Use of Iodogen and sulfosuccinimidobiotin to identify and isolate cuticular proteins of the filarial parasite Brugia malayi.
The cuticle of filarial nematodes is a dynamic structure which may be an important target for protective host immune responses. Prior studies have employed radioiodination of intact parasites to demonstrate that the collagenous cuticle of filariids contains relatively few exposed proteins, some of which are stage and/or species-specific. In the present study, we have used sulfo-NHS-biotin to label and affinity purify cuticular components of living adult Brugia malayi. Results obtained by this method were compared with the widely used Iodogen method of surface radioiodination by SDS-PAGE analysis of detergent-solubilized worms and by ultrastructural analysis. Both labeling methods produced very similar electrophoretic patterns with major doublets at 70 and 100 kDa, a major band at 25 kDa, and minor bands between 60-200 kDa. Ultrastructural analysis showed that both methods labeled components throughout all levels of the parasite cuticle; underlying somatic tissues were not labeled. The biotinylated components were isolated from the total parasite extract by affinity chromatography on an avidin matrix. Further characterization of these surface-associated proteins may lead to improved methods for the control of filariasis.
Alvarez
R M
RM
Division of Infectious Diseases, Jewish Hospital, Washington University Medical Center, St. Louis, MO 63110.
Henry
R W
RW
Weil
G J
GJ
D000818:Animals; D001710:Biotin / Q000031:analogs & derivatives*; D002009:Brugia / Q000032:analysis* / Q000648:ultrastructure; D002846:Chromatography, Affinity; D002847:Chromatography, Agarose; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D008565:Membrane Proteins / Q000032:analysis* / Q000302:isolation & purification; D013388:Succinimides; D014508:Urea / Q000031:analogs & derivatives*
D008565:Membrane Proteins; D013388:Succinimides; C053827:biotinyl-N-hydroxysulfosuccinimide ester; C012385:1,3,4,6-tetrachloro-3 alpha,6 alpha-diphenylglycoluril; D001710:Biotin; D014508:Urea