Analysis of murine C-type virus structural proteins by rocket and crossed immunoelectrophoresis

Georgiades, J; Billiau, Alfons; Verbruggen, R.; Somer, P. De

doi:10.1016/0022-1759(77)90088-6

Cited by 2 publications

(1 citation statement)

References 3 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In addition, EID is able to resolve multiple antigens in a mixture and to establish relationships among antigens when a polyvalent antiserum is used (Svendsen, 1973; Sweet et al 1973 ;Weeke, 1973 ;Georgiades et al I977). We have used the latter capabilities in the present investigation on the number of antigens produced in cells infected with human cytomegalovirus (CMV) and on the antigenic relationships among six strains of the virus.…”

Section: Discussionmentioning

confidence: 99%

Antigens of Human Cytomegalovirus: Electroimmunodiffusion Assay and Comparison Among Strains

Sweet¹,

Tegtmeier²,

Bayer³

1979

Journal of General Virology

View full text Add to dashboard Cite

SUMMARYThe antigens of strain AD169 of human cytomegalovirus (CMV) were extracted by various methods and at different times following the appearance of cytopathic effects (c.p.e.) in infected fibroblasts. Assay with a pooled human serum in electroimmunodiffusion (EID) revealed that the most reactive preparations were obtained by shell-freeze (SF) extraction on the fourth day after 4÷c.p.e. As many as 20 antigens could be detected in the original gels, most of which were stable upon storage at 4 °C for up to 4 weeks; of these, about r4 can be reproducibly seen on photographs. EID runs on day 4 SF preparations from high-passage CMV strains C87 and Davis and low passage recent isolates VDI4, I694 and 1723 resolved, respectively, I5, 15, I3, II and 11 antigens in the original gels ( I I , 9, II, 8 and 9 are visible in photographs). Strains 1694 and 1723 shared fewer antigens with one another and with high passage strains than were shared among the latter, whereas VDI 4 had relatively large numbers of antigens common to both low and high passage strains. At least six antigens were common to all strains.Electroimmunodiffusion (EID), in which antigen is electrophoresed into agarose that contains monospecific antiserum (Laurell, I966), has been widely used as a simple, rapid and sensitive technique for quantification of antigen (reviewed by Verbruggen, I975). In addition, EID is able to resolve multiple antigens in a mixture and to establish relationships among antigens when a polyvalent antiserum is used (Svendsen, 1973; Sweet et al. 1973 ;Weeke, 1973 ;Georgiades et al. I977). We have used the latter capabilities in the present investigation on the number of antigens produced in cells infected with human cytomegalovirus (CMV) and on the antigenic relationships among six strains of the virus.CMV high-passage strains AD169, C87 and Davis, and recent isolates I694, 1723 (both from leucocytes of patients with heterophile-negative mononucleosis) and VD I4 (from the urine of a patient who reportedly had syphilis) were used to infect monolayers of human embryonic tonsil fibroblasts at an approximate m.o.i, of o.25. The viruses were allowed to adsorb to monolayers in 64 oz roller bottles at 37 °C for I h, after which IOO ml of Eagle's minimal essential medium containing 2 %1 foetal bovine serum, Ioo units penicillin/ml and IOO/zg streptomycin/ml were added to each bottle. Monolayers showed > 90 ~o cytopathic effects (4+c.p.e.) within 24 h when infected with AD169, within 2 to 3 days when infected with C87, VD14, 1694 or I723 and within 3 to 5 days when infected with Davis.Antigens were prepared from cultures on the first and fourth days (all CMV strains) and on the seventh day (ADI69) after 4+c.p.e. A shell-frozen (SF) antigen was prepared (we thank Dr John Stewart of the Center for Disease Control, Atlanta, Ga., for the procedure for SF antigen preparation) by pouring culture medium from each bottle, washing the remaining monolayer twice with 2o ml of phosphate buffered saline (PBS, o'85 % NaCI buffered to pH 7"3 with o.oI M...

show abstract

Section: Discussionmentioning

confidence: 99%

Antigens of Human Cytomegalovirus: Electroimmunodiffusion Assay and Comparison Among Strains

Sweet¹,

Tegtmeier²,

Bayer³

1979

Journal of General Virology

View full text Add to dashboard Cite

show abstract

Immunochemical characterization and quantification of larval brine shrimp malate dehydrogenase

Hand

Conte

1982

J. Exp. Zool.

View full text Add to dashboard Cite

The cytosolic malate dehydrogenase (s-MDH) from larval brine shrimp has been immunochemically characterized by quantitative rocket immunoelectrophoresis (IEP) and antibody catalytic inhibition studies. Monospecific rabbit antiserum produced against purified naupliar s-MDH was used for rocket IEP as a quantitative assay for the cytoplasmic isozyme, Since cross-reactivity was not observed against the mitochondrial MDH (m-MDH), the assay allows specific measurement of the s-MDH in crude naupliar supernatants in the presence of contaminating m-MDH. The assay has a sensitivity of approximately 100 nanograms s-MDH protein using either purified enzyme solutions or crude supernatant preparations. Catalytic inhibition studies using the monospecific antiserum gave 85% inhibition of s-MDH but did not give significant inhibition of the m-MDH. Lack of complete s-MDH inhibition by the antiserum suggests a difference between the enzyme active site and immunological binding site. Although the antiserum was produced against s-MDH from nauplii of the San Francisco Bay population, the antiserum inhibits equally well the s-MDH of Great Salt Lake (Utah) nauplii, indicating little if any structural difference in the immunological sites of the s-MDHs from these two geographical sources. Porcine (s- and m-MDH), bovine (m-MDH), and pigeon (m-MDH) were also tried as antigens against the brine shrimp s-MDH antiserum, but antigen-antibody reactivity was not observed during rocket IEP or catalytic inhibition experiments.

show abstract

Analysis of murine C-type virus structural proteins by rocket and crossed immunoelectrophoresis

Cited by 2 publications

References 3 publications

Antigens of Human Cytomegalovirus: Electroimmunodiffusion Assay and Comparison Among Strains

Antigens of Human Cytomegalovirus: Electroimmunodiffusion Assay and Comparison Among Strains

Immunochemical characterization and quantification of larval brine shrimp malate dehydrogenase

Contact Info

Product

Resources

About