen 2 to 4 weeks after infection in NZB x NZWF,, BALB/c, C.B-17 and ICR mice. On the other hand, a more than five-fold increase in LDH activity was detected 2 days after LDV infection ( 104’51D,,) in BALB/c, C.B-17, SCID (Hayashi et al., 1992), NZB x NZWF, and ICR mice (data not shown). In addition, it has been reported that the plasma LDH activity of mice that had received a low dose of virus ( 10”51D,,) was equal to that of the mice that had received a high dose ( 107’51D5,) (Notkins, 1965). Thus in the early stage of infection in mice, detection of LDH activity may be applicable for LDV infection. As shown in Fig. 2, the antibody titre was higher in NZB x NZWF, mice than in the other three strains of mice. NZB x NZWF, mice are known to have for induction of the a large number of Ia+ macrophages, which are important humoral immune response (Isakov, Feldmann and Segal, 1982)) compared with other strains of mice (Kelly and Roths, 1982). Decreased suppressor T cell function (Gerber, Hardin, Chused and Steinberg, 1974) and B cell activation in NZB x NZWF, mice are also reported by Prud’homme, Balderas, Dixon and
Antibody
183
in LDV mice
Theofilopoulos (1983). These features might have contributed to the high antibody response seen in this strain of mice. Further studies including the comparative degrees of infection in the different strains of mice are needed to clarify this point. References
Bosma, G. C., Custer, R. R. and Bosma, M. J. (1983). A severe combined immunodeficiency mutation in mice. Nature (London), 301, 527-530. Brinton-Darnell, M., Collins, J. D. and Plagemann, P. G. W. (1975). Lactate dehydrogenase-elevating virus replication, maturation and RNA synthesis in primary mouse macrophage cultures. Virology, 65, 187-195. Cafruny, W. A. and Plagemann, P. G. W. (1982). Immune response to lactate dehydrogenase-elevating virus: isolation of infectious immunoglobulin G complexes and quantitation of specific antiviral immunoglobulin G responsein wild-type and nude mice. Infection and Immunity, 37, 1001-1006. Gerber, N. L., Hardin, J. A., Chused, T. M. and Steinberg, A. D. (1974). Losswith age in NZB/W mice of thymic suppressorcells in the graft-vs-host reaction. Journal of Immunology, 113, 1618-1625. Hayashi, T., Mori, I., Ozaki, M., Saito, M., Itoh, T. and Yamamoto, H. (1992). Enhanced lactic dehydrogenase-5 in severe combined immunodeficiency mice: effect of lactic dehydrogenase virus on enzyme clearance. International Journal of Experimental
Pathology, 73, 173-18 1.
Hayashi, T., Salata, K., Kingman, A. and Notkins, A. L. (1988). Regulation of enzyme levels in the blood: influence of environmental and genetic factors on enzyme clearance. American 3ournal of Pathology, 132, 503-5 11. Heremans, H., Billiau, A., Coutelier, J. P.and DeSomer, P. (1987). The inhibition of endotoxin-induced local inflammation by LDH virus or LDH virus-infected tumors is mediated by interferon. Proceedings of the Societyfor Experimental Biology and Medicine, 185, 6-15. Isakov, N., Feldmann, M. and Segal, S. (1982). Acute infection of mice with lactic dehydrogenase virus (LDV) impairs the antigen-presenting capacity of their macrophages. Cellular Immunology, 66, 3 17-332. Kelly, V. E. and Roths, J. B. (1982). Increase in macrophage Ia expression in autoimmune mice: role of the lpr gene. Journal of Immunology, 129, 923-925. Notkins, A. L. (1965). Lactic dehydrogenasevirus. Bacteriological Reviews, 29, 143-160. Notkins, A. L., Mahar, S., Scheele, C. and Goffman, J. (1966). Infectious virus-antibody complex in the blood of chronically infected mice. Journal of Experimental Medicine, 124, 8 l-97. Oldstone, M. B. A., Yamazaki, S., Niwa, A. and Notkins, A. L. (1973). In vitro detection of cells infected with lactic dehydrogenase virus (LDV) by fluorescine-labeled antibody to LDV. Intervirology, 2, 261-265. Porter, D. D., Porter, H. G. and Deerhake, B. B. (1969). Immunofluorescence assayfor antigen and antibody in lactic dehydrogenase virus infection. Journal oj Immunology,
102,431-436.
Prud’homme, G. J., Balderas, R. S., Dixon, F. J. and Theofilopoulos, A. N. (1983). B cell dependence on and responseto accessory signals in murine lupus strains. Journal of Experimental Medicine, 157, 18181827. Rowson, K. E. K. and Mahy, B. W. J. (1985). Lactate dehydrogenase-elevating virus. Journal of General Virology, 66, 2297-2312. Stevenson, M. M., Rees,J. C. and Meltzer, M. S. (1980). Macrophage function in tumor-bearing mice: evidence for lactic dehydrogenase-elevating virus associated changes. Journal of Immunology, 124, 2892-2894.
1
Received, March 9th, 1992 Accepted, May 2Oth, 1992
Path.
1992 Vol.
107, 179-183
Immunofluorescent Dehydrogenase T. Hayashi’, Laboratory
Antibody Response to Lactic Virus in Different Strains of Mice
I. Mori,
Y. Noguchi,
T. Itoh*
and
M. Saitoh*
of Veterinary Pathology, University of Yamaguchi, Yamaguchi and *Central Institute for Experimental Animals, Kawasaki, Japan
Summary
The development of antibody against lactic dehydrogenasevirus in five strains of mice (NZB x NZWF,, BALB/c, C.B-17, ICR and C.B-17 scid or SCID mice) was examined by indirect immunofluorescence (IIF) of infected liver sections. IIF antibody appeared 1 to 3 weeks and rose progressively 2 to 4 weeksafter infection in four strains of mice (NZB x NZWF,,BALB/c, C.B-I 7 and ICR mice). SCID mice did not develop antibody. These results suggest that IIF may be applicable for detecting LDV infection in many other ordinary strains of mice.
Introduction Lactic dehydrogenase virus (LDV) is a member of the unclassified Togaviridae with the mouse as host (Rowson and Mahy, 1985). Infected mice have an inapparent infection and develop persistent elevation of enzyme activity without any clinical signs (Notkins, 1965). The virus contaminates several different types of mouse transplantable tumour (Notkins, 1965). Infected mice exhibit altered humoral and cell-mediated immunity (Notkins, 1965; Rowson and Mahy, 1985). Because contamination with LDV might affect the results and interpretation of experiments through viral action on the host immune system (Notkins, 1965), it is important to be able to detect serum antibody to LDV. However, there are limited reports concerning the development of antibody after LDV infection in mice (Cafruny and Plagemann, 1982; Notkins, Mahar, Scheele and Goffman, 1966; Porter, Porter and Deerhake, 1969). In this brief report, the antibody response to LDV in several strains of mice after infection is described. Materials
and Methods
Animals
Five-week-old male BALB/c, NZB x NZWF, and ICR mice were obtained from SLC Co. (Shizuoka, Japan). C.B-17 scid (designated as SCID) and C.B-17 mice were bred under specific pathogen-free conditions at the Central Institute for Experimental Address for correspondence: University of Yamaguchi, 002 l-9975/92/060179
Dr T. Hayashi, Laboratory 1-1677 Yoshida, Yamaguchi
+ 05 $0&00/O
of Veterinary 753, Japan.
Pathology,
0
Faculty
1992 Academic
of Agriculture.
Press Limited
180
T. Hayashi
et al.
Animals (Kawasaki, Japan) as described previously (Hayashi, Mori, Ozaki, Saito, Itoh and Yamamoto, 1992). Mice were housed at Yamaguchi University in microisolator cages and given sterile commercial food (Oriental MF, Oriental Yeast Co., Tokyo, Japan) and water freely. Each group consisted of five to ten mice. Plasma Plasma was collected by retro-orbital bleeding with heparinized micropipettes erumo Japan) before and 1,3,5,7,14,21 and 28 days, and 2,3 and 4 months after !Zection. Virus A stock preparation of LDV kindly supplied by Dr A. L. Notkins (NIDR, NIH, USA) was used (Hayashi, Saiata, Kingman and Notkins, 1988). Mice were infected with virus at the age of 6 weeks by intraperitoneal (i.p.) injection of 104’5median infectious doses (ID,,). Antibody Titration Antibody titre was determined by a simple modification of the method described previously (Porter et al., 1969). In brief, 24 h after infection, liver tissues from ICR mice were obtained and cryostat sections used for antibody titration. Four-fold diluted samples of plasma were incubated with liver sections for 30 min at 37°C. After washing, tissues were incubated with fluorescein isothiocyanate (FITC)-labelled rabbit anti-mouse immunoglobulins (G, M and A; Cappel, PA, USA) for 30 min at 37°C. The indirect immunofluorescence (IIF) titre was expressed as the reciprocal of the highest dilution of test sample showing positive fluorescence (Fig. 1). The specificity of the reaction was confirmed by the method described previously (Hayashi et al., 1992). Statistics The titre was transferred to log,, and standard errors (S.E.) of the means were determined. Differences between means with P values of less than 0*05 (P < 0.05) were considered to be significant. Results
and Discussion
Detection of Lactic Dehydrogenase Virus (LDV) Mice after Infection
Antibody in Di$erent Strains of
As shown in Fig. 2, antibody was produced in four strains of mice (NZB x NZWF,, BALB/c, C.B-17 and ICR mice) after infection, but there was no antibody
production
in SCID
mice. Antibody
first appeared
1 to 3 weeks
and then rose dramatically 2 to 4 weeks after infection. The titre of nearly 2-3 log,, at 1 month increased slightly up to 4 months (to a titre of 3 log,,) in three strains (BALB/c, C.B-17 and ICR mice). In NZB X NZWF, mice the antibody titre of 3 log,, at 1 month increased further up to 4 months (to a titre of 4.2 log,,). The titre in NZB X NZWF, mice was greater than that in the three other strains
of mice
(PC 0.01, comparison
at 2 weeks
and 1 to 4 months
after
infection). Detection
of LDV
infection
in mice is important
for researchers
using mice
Antibody
Fig.
1.
181
in LDV mice
.~~ Virus antigen-positive cells in the cytoplasm of liver tissue obtained 24 h after lactic dehydrogenase virus (LDV) infection ( 10’51D,). Antibody titre was determined by indirect immunofluorescence (IIF). The antigen-positive cells were scattered throughout the liver. Staining was limited to the cytoplasm of typical Kupffer cells and no fluorescence was observed in hepatocytes or endothelial cells. A sample from NZB x NZWF’, mouse at 3 months after infection showing positive fluorescence. x 600.
A
/
4.0 NZBxNZW
Months Fig. 2.
The development of antibody represents the mean antibody
1
F,
after
infection
to LDV in five strains of mice infected with lO”sID,. Each point titre from 5 to 10 micef SE. Antibody titre was assayed by IIF.
182
T. Hayashi
et al.
because those infected have persistent infection without obvious clinical signs other than elevation of several kinds of enzyme activity (No&ins, 1965; Rowson and Mahy, 1985). Methods for detecting infection are as follows. ( 1) Isolation of virus from infected tissues and/or blood. This is not usually used, since there are no available cell lines (Rowson and Mahy, 1985). Primary macrophages or primary mouse embryo cells are of limited use, since only a few per cent of cells are infected (Oldstone, Yamazaki, Niwa and Notkins, 1973) and there are no cytopathic effects (Brinton-Darnell, Collins and Plagemann, 1975). (2) Detection of neutralizing antibody is also limited, since the titre is very low and the antibody appears 10 weeks after infection (Notkins et al., 1966). (3) Detection of enzyme activity is widely employed but the method lacks specificity. Therefore, the present as well as previous reports (Porter et al., 1969) indicate a simple, easy and rapid method for detecting LDV infection in mice. The IIF method described here is also useful compared with that using infected spleen as target tissue (Porter et al., 1969), since spleen contains many B cells which may react with a second antibody (FITC-labelled anti-mouse immunoglobulins). SCID mice did not develop antibody against LDV though they were persistently infected by the virus (data not shown). SCID mice are used for allogeneic and/or xenogeneic tumour implant experiments, since they lack T and B cells (Bosma, Custer and Bosma, 1983). Therefore SCID mice may acquire virus infection, since many mouse transplantable tumours contain LDV (Notkins, 1965; Stevenson, Rees and Meltzer, 1980; Heremans, Billiau, Coutelier and DeSomer, 1987). In SCID mice, examination of LDH activity in the blood is the best way to detect infection. In the present work, mice were infected with a relatively high dose of infective virus ( 104.51D5,,). A low infective dose of virus may be anticipated as a realistic contamination. It has been reported that mice could be infected by a low dose of virus ( 10”51D50) (Notkins, 1965). It seems likely that a low infective dose of virus contamination in transplantable tumours may cause infection in mice. Further study is needed to clarify this point. Initially, antibody development was very slow but a marked increase of titre was seen 2 to 4 weeks after infection in NZB x NZWF,, BALB/c, C.B-17 and ICR mice. On the other hand, a more than five-fold increase in LDH activity was detected 2 days after LDV infection ( 104’51D,,) in BALB/c, C.B-17, SCID (Hayashi et al., 1992), NZB x NZWF, and ICR mice (data not shown). In addition, it has been reported that the plasma LDH activity of mice that had received a low dose of virus ( 10”51D,,) was equal to that of the mice that had received a high dose ( 107’51D5,) (Notkins, 1965). Thus in the early stage of infection in mice, detection of LDH activity may be applicable for LDV infection. As shown in Fig. 2, the antibody titre was higher in NZB x NZWF, mice than in the other three strains of mice. NZB x NZWF, mice are known to have for induction of the a large number of Ia+ macrophages, which are important humoral immune response (Isakov, Feldmann and Segal, 1982)) compared with other strains of mice (Kelly and Roths, 1982). Decreased suppressor T cell function (Gerber, Hardin, Chused and Steinberg, 1974) and B cell activation in NZB x NZWF, mice are also reported by Prud’homme, Balderas, Dixon and
Antibody
183
in LDV mice
Theofilopoulos (1983). These features might have contributed to the high antibody response seen in this strain of mice. Further studies including the comparative degrees of infection in the different strains of mice are needed to clarify this point. References
Bosma, G. C., Custer, R. R. and Bosma, M. J. (1983). A severe combined immunodeficiency mutation in mice. Nature (London), 301, 527-530. Brinton-Darnell, M., Collins, J. D. and Plagemann, P. G. W. (1975). Lactate dehydrogenase-elevating virus replication, maturation and RNA synthesis in primary mouse macrophage cultures. Virology, 65, 187-195. Cafruny, W. A. and Plagemann, P. G. W. (1982). Immune response to lactate dehydrogenase-elevating virus: isolation of infectious immunoglobulin G complexes and quantitation of specific antiviral immunoglobulin G responsein wild-type and nude mice. Infection and Immunity, 37, 1001-1006. Gerber, N. L., Hardin, J. A., Chused, T. M. and Steinberg, A. D. (1974). Losswith age in NZB/W mice of thymic suppressorcells in the graft-vs-host reaction. Journal of Immunology, 113, 1618-1625. Hayashi, T., Mori, I., Ozaki, M., Saito, M., Itoh, T. and Yamamoto, H. (1992). Enhanced lactic dehydrogenase-5 in severe combined immunodeficiency mice: effect of lactic dehydrogenase virus on enzyme clearance. International Journal of Experimental
Pathology, 73, 173-18 1.
Hayashi, T., Salata, K., Kingman, A. and Notkins, A. L. (1988). Regulation of enzyme levels in the blood: influence of environmental and genetic factors on enzyme clearance. American 3ournal of Pathology, 132, 503-5 11. Heremans, H., Billiau, A., Coutelier, J. P.and DeSomer, P. (1987). The inhibition of endotoxin-induced local inflammation by LDH virus or LDH virus-infected tumors is mediated by interferon. Proceedings of the Societyfor Experimental Biology and Medicine, 185, 6-15. Isakov, N., Feldmann, M. and Segal, S. (1982). Acute infection of mice with lactic dehydrogenase virus (LDV) impairs the antigen-presenting capacity of their macrophages. Cellular Immunology, 66, 3 17-332. Kelly, V. E. and Roths, J. B. (1982). Increase in macrophage Ia expression in autoimmune mice: role of the lpr gene. Journal of Immunology, 129, 923-925. Notkins, A. L. (1965). Lactic dehydrogenasevirus. Bacteriological Reviews, 29, 143-160. Notkins, A. L., Mahar, S., Scheele, C. and Goffman, J. (1966). Infectious virus-antibody complex in the blood of chronically infected mice. Journal of Experimental Medicine, 124, 8 l-97. Oldstone, M. B. A., Yamazaki, S., Niwa, A. and Notkins, A. L. (1973). In vitro detection of cells infected with lactic dehydrogenase virus (LDV) by fluorescine-labeled antibody to LDV. Intervirology, 2, 261-265. Porter, D. D., Porter, H. G. and Deerhake, B. B. (1969). Immunofluorescence assayfor antigen and antibody in lactic dehydrogenase virus infection. Journal oj Immunology,
102,431-436.
Prud’homme, G. J., Balderas, R. S., Dixon, F. J. and Theofilopoulos, A. N. (1983). B cell dependence on and responseto accessory signals in murine lupus strains. Journal of Experimental Medicine, 157, 18181827. Rowson, K. E. K. and Mahy, B. W. J. (1985). Lactate dehydrogenase-elevating virus. Journal of General Virology, 66, 2297-2312. Stevenson, M. M., Rees,J. C. and Meltzer, M. S. (1980). Macrophage function in tumor-bearing mice: evidence for lactic dehydrogenase-elevating virus associated changes. Journal of Immunology, 124, 2892-2894.
1
Received, March 9th, 1992 Accepted, May 2Oth, 1992