Production and utilization of polyclonal antibodies against nisin in an ELISA and for immuno‐location of nisin in producing and sensitive bacterial strains

• Published in: Journal of applied microbiology Vol. 87; no. 4; pp. 500 - 510
• Main Authors: Bouksaim, M, Lacroix, C, Bazin, R, Simard, R.E
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Science Ltd 01.10.1999; Blackwell Science
• Subjects: Animals; Bacteriocins - analysis; Bacteriology; Biological and medical sciences; Chromatography, Affinity - methods; Enzyme-Linked Immunosorbent Assay - methods; Female; food preservatives; Fundamental and applied biological sciences. Psychology; hemocyanins; Immune Sera - biosynthesis; Immune Sera - isolation & purification; Immunoglobulin G - biosynthesis; keyhole limpet hemocyanin; lactococcus diacetylactis; Lactococcus lactis - chemistry; Lactococcus lactis lactis; Lactococcus lactis subsp. lactis; Lactococcus lactis subsp. lactis bv. diacetylactis; Microbiology; Microscopy, Electron; Milk - microbiology; Nisin - analogs & derivatives; Nisin - analysis; Nisin - immunology; nisin Z; Pathogenicity, virulence, toxins, bacteriocins, pyrogens, host-bacteria relations, miscellaneous strains; Rabbits; Nisin; Antibody; Bacteriocin; Method; Biological activity; Antimicrobial agent; Streptococcaceae; Lactococcus lactis subsp. lactis; Bacteria; Micrococcales; Detection; Localization; ELISA assay
• ISSN: 1364-5072; 1365-2672
• DOI: 10.1046/j.1365-2672.1999.00842.x

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 μg 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.