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Immunoassays with specific antibodies offer higher sensitivity than do bioassays with indicator strains in the detection and quantification of several bacteriocins. Here we present the purification of lacticin RM and the production of specific polyclonal antibodies to a synthetic peptide resembling an internal fragment of the mature bacteriocin. The specificity and sensitivity of the generated polyclonal antibodies were evaluated in various immunoassays. The detection limits of lacticin RM were found to be 1.9, 0.16, and 0.18 g ml ؊1 for Western blot, immuno-dot blot, and noncompetitive indirect enzyme-linked immunosorbent assays, respectively. Immunoassay sensitivities were 12.5-fold higher than that of the agar diffusion test (ADT). The production of lacticin RM showed temperature dependency, with 3, 4.2, 12.7, 28.9, 37.8, and 12 g ml ؊1 at 37, 30, 20, 15, 10, and 4°C, respectively. Temperature-stability analysis demonstrated that lacticin RM is sensitive to mild temperature, but the loss of activity does not seem to result from protein degradation. Tween 80 increased the concentration of lacticin RM eightfold and probably affected the results of the ADT either by enhancing the activity of lacticin RM or by increasing the sensitivity of the indicator strain. The use of antibodies for the specific detection and quantification of lacticin RM can expand our knowledge of its production and stability, with important implications for further investigation and future application.Bacteriocins are antibacterial proteins produced by bacteria that kill or inhibit the growth of closely related bacteria. Many lactic acid bacteria (LAB) produce a high diversity of bacteriocins, with a fairly broad inhibitory spectrum (19). In recent years, many bacteriocins from LAB belonging to different groups have been described, characterized, and purified. Nisin, produced by several strains of Lactococcus lactis, is the most studied bacteriocin, and it is used as a commercial food preservative (6,11,16). The role of LAB and their bacteriocins as food biopreservatives is expected to grow in the future as a result of consumer awareness of the potential risks derived from food-borne pathogens as well as from the artificial chemical preservatives currently used to control them (14).Lacticin RM is a bacteriocin produced by Lactococcus lactis subsp. lactis EZ26 (DSM ID-95-131), isolated from goat's milk. It displays a wide range of inhibition, including Listeria monocytogenes and Staphylococcus aureus (36). The operon encoding lacticin RM is located on a 2.5-kb fragment in the plasmid pHU1, and it is rather simple in comparison to other bacteriocin systems, including only four genes. lacA is the bacteriocin structural gene, while lacF, lacG, and lacI participate in immunity against lacticin RM (36). Preliminary results indicate that lacticin RM is optimally produced at low growth temperatures, with maximal inhibition zones against an indicator strain at 10 and 15°C. Although most of the characterized bacteriocins are produced at the producer...

Clonidine has been reported to exert anti-anxiety effects in animals and man similar to those of benzodiazepines. The present experiment examined the effects of clonidine administration on the partial reinforcement extinction effect (PREE) which is known to be sensitive to benzodiazepine action. Two groups of rats were trained to run in a straight alley. The continuously reinforced (CRF) group received food reward on every trial. The partially reinforced (PRF) group was rewarded on a quasi-random 50% schedule. All animals were then tested in extinction. Clonidine 50 micrograms/kg was administered in a 2 X 2 design, i.e., drug-no drug in acquisition and drug-no drug in extinction. The PREE, i.e., increased resistance to extinction exhibited by PRF animals as compared to CRF animals, was obtained in animals that received saline in acquisition, independently of drug treatment in extinction, as well as in animals that received clonidine in both acquisition and extinction, but not in animals that received clonidine in acquisition alone. The administration of clonidine in extinction alone increased resistance to extinction in both the CRF and PRF animals. The increase in resistance to extinction, typically obtained with benzodiazepine treatment, indicates that clonidine exerts anxiolytic effects, supporting the involvement of the noradrenergic system in anxiety. However, clonidine did not fully reproduce the effects of benzodiazepines on the PREE, suggesting that the two classes of drugs may act via different noradrenergic mechanisms.

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