MYCOPLASMA PNEUMONIAE ANTIGENS DEMONSTRATED BY CROSSED IMMUNOELECTROPHORESIS

SummaryHuman endothelial cells isolated from umbilical cordswere solubilized in Triton X-100 and examined by crossedimmunoelec-trophoresis using rabbit antiserum against endothelial cells. Endogenous labelling of the endothelialcell proteins with 14Cmannose followed by crossed immunoelectrophoresis and autoradiography revealed about 10 immunoprecipitates. Four of these endothelial cell glycoproteins were labelled by lactoperoxidase catalyzed iodination and thus were surface located. Three of the surface located glycoproteins showed reduced electrophoretic mobility after incubation of the endothelial cells with neuraminidase and were therefore sialoglycoproteins. Amphiphilicity of endothelial cell glycoproteins was studied by crossed hydrophobic interaction immunoelectrophoresis with phenyl-Sepharose in the intermediate gel. Amphiphilic proteins also show increasing electrophoretic migration velocity with decreasing concentration of Triton X-100 in the first dimension gels. Five of the endothelial cell glycoproteins were shown to be amphiphilic using these two techniques.Two monoclonal antibodies against the platelet glycoprotein complex Ilb-IIIa and glycoprotein IlIa, respectively, reacted with the same precipitate of endothelial cells. When a polyclonal antibody against the platelet glycoprotein complex Ilb-IIIa was incorporated into the intermediate gel the position of two endothelial cell precipitates were lowered. One of these was a sialoglycoprotein.

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The Interaction of β2 Glycoprotein-I and Heparin and Its Effect on β2 Glycoprotein-I Antiphospholipid Antibody Cofactor Function in Plasma

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648918 ◽

1994 ◽

Vol 72 (04) ◽

pp. 578-581 ◽

Cited By ~ 3

Author(s):

T McNally ◽

S E Cotterell ◽

I J Mackie ◽

D A Isenberg ◽

S J Machin

Keyword(s):

Solid Phase ◽

Pharmaceutical Preparations ◽

Enzyme Linked Immunosorbent Assay ◽

Antiphospholipid Antibody ◽

Biological Functions ◽

Dermatan Sulphate ◽

Glycoprotein I ◽

Crossed Immunoelectrophoresis ◽

Calcium Heparin ◽

Cofactor Activity

Summaryβ2 glycoprotein-I (β2GPI), a cofactor for antiphospholipid antibody (aPA) binding, binds to many anionic macromolecules including heparin. The nature of this interaction with heparin is not well understood and its effect on the purported biological functions of β2GPI is unknown.We have examined the interactions of dermatan sulphate (DS) and different pharmaceutical preparations of heparin with β2GPI by crossed immunoelectrophoresis (CIE) and investigated the effect of these agents on plasma levels of p2GPI antigen (β2GPI: Ag) by a standardised enzyme linked immunosorbent assay (ELISA). P2GPI aPA cofactor activity (β2GPI:Cof) was also measured using a modified solid phase an-ti-phosphatidylserine (aPS) ELISA. CIE results confirmed a heparin-β2GPI interaction with unfractionated (UF) heparin. β2GPI:Ag levels were unaffected by any of the preparations investigated. There were no significant differences in β2GPI:Cof activities of the samples containing LMW heparins or DS but levels of β2GPI:Cof were increased in samples containing UF sodium and calcium heparin preparations (0.5 IU/ml Monoparin, p <0.05, and 10 IU/ml Liquemin and Calcipa-rine, p <0.05).

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Binding of Human Platelet Glycoprotein lb and Actin to Fragments of Actin-Binding Protein

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648421 ◽

1992 ◽

Vol 67 (02) ◽

pp. 252-257 ◽

Cited By ~ 9

Author(s):

Anne M Aakhus ◽

J Michael Wilkinson ◽

Nils Olav Solum

Keyword(s):

Actin Filaments ◽

High Speed ◽

Binding Protein ◽

Protein A ◽

Actin Binding ◽

Platelet Glycoprotein ◽

Actin Binding Protein ◽

Crossed Immunoelectrophoresis ◽

Triton X ◽

Calpain Inhibitors

SummaryActin-binding protein (ABP) is degraded into fragments of 190 and 90 kDa by calpain. A monoclonal antibody (MAb TI10) against the 90 kDa fragment of ABP coprecipitated with the glycoprotein lb (GP lb) peak observed on crossed immunoelectrophoresis of Triton X-100 extracts of platelets prepared without calpain inhibitors. MAb PM6/317 against the 190 kDa fragment was not coprecipitated with the GP lb peak under such conditions. The 90 kDa fragment was adsorbed on protein A agarose from extracts that had been preincubated with antibodies to GP lb. This supports the idea that the GP Ib-ABP interaction resides in the 90 kDa region of ABP. GP lb was sedimented with the Triton-insoluble actin filaments in trace amounts only, and only after high speed centrifugation (100,000 × g, 3 h). Both the 190 kDa and the 90 kDa fragments of ABP were sedimented with the Triton-insoluble actin filaments.

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