Immunologic characterization and specificity of three monoclonal antibodies against the 58-kilodalton protein of Legionella pneumophila

Abstract

Three monoclonal antibodies against the Legionella pneumophila 58-kDa protein were produced. By using immunoblot analysis, the percentages of reactivity against 47 serogroups of Legionella representing 29 species were determined to be 80.9, 87.2, and 95.6 for monoclonal antibodies GB5BE8, GB5AF6, and CA4AF5, respectively. Specificities obtained from testing 63 heterologous organisms representing 22 genera and 46 species were 90.7, 92.2, and 95.3% for monoclonal antibodies GB5BE8, GB5AF6, and CA4AF5, respectively. No single heterologous strain was reactive with all three monoclonal antibodies. These monoclonal antibodies successfully identified all 10 clinical isolates of Legionella examined in a dot blot assay and should be excellent reagents for use in genuswide diagnostic immunoassays.

Purification and seroreactivity of pneumococcal surface adhesin A (PsaA)

“…Following a final dialysis against 150 mM glycine buffer (pH 10) to solubilize insoluble components resulting from dialysis, the protein solution was centrifuged first at 12,000 ϫ g at 4ЊC for 20 min to remove minute, visible particles and then at 105,000 ϫ g for 1 h at 4ЊC. A small pellet was discarded and the supernatant was analyzed for PsaA by sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE) and Western blot (immunoblot) analysis as described previously (11).…”

“…Focusing of the proteins was carried out for 5.5 h with cooling at 12 W of constant power with a 2,000-V limit. Twenty fractions (2.5 ml each) were collected, separated by SDS-PAGE, and silver stained (11) to determine the locations of the proteins. The PsaA focused into fractions 1 to 6 and could be seen in a silver-stained gel (Fig.…”

Τυποποίηση στελεχών λεγεωνέλλας pneumophila ορότυπων 2-14 και ανίχνευση γονιδίων παθογονικότητας

“…Μνλνθισληθά αληηζψκαηα έλαληη αληηγφλσλ ηνπ γέλνπο έρνπλ παξαρζεί γηα λα δηεπθνιχλνπλ ηνλ πξνζδηνξηζκφ ησλ εηδψλ. Αξθεηνί εξεπλεηέο έρνπλ παξάμεη κνλνθισληθά αληηζψκαηα έλαληη ηεο Legionella heatshock πξσηεΐλεο κε πνηθίιινπ βαζκνχ εηδηθφηεηα (Sampson et al, 1991;Steinmetz et al,1991). Οη Helbig et al παξήγαγαλ κνλνθισληθά αληηζψκαηα έλαληη ηεο πξσηεΐλεο mip ηα νπνία αληέδξαζαλ κε 82 ζηειέρε Legionella πνπ αληηπξνζψπεπαλ 34 είδε (Helbig et al, 1995).…”

“…Έλα εκπνξηθά δηαζέζηκν κνλνθισληθφ αληίζσκα (Genetic Systems, Seattle, Wash) ην νπνίν αληηδξά κε πξσηεΐλε ηεο εμσηεξηθήο κεκβξάλεο θαη αληρλεχεη φινπο ηνπο νξνηχπνπο ηεο L.pneumophila κπνξεί λα ρξεζηκνπνηεζεί γηα ηελ αλίρλεπζε ηεο Λεγεσλέιιαο ζε θιηληθά δείγκαηα (Gosting et al, 1984 (Edelstein etal, 1985). Μνλνθισληθά αληηζψκαηα γηα ην γέλνο έρνπλ παξαρζεί γηα λα αληρλεχνπλ είηε ηελ heat shock πξσηεΐλε ή ηελ mip πξσηεΐλε, αιιά δε θαίλεηαη λα είλαη ρξήζηκα γηα ηελ αλίρλεπζε ηεο Λεγεσλέιιαο ζε θιηληθά δείγκαηα ( Helbig et al, 1995;Helsel et al,1988;Sampson et al, 1991;Steinmetz et al,1991). (Farshy et al,1979;Farshy et al,1978;Klein et al, 1979).…”