Isolation of the dengue virus envelope glycoprotein from membranes of infected cells by concanavalin A affinity chromatography

Stohlman, S. A.; Eylar, Ollie R.; Wisseman, Charles L.

doi:10.1128/jvi.18.1.132-140.1976

Cited by 26 publications

(6 citation statements)

References 30 publications

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“…The importance of these parameters is also reflected in different reports on the isolation of immunogenic subunits from flaviviruses. Stohlman et al (20) compared the antigenicity of membrane-bound V3 from dengue virus with that after purification by binding to concanavaiin A and elution in a sodium deoxycholatecontaining buffer. The solubilized form of the glycoprotein had lost its capability to block neutralizing antibodies, and, as suggested by the authors, this is most likely due to conformational changes caused by detergent binding.…”

Section: Discussionmentioning

confidence: 99%

Antigenic and immunogenic properties of defined physical forms of tick-borne encephalitis virus structural proteins

Heinz¹,

Tuma²,

Künz³

1981

Infect Immun

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Polymeric, delipidated glycoprotein complexes of defined size and composition were prepared from tick-borne encephalitis virus by solubilization with Triton X-100 or cetyltrimethylammonium bromide, followed by centrifugation into detergent-free sucrose density gradients. The antigenic reactivities and immunogenicities of these complexes were compared with those of complete inactivated virus. These glycoprotein preparations induced hemagglutination-inhibiting and neutralizing antibodies which proved to be protective in passive mouse protection tests and monospecifically reacted only with the viral envelope and not with the internal core. In a competitive radioimmunoassay the glycoprotein complexes revealed about 10-fold higher antigenicity than whole virus when tested at equal protein concentrations. The important implications of these results with respect to antigen quantification in vaccines are discussed. As shown in the mouse challenge potency test, glycoprotein complexes prepared after Triton X-100 solubilization actively protected mice almost as well as did complete inactivated virus at the same protein concentration, whereas those prepared after cetyltrimethylammonium bromide solubilization had a somewhat lower protective activity per microgram of protein.

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Section: Discussionmentioning

confidence: 99%

Antigenic and immunogenic properties of defined physical forms of tick-borne encephalitis virus structural proteins

Heinz¹,

Tuma²,

Künz³

1981

Infect Immun

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“…Dalrymple et al (10) recently reported the use of a similar procedure for isolating alphavirus structural proteins in an antigenically active form. Ultracentrifugation of nonionic detergent-treated flaviviruses permits separation of the nucleocapsid from the glycoprotein and membrane but does not facilitate separation of the viral envelope components (19,33,34,37,38). SDS solubilization of JE virus permitted chromatographic separation of the envelope glycoprotein but did not resolve the membrane 1to…”

Section: Resultsmentioning

confidence: 99%

“…In both CF and immunodiffusion tests, this isolated glycoprotein is serologically broadly cross-reactive (19,26,27). Dengue virus envelope glycoprotein isolated by concanavalin A chromatography is type specific by CF and does not react with neutralizing antibodies (34). We now report additional studies on the antigenic properties of purified flavivirus proteins: the large glycosylated envelope protein with a molecular weight of 53,000 to 58,000, and the nucleocapsid protein with a molecular weight of about 14,000 (32-39, 41, 42).…”

mentioning

confidence: 93%

“…The major envelope protein of JE and SLE viruses, released from the visions or infected cells after detergent treatment, binds to erythrocytes (17,34), reacts with antibodies in the CF and immunodiffusion tests (13,17,24,42), and induces the formation of neutralizing antibodies (12,26,27). In both CF and immunodiffusion tests, this isolated glycoprotein is serologically broadly cross-reactive (19,26,27).…”

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confidence: 99%

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Antigenic characterization of flavivirus structural proteins separated by isoelectric focusing

Trent¹

1977

J Virol

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Isoelectrofocusing of nonionic-detergent-disrupted flaviviruses separated the envelope glycoprotein of 53,000 to 58,000 daltons and the nucleocapsid protein of 14,000 daltons. The envelope protein and nucleocapsid protein were isolated at isoelectric points of pI 7.8 and 10.3, respectively. The antigenic determinants of St. Louis encephalitis, Japanese encephalitis, and dengue virus envelope and nucleocapsid proteins were examined by solid-phase competition radioimmunoassay. By the appropriate selection of antiserum and competing proteins, it was possible to distinguish type-specific, complex-reactive and flavivirus groupreactive antigenic determinants. The envelope glycoproteins of St. Louis encephalitis, Japanese encephalitis, and dengue viruses were found to contain each of these three classes of antigenic determinants. Most of the determinants on the envelope protein were type specific, some were complex reactive, and a small fraction were flavivirus group reactive. The nucleocapsid protein contained only flavivirus group-reactive antigenic determinants. Purified flaviviruses contain three classes of antigenic determinants: type specific, complex reactive, and flavivirus group reactive. Type-MATERIALS AND METHODS Viruses and cells. Virus strains used in these experiments were the Tampa Bay human-28 strain of SLE and the Nakayama, JaGAr-01, and Yokoshiba strains of JE viruses. Four prototype dengue 608

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“…ConA-Sepharose column chromatography. Affinity chromatography of glycoproteins on ConA-Sepharose columns was carried out as described by Stohlman et al (26), with the following modifications. All steps were at 4°C.…”

Section: Ad2 Early Glycoprotein 315mentioning

confidence: 99%

Evidence for an Adenovirus Type 2-Coded Early Glycoprotein

Jeng

Wold

Green

1978

J Virol

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We have identified an adenovirus type 2 (Ad2)-induced early glycopolypeptide with an apparent molecular weight of 20,000 to 21,000 (20/21K), as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The 20/21K polypeptide could be labeled in vivo with [3H]glucosamine. [35S]methionineand [3H]glucosamine-labeled 20/21K polypeptides bound to concanavalin A-Sepharose columns and were eluted with 0.2 M methyl-aD -mannoside. The pulse-labeled polypeptide appeared as a sharp band with an apparent molecular weight of 21K, but after a chase it converted to multiple bands with an average molecular weight of 20K. This variability in electrophoretic mobility is consistent with glycosylation or deglycosylation of the 20/21K polypeptide. Analysis of the pulse and pulsechase-labeled forms by using partial proteolysis indicated that the polypeptides were highly related chemically, but not identical. Most of the 20/21K polypeptide is localized in the cytoplasm fraction of infected cells lysed by Nonidet P-40. The 20/21K polypeptide and a 44K polypeptide, labeled with [35S]methionine or [3H]glucosamine in Ad2-infected human cells, were precipitated by a rat antiserum against an Ad2-transformed rat cell line (T2C4), but not by antisera against three other Ad2-transformed rat cell lines, or by serum from nonimmune rats. The partial proteolysis patterns of the 20/21K and the 44K polypeptides were indistinguishable, indicating that the two polypeptides are highly related, and suggesting that the 44K polypeptide might be a dimer of the 20/21K polypeptide. The 20/21K polypeptide was also induced in Ad2-early infected monkey and hamster cells. These results imply that the 20/21K polypeptide is synthesized in Ad2-infected human, monkey, and hamster cells, and in one but not all Ad2transformed rat cells. Thus, the 20/21K polypeptide is probably viral coded rather than cell coded and viral induced.

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Isolation of the dengue virus envelope glycoprotein from membranes of infected cells by concanavalin A affinity chromatography

Cited by 26 publications

References 30 publications

Antigenic and immunogenic properties of defined physical forms of tick-borne encephalitis virus structural proteins

Antigenic and immunogenic properties of defined physical forms of tick-borne encephalitis virus structural proteins

Antigenic characterization of flavivirus structural proteins separated by isoelectric focusing

Evidence for an Adenovirus Type 2-Coded Early Glycoprotein

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