Lectin‐Reactive Components in White Matter Membranes from Normal and Multiple Sclerosis Brains

Abstract

Polypeptides derived from human white matter membranes reacted with the radioiodinated lectins concanavalin A, Lens culinaris phytohemagglutinin, Ricinus communis agglutinin and wheat germ agglutinin after electrophoresis in polyacrylamide pore gradient gels. The molecular weights of these lectin-reactive bands were estimated by comparison with radioiodinated protein standards by using the linear relationship between log of the molecular weight and log of the gel concentration reached by the protein after electrophoresis in a polyacrylamide gradient gel. The molecular weight estimates for components reactive with concanavalin A were 176,800, 141,200, 72,800, 52,800, 44,700, 40,000, 24,800 and 23,900. The molecular weights of the bands reactive with both wheat germ agglutinin and Lens culinaris phytohemagglutinin were 138,000, 113,500, 92,100, 52,800, 44,700, 24,800 and 23,900. Wheat germ agglutinin was bound also to a band with a molecular weight of 72,800. Ricinus communis agglutinin bound to bands with estimated molecular weights of 138,000, 72,800, 52,800, 44,700, 24,800 and 23,900. The electrophoretic pattern of lectin-reactive polypeptides derived from normal-appearing white matter of multiple sclerosis brains was not qualitatively different from the lectin-binding pattern of control brain membrane polypeptides.

Methods for the Identification and Characterization of Glycoproteins in Central and Peripheral Myelin

“…9), one of the glycoproteins detected corresponded to MAG that was labeled with [ 14 C]fucose, but more than ten other glycoproteins were also detected. Other investigators have used overlay techniques with radioiodinated 1ectins (Poduslo et al, 1980;Hukkanen, 1982) to detect glycoproteins on SDS slab gels or incubation with fluoroscein isothiocyanate (FITC)-labeled lectins to detect glycoproteins on microgels in 1O-j-L1 capillaries (Lane et al, 1979;Linnington and Waehneldt, 1981). The methods involving binding of lectins to myelin proteins generally reveal a more complex population of glycoproteins in isolated myelin than is revealed by periodic acid-Schiff staining or by in vivo labeling.…”

Antibodies to White Matter Proteins and Viral Antigens in Multiple Sclerosis

“…The viruses were grown and labeled by 35S-methionine as described by Vainionpas et al (1978). Electrophoresis of the immunoprecipitates was performed in 8.0-18.9% acrylamide gradient gels as described previously (Hukkanen 1982). The virus-infected cell lysates and the lectin-isolated glycoprotein fractions were solubilized with 1% Triton X-100 (Tx-100) for imunoprecipitation.…”

“…The membrane fractions labeled by Na125I using the chloramine T method (Hunter and Greenwood 1962) had specific activities ranging from 0.1 to 0.2 mCi/mg of protein. Electrophoresis of the immunoprecipitates was performed in 8.0-18.9% acrylamide gradient gels as described previously (Hukkanen 1982). The viruses were grown and labeled by 35S-methionine as described by Vainionpas et al (1978).…”

“…In the precipitation of labeled glycoproteins the staphylococci were replaced by Protein A-Sepharose ( P h a mcia, Sweden), and the Sepharose gel was washed four times before extraction of the labeled antigen by 2% SDS and 5% beta-mercaptoethanol in 0.1 M Tris-HC1, pH 6.8. Electrophoresis of the immunoprecipitates was performed in 8.0-18.9% acrylamide gradient gels as described previously (Hukkanen 1982).Electrophoresis of the imunoprecipitates suggested that no specific binding of CSF IgG to the unfractionated membrane polypeptides occurs and qt? MS-associated autoantigen was revealed from MS white matter.…”

Immunochemical Analysis of Antigenic Determinants of Chlamydia trachomatis by Monoclonal Antibodies

“…Monoclonal antibody Ll/Al lost the avidity for chlamydial L1 typespecific antigen almost completely after ' 251 labelling. (Hukkanen, 1982). Each sample (60 pg) was dissociated for 3 min at 100 "C in 0.1 M-Tris/HCI pH 6.8 containing 2% (w/v) SDS, 5% (v/v) /J-mercaptoethanol, 5 % (v/v) glycerol and 0.005 7; (w/v) bromphenol blue.…”