Production and utilization of polyclonal antibodies against nisin in an ELISA and for immuno-location of nisin in producing and sensitive bacterial strains.
Specific nisin polyclonal antibodies (PAb) were produced in rabbits using nisin Z produced by Lactococcus lactis subsp. lactis biovar diacetylactis UL 719. Antisera were obtained from white female New Zealand rabbits that were first immunized with a nisin Z-keyhole limpet haemocyanin conjugate and boosted with free nisin Z. Nisin-specific PAb were purified by affinity chromatography with a yield of 15 mg specific antinisin 100 ml-1 serum. The detection limit of the ELISA test for nisin Z was 0.75 ng ml-1 in buffer but was 1.7 and 3.5 ng ml-1 in milk and complex media broth spiked (5, 10, 20 microg ml-1) with nisin Z, respectively. In nisin Z-spiked samples, the average concentration was between 90 and 107% of actual added amount. In contrast, when the bioassay (microtitration method) was used, only 50-63% of nisin Z biological activity could be detected. In addition, the affinity-purified nisin PAb, antirabbit IgG gold conjugate and transmission electron microscopy were successfully used to locate nisin Z on producing cells and to observe its bactericidal effects against sensitive cells.
Bouksaim
M
M
Centre de recherche en sciences et technologie du lait, Université Laval, Sainte-Foy, Québec, Canada.
Lacroix
C
C
Bazin
R
R
Simard
R E
RE
D000818:Animals; D001430:Bacteriocins / Q000032:analysis; D002846:Chromatography, Affinity / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D007106:Immune Sera / Q000096:biosynthesis* / Q000302:isolation & purification; D007074:Immunoglobulin G / Q000096:biosynthesis; D013294:Lactococcus lactis / Q000737:chemistry*; D008854:Microscopy, Electron; D008892:Milk / Q000382:microbiology; D009561:Nisin / Q000031:analogs & derivatives* / Q000032:analysis / Q000276:immunology; D011817:Rabbits
D001430:Bacteriocins; D007106:Immune Sera; D007074:Immunoglobulin G; D009561:Nisin; C071364:nisin Z