Isolation of specific antibody under conditions of low ionic strength

Hardie, Gwendoline; Regenmortel, M.H.V. Van

doi:10.1016/0022-1759(77)90092-8

Cited by 63 publications

(18 citation statements)

References 8 publications

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“…Blood samples were taken at weekly intervals for 2 months. The serum fractions were assayed in indirect antigen-coated plate-ELISA with alkaline phosphatase-labeled goat anti-rabbit globulins (Biosys), with a CRY solution (60 ,ug/mL) directly adsorbed onto the support; the most active fractions were pooled and the antibodies purified by the rivanol precipitation method (8). In a subsample, a conjugate was prepared by coupling the IgG to alkaline phosphatase by means of glutaraldehyde (1).…”

Section: Methodsmentioning

confidence: 99%

Migration of the Fungal Protein Cryptogein within Tobacco Plants

Devergne¹,

Bonnet²,

Panabieres³

et al. 1992

Plant Physiol.

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Cryptogein (CRY), a protein secreted by Phytophthora cryptogea, causes necrosis on tobacco (Nicotiana tabacum) plants at the site of application (the stem or the roots) and also on distant leaves. Autoradiography of plantlets after root absorption of radioiodinated CRY demonstrated a rapid migration of the label to the leaf lamina via the veins. Using an anti-CRY antiserum, a CRY-related antigen was detected in the stem and leaves of CRY-treated plants at a distance from the site of application. This antigen had the same molecular weight as CRY and was detected in the leaves as early as 1 hour after stem treatment, i.e. long before necrosis was detectable. The antigen was also detected in plants inoculated with P. cryptogea. The distant location of the necrosis induced by the fungus or by CRY can be ascribed to the migration of this protein, which is toxic to tobacco cells. It is proposed that CRY, which also elicits defense reactions in tobacco, might contribute to the hypersensitive response of tobacco to P. cryptogea.The interaction between the phytopathogenic fungus Phytophthora cryptogea and the nonhost plant tobacco (Nicotiana tabacum) leads to an HR2 characterized by plant tissue necrosis and a restricted growth of the fungus (5). An unusual feature of this HR is that necrosis occurs not only at the inoculation court but also on the leaves at a distance from the fungus. In culture, P. cryptogea secretes a protein, CRY (mol wt 10,323), which causes the rapid death of tobacco cells and is an elicitor of phytoalexin accumulation (4,6,11,12). When applied to the wounded stem or roots of a tobacco plant, CRY induces necrosis within 24 h at the point of application and also on distant leaves (14). This raises two questions: (a) does this elicitor actually move within the plant; and (b) interaction, then it should be possible to detect this proteinaceous elicitor in noninfected tissues from inoculated plants.We have investigated the possibility that CRY migrates in tobacco by two approaches: (a) autoradiography of plantlets after absorption of radioiodinated CRY by the roots; and (b) immunodetection of the protein in the stem and the leaves after application of exogenous CRY to the stem or to the roots, or after stem-inoculation with P. cryptogea. In addition, the behavior of CRY was compared with that of CAP, a related protein produced by Phytophthora capsici, which has slightly different biological effects on tobacco (14). MATERIALS AND METHODSTobacco plants (Nicotiana tabacum cv Xanthi nc) grown in a greenhouse for 30 d were transferred into a growth chamber (240C, 16 h daylength), and CRY was applied 10 d later to the stem or to the roots. Stem treatments were as previously described (14): the plant was decapitated above the third highest fully expanded leaf, and the wounded stem received 2.5 ,ug of elicitor as a 20-,uL drop of aqueous solution or a plug taken from a malt-agar culture of Phytophthora cryptogea; only the two upper remaining leaves (Li and L2) were left attached. In root treatments, t...

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Section: Methodsmentioning

confidence: 99%

Migration of the Fungal Protein Cryptogein within Tobacco Plants

Devergne¹,

Bonnet²,

Panabieres³

et al. 1992

Plant Physiol.

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“…Equivalent groups of virus-free adult X index from populations maintained on fig roots in a greenhouse were taken as controls. The immunoglobulins (IgG) used for ELISA were purified from anti-GFLV serum according to the rivanol precipitation method (Hardie and Van Regenmortel, 1977). The ELISA plates were coated with 0.05 μg/ml IgG for 3 h at 38 °C, then rinsed 3 times.…”

Section: Introductionmentioning

confidence: 99%

Vertical distribution and infectious potential of Xiphinema index (Thorne et Allen, 1950) (Nematoda: Longidoridae) in fields affected by grapevine fanleaf virus in vineyards in the Champagne region of France

Esmenjaud

Walter²,

Valentin

et al. 1992

Agronomie

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“…injections of 200 ~tg antigen (middle component and BPMV protein obtained from J. E. Johnson) in Freund's incomplete adjuvant. Immunoglobulins were prepared by rivanol precipitation using the method of Hardie & Van Regenmortel (1977). Antisera to peptides were obtained from rabbits immunized by a series of i.m.…”

Section: Methodsmentioning

confidence: 99%

Influence of the C terminus of the small protein subunit of bean pod mottle virus on the antigenicity of the virus determined using monoclonal antibodies and anti-peptide antiserum

Joisson

Regenmortel²

1991

Journal of General Virology

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Middle component particles of bean pod mottle virus (BPMV) containing small protein subunits with a cleaved C terminus were used to produce monoclonal antibodies (MAbs). All MAbs were specific for cryptotopes, i.e. epitopes present only on dissociated BPMV protein. The MAbs reacted more strongly with virus protein preparations containing the cleaved form of the small subunit than with preparations containing only the uncleaved form. It seems that the presence of additional residues at the C terminus of the intact small subunit interferes with antibody binding. Antibodies raised against synthetic peptides corresponding to the C terminus of the uncleaved small subunit reacted with both intact virions and dissociated subunits. This Cterminal region seems to play a dominant role in the antigenicity of the virus.

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Isolation of specific antibody under conditions of low ionic strength

Cited by 63 publications

References 8 publications

Migration of the Fungal Protein Cryptogein within Tobacco Plants

Migration of the Fungal Protein Cryptogein within Tobacco Plants

Vertical distribution and infectious potential of Xiphinema index (Thorne et Allen, 1950) (Nematoda: Longidoridae) in fields affected by grapevine fanleaf virus in vineyards in the Champagne region of France

Influence of the C terminus of the small protein subunit of bean pod mottle virus on the antigenicity of the virus determined using monoclonal antibodies and anti-peptide antiserum

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