Production and characterization of anti-nisin Z monoclonal antibodies: suitability for distinguishing active from inactive forms through a competitive enzyme immunoassay

As a pre-requisite to monoclonal antibody development, an efficient purific ation strategy was devised that yielded 72 mg of nisin Z from 14.5 1 of Lac tococcus lactis subsp. lactis biovar. diacetylactis UL 719 (L. diacetylacti s UL719) culture in supplemented whey permeate. Specific monoclonal antibod ies (mAbs) were produced in mice against the purified nisin Z using keyhole limpet hemocyanin as a carrier protein. These antibodies did not recognize nisin A, suggesting that the asparagine residue at position 27 is involved in antibody recognition to nisin Z. However, the high reactivity of mAbs a gainst biologically inactive nisin Z degradation products, produced during storage of freeze-dried pure nisin Z at -70 degreesC, indicated that the de hydroalanine residue at position 5 (Dha(5)), required for biological activi ty, is not necessary in nisin Z recognition by the mAb. A competitive enzym e immunoassay (cEIA) using the specific anti-nisin Z mAb was developed and used for rapid and sensitive detection and quantification of nisin Z in fre sh culture supernatant, milk and whey. Detection limits of 78 ng/ml in phos phate-buffered saline, 87 ng/ml in culture supernatant, 106 ng/ml in milk a nd 90.5 ng/ml in whey were obtained for this assay. The cEIA using specific mAbs can be used to quantify nisin Z in food products.