Production and characterization of monoclonal antibodies specific to chicken muscle soluble proteins

Three stable hybridoma cell lines (AH4, BC9 and CF2) have been produced which secrete monoclonal antibodies specific for chicken and turkey muscle proteins. Partial characterization by ELISA and SDS-PAGE immunoblotting indicated that the antibodies failed to cross-react with similar extracts of pork, beef, lamb, horse or rabbit. One of the cell lines (AH4) secreted a monoclonal antibody that was also capable of distinguishing between chicken and turkey by indirect ELISA.     

Psychrotrophic clostridia causing spoilage in cooked meat and poultry products.

Certain types of commercially produced noncured turkey breast and roast beef are precooked in situ, stored at 4 degrees C or below, and typically given use by dates of greater than 50 days. While of rare, sporadic occurrence, an unpleasant spoilage characterized by strong H2S odor and gas production has been observed in these products. This spoilage is due to the growth of psychrotrophic anaerobic sporeformers. Isolates from roast beef resemble Clostridium laramie while isolates from uncured turkey have been designated C. ctm for cooked turkey meat. The turkey breast isolates were characterized by temperature growth ranges, carbohydrate fermentations, and other biochemical reactions. Growth of all isolates was inhibited in broth media by 3.0% NaCl, 100 ppm nitrite, 2.0% sodium lactate, or 0.2% sodium diacetate. Inoculated studies were performed with three isolates in cooked turkey product. All three isolates grew and spoiled product at 10 and 3.3 degrees C, and one isolate grew at 0.5 and -3 degrees C. Some differences in growth were observed with the lactate and diacetate treatments in turkey meat among the three isolates. One isolate appeared to utilize the lactate, two were inhibited. Overall, 0.1% diacetate consistently delayed growth, although to different degrees, for all isolates.     

Predictive model for growth of Clostridium perfringens during cooling of cooked uncured meat and poultry

Comparison of Clostridium perfringens spore germination and outgrowth in cooked uncured products during cooling for different meat species is presented. Cooked, uncured product was inoculated with C. perfringens spores and vacuum packaged. For the isothermal experiments, all samples were incubated in a water bath stabilized at selected temperatures between 10 and 51 °C and sampled periodically. For dynamic experiments, the samples were cooled from 54.4 to 27 °C and subsequently from 27 to 4 °C for different time periods, designated as x and y hours, respectively. The growth models used were based on a model developed by Baranyi and Roberts (1994. A dynamic approach to predicting bacterial growth in food. Int. J. Food Micro. 23, 277-294), which incorporates a constant, referred to as the physiological state constant, q₀. The value of this constant captures the cells’ history before the cooling begins. To estimate specific growth rates, data from isothermal experiments were used, from which a secondary model was developed, based on a form of Ratkowsky’s 4-parameter equation. The estimated growth kinetics associated with pork and chicken were similar, but growth appeared to be slightly greater in beef; for beef, the maximum specific growth rates estimated from the Ratkowsky curve was about 2.7 log₁₀ cfu/h, while for the other two species, chicken and pork, the estimate was about 2.2 log₁₀ cfu/h. Physiological state constants were estimated by minimizing the mean square error of predictions of the log₁₀ of the relative increase versus the corresponding observed quantities for the dynamic experiments: for beef the estimate was 0.007, while those for pork and chicken the estimates were about 0.014 and 0.011, respectively. For a hypothetical 1.5 h cooling from 54 °C to 27° and 5 h to 4 °C, corresponding to USDA-FSIS cooling compliance guidelines, the predicted growth (log₁₀ of the relative increase) for each species was: 1.29 for beef; 1.07 for chicken and 0.95 log₁₀ for pork. However, it was noticed that for pork in particular, the model using the derived q₀ had a tendency to over-predict relative growth when the observed amount of relative growth was small, and under-predict the relative growth when the observed amount of relative growth was large. To provide more fail-safe estimate, rather than using the derived value of q₀, a value of 0.04 is recommended for pork.     

Isolation and characterization of Carnobacterium, Lactococcus, and Enterococcus spp. from cooked, modified atmosphere packaged, refrigerated, poultry meat

The microbiota of commercially produced, cooked and modified atmosphere packaged poultry meat was followed during storage at 3.5 degrees C for up to 7 weeks. The dominant microbiota consisted of Lactococcus raffinolactis (117 isolates), Carnobacterium divergens (61 isolates), Carnobacterium piscicola (11 isolates), Lactococcus garvieae (four isolates), Lactococcus lactis (one isolate) and Enterococcus faecalis (three isolates). All isolates were screened for production of bacteriocins. Only C. piscicola isolates produced an inhibitory substance active against other lactic acid bacteria and against several Listeria spp. Species-specific polymerase chain reaction (PCR) primers were used for the differentiation of Carnobacterium, L. raffinolactis, L. lactis, and L. garvieae strains associated with the modified atmosphere packaged poultry products. No false PCR products were observed with other closely related bacterial species.     

Production of monoclonal antibodies to bovine leukocyte cell-surface components

Fusion of murine myeloma cells with syngeneic spleen lymphocytes has led to development of hybridomas secreting antibodies. Hybrid cells retain the immortality and clonability of myeloma parents as well as the antibody-producing property of lymphocytes. Specificity of monoclonal antibody produced is based on the one lymphocyte-one antibody phenomenon and represents the most effective process for producing specific antisera. spleen cells from mice immunized with desired antigen are hybridized with nonsecreting mouse myeloma cells. Resulting hybrids are cloned and culture fluids tested for specific antibody activity to the antigen. Positive clones are cultivated in vitro and injected into mice for monoclonal antibody production. This technology has been extended to the bovine species to obtain monoclonal antisera to immunoglobulins and cell-surface components of leukocytes for study of mammary gland immunity. Recent progress in monoclonal research has led to interspecific fusion of murine myelomas with bovine lymphocytes, resulting in hybridomas that produce monoclonal bovine immunoglobulins. Monoclonal antibodies will be useful in investigations applicable to bovine research including purification of immunoglobulins, determining immunoglobulin concentration in colostrum and milk, reference reagents for bovine serology, antibody localization in tissue, gene sequencing, characterizing histocompatibility antigens, distinguishing and quantitating cell types in blood, milk, and udder tissue, and elucidating role of cell subpopulations in the immune response.     

Antioxidative effect of added tea catechins on susceptibility of cooked red meat,poultry and fish patties to lipid oxidation

The comparative antioxidant activity of added tea catechins on susceptibility of cooked and overwrapped red meat (beef and pork), poultry (chicken, duck and ostrich) and fish (whiting and mackerel) to lipid oxidation was investigated. Fresh meats, poultry and fish, purchased from a local market, were trimmed to remove bones, skin and visible fat and minced through a 4-mm plate. The minced muscle from each species was treated with either 1% NaCl (S), 300 mg tea catechins kg⁻¹ minced muscle (TC) or 1% NaCl plus 300 mg tea catechins kg⁻¹ minced muscle (TCS). Control minced muscle samples (C) contained neither NaCl nor tea catechins. Patties (50 g), prepared from treated and untreated minced muscle, were cooked until the core temperature reached 75°C, cooled down to room temperature and held in a refrigerated (4°C) and illuminated (616 lux) display cabinet for 10 days. Oxidative stability (TBARS) was measured at 3-day intervals. The susceptibility of cooked patties to lipid oxidation was closely related to lipid content, concentration of unsaturated fatty acids and presence of iron in different species. Addition of NaCl to raw minced muscle significantly (P<0.05) promoted lipid oxidation for cooked patties regardless of species sources. Tea catechins added at a level of 300 mg kg⁻¹ minced muscle significantly (P<0.01) inhibited the pro-oxidation caused by NaCl and controlled lipid oxidation for all cooked muscle patties examined. Tea catechins at concentrations greater than 300 mg kg⁻¹ were necessary to reduce oxidation for mackerel patties containing high levels of lipids and unsaturated fatty acids. The high affinity of tea catechins for the lipid bilayers of muscle and the radical scavenging abilities of tea catechins may be possible mechanisms to explain the oxidative stability in cooked muscle foods.     

Species-specific expression of various proteins in meat tissue: Proteomic analysis of raw and cooked meat and meat products made from beef,pork and selected poultry species

The aim was to search for proteins differentiating the six species (cattle, pig, chicken, turkey, duck and goose) and relatively stable during the meat aging and only slightly degraded in ready-made products. The two-dimensional electrophoresis was used for analysis of the protein profiles from raw meat and frankfurters and sausages (15 products). The observed species-specific differences in protein expression in raw meat were retained in processed products after finishing the entire technological process. Regulatory proteins, metabolic enzymes, some myofibrillar and blood plasma proteins were identified, which were characterised by the electrophoretic mobility specific to the given species. Large differences in the primary structure were observed in serum albumin, apolipoprotein B, HSP27, H-FABP, ATP synthase, cytochrome bc-1 subunit 1 and alpha-ETF. Some of these proteins have potential to be used as markers in authentication of meat products.     

如何保证熟肉制品杀菌彻底

熟肉制品以其品种多样、食用方便深受消费者的青睐.但是夏季熟肉制品的胀袋、变质现象,是制约肉制品企业发展的一个瓶颈.而导致产品出现胀袋、变质现象的主要原因为产品杀菌不彻底.     

The kinetics of cooked meat haemoprotein formation in meat and model systems

The rate of cooked meat haemoprotein formation (measured as the rate of loss of myoglobin solubility) was found, at least initially, to obey first order kinetics in meat, aqueous muscle extracts and mixtures of myoglobin and bovine serum albumin. In meat at 60 °C the rate was dependent on the species, (the pigment was formed significantly faster in lamb m. longissimus dorsi than in beef m. longissimus dorsi) and anatomical location (cooked meat haemoprotein was formed in beef m. 1. dorsi about twice as rapidly as in both beef shin and chuck (shoulder) muscle of similar pH). The rate of formation was similar in aqueous muscle extracts to that found in meat and in these systems increased with decreasing pH. The activation energies for all beef systems studied were similar and typical of those associated with protein denaturation (˜300 KJ mol⁻¹); however, that from lamb appeared to be lower (˜200 KJ mol⁻¹). The problems of using colour as an index of temperature reached, either for microbial safety (E. Coli 0157:H7 destruction) or quality are discussed in the light of these results.     

Oxymyoglobin formation in meat and poultry

Display-packaged samples of beef steak and of ground beef were delivered to the homes of 120 panelists. Each panelist received a set of samples of each meat. Each set consisted of one sample freshly prepared for retail display from vacuum-packaged product, one retail-packaged and then stored in a master pack under N₂ and one retail-packaged and the stored under CO₂. The samples in each set were prepared from the same strip loin or batch of coarsely ground beef and had been stored for between 21 and 23 days at 2°C. The panelists completed questionnaires on the attributes of the meats while they were packaged, when they were unpackaged for cooking, and when they were eaten. The responses to each question were tabulated, and the probability of the χ² statistics was calculated for each table.

There were no significant differences in the general acceptability of the steaks from the three storage treatments. Significant numbers of panelists judged that ground beef prepared from vacuum-packaged product was of better colour and had less exudate, but was of poorer eating quality than the product from master packs. However, the majority of panelists did not distinguish between ground beef from the three storage treatments. The consumer responses indicate that controlled-atmosphere master packaging of display packs may offer a means of preserving display-ready beef for times that would allow wide distribution of the product through current commercial systems.     

Extended shelf-life of unrefrigerated prerigor cooked meat

Five experiments were conducted to evaluate the microbial quality of unrefrigerated cooked prerigor beef after the application of oxygen-permeable packaging. Specific objectives were to combine the beneficial effects of aerobic packaging, meat surface acidification and prerigor rapid cooking rates on meat storage stability at ambient temperature. In the experiments, the triceps brachii muscle was dissected from one side 45 min after exanguination of the animal, and samples of 2 × 3 × 5 cm were prepared. Bags made of a strong barrier, plastic film, and a highly oxygen permeable oriented polypropylene, were used. The cooking of the packaged samples covered a number of treatments ranging from cooking in a 70°C water bath to an internal sample temperature of 65°C to cooking in 100°C water for 40 min. Reheating and multiple heat treatments were also included. The pH values of the cooked samples were determined and aerobic plate counts (log(10)/g) were determined for the cooked samples at various intervals during two weeks of storage at 22°C or at 3°C for the control samples. The results indicate that heat treatment alone did not improve shelf stability at 22°C. However, dipping the samples in 80°C solutions of 0·7% citric acid or 1·25% lactic acid for 1 min and draining for 1 min followed by packaging using oxygen permeable bags and cooking in 100°C water for 40 min consistently resulted in shelf stable products at all 22°C storage intervals. The lactic acid treatment was superior to the citric acid treatment because it completely decontaminated the samples and delayed spoilage, especially at 3°C.     

Molecular characterization of Listeria monocytogenes isolated from a poultry further processing facility and from fully cooked product

This study was undertaken to explore environmental sources of Listeria monocytogenes in a commercial chicken further processing facility and to compare the isolates obtained from this facility with others obtained from fully cooked product. In a survey conducted at the processing facility, 40 environmental sites (encompassing two production lines and representing areas in which raw and cooked products are processed) were cultured for L. monocytogenes. The resulting isolates were subjected to molecular subtyping by ribotyping, and these isolates were compared with 25 isolates collected by plant personnel from product contact surfaces and from fully cooked product. Eighty-nine environmental and product isolates were divided into 15 distinct ribogroups. Two ribogroups included isolates from fully cooked product; the members of these two ribogroups were subjected to further analysis by pulsed-field gel electrophoresis, resulting in four clusters. L. monocytogenes isolates from fully cooked product produced on line 1 were found to be indistinguishable from isolates collected from (i) drains on the raw-product side of line 1 and (ii) the floor surface in the cooked-product area of line 1. L. monocytogenes isolates from fully cooked product from line 2 were found to be indistinguishable from isolates collected from (i) the spiral freezer exit conveyor on line 2, (ii) raw product contact surfaces on line 1, and (iii) drains in the cooked-product area of line 1. These data suggest that L. monocytogenes can colonize a poultry further processing facility and eventually be transferred to fully cooked product.     

Evaluation of protein aggregation in cooked meat

The effect of meat cooking on protein aggregation was measured in pig M.Longissimus dorsi. Muscles were aged 4 days in air and then cooked at 100 °C for 10 or 30 min. Meat was ground in a KCl solution and the whole extract was delipidated with a mixture of butanol and di-isopropyl ether in a 40:60 v/v ratio. Protein aggregation induced by cooking was evaluated with a laser granulometer, which enabled reliable and reproducible characterisation of particle size distributions, and particle shape distributions using automated imaging techniques. Cooking significantly decreased the level of big particles, while the number of small particles remained stable. Cooking also affected the form and size of the particles. In order to better understand the mechanisms implicated in the aggregation process the level of protein oxidation and the protein surface hydrophobicity were evaluated in parallel with the granulometry measurements. Significant correlations were observed between the granulometry parameters and some of the protein oxidation indices. The protein surface hydrophobicity was also correlated with the granulometry parameters demonstrating the impact of thermal denaturation on the aggregation process.     

Isolation and molecular characterization of antibiotic-resistant lactic acid bacteria from poultry and swine meat products

The transfer via the food chain from animals to humans of microbes that are resistant to antimicrobial agents is of increasing concern. To determine the contributions of nonpathogenic microflora to the occurrence and spread of antibiotic resistance (AR) genes in the food chain, 123 lactic acid bacteria were isolated from 29 samples of raw and processed pork and chicken meat products that had previously tested positive for one or more AR genes that encode clinically relevant ARs: tet(M), tet(O), tet(K), erm(A), erm(B), erm(C), aac (6')-Ie aph (2")-Ia, mecA, and blaZ. All of the isolates were initially tested for their AR gene profiles by PCR. The 59 isolates carrying a tet, erm, or blaZ gene were taken through molecular identification, analyzed by determination of the MIC, and subjected to genetic fingerprinting. Lactococcus garvieae was the predominant species (28 isolates), followed by Lactobacillus plantarum (11 isolates) and L. salivarius (6 isolates), whereas Lactococcus lactis subsp. lactis, Lactobacillus johnsonii, L. reuteri, L. crispatus, and L. brevis were identified at lower frequencies. The tet(M) and erm(B) genes were the most frequently detected. Assessment of multiple resistances in 18 tet positive (tet+) isolates revealed that tet(M) plus erm(B) and tet(K) plus erm(B) were the most frequent AR gene patterns. Partial sequencing of the tet(M) open reading frame of three selected strains showed high sequence similarities (> 99%) with tet(M) genes previously found in human pathogens (Listeria monocytogenes and Neisseria meningitidis). Southern hybridization with plasmid profiles revealed these strains contained tet(M)-carrying plasmids.     

Isoelectric variants of actin in raw and cooked meat

Isoelectric focusing of myofibrillar proteins from raw meat gave one major and several minor actin bands. Extending the time, or raising the temperature, of a heat treatment progressively increased the proportion of variants of isoelectric point lower than the primary component. Because of the concomitant release of ammonia on heating, it was concluded that these variants arose from the hydrolysis pf amide groups of asparagine and/or glutamine. Determination of the relative proportion of deamidated actin components may provide a measure of the severity of heat treatment given to a sample of cooked meat.     

高特异性黄曲霉毒素B_1单克隆抗体的制备及特性研究

本研究将黄曲霉毒素B1转化为半缩醛B2a,在硼氢化钠（NaBH4）还原作用下与载体蛋白偶联制备完全抗原。将制备的完全抗原免疫Balb/c小鼠,经4次免疫后取其脾脏与小鼠骨髓瘤细胞Sp2/0细胞融合,采用半固体培养基筛选后鉴定,获得杂交瘤细胞株3A12,抗体的灵敏度可达6.1±0.025ng/mL,抗体与其它黄曲霉毒素B2、G1及G2的交叉反应率依次为7.8%、20.2%及0.6%,与黄曲霉毒素M1交叉反应率小于0.1%。本研究为研发花生等农产品黄曲霉毒素B1特异性免疫分析技术及产品奠定了重要基础。     

Prevalence of Campylobacter spp. in poultry and poultry meat in Germany

Of 509 samples from poultry flocks, 209 isolates (41.1%) were Campylobacter positive. The number of positive cases in broiler carcasses was 45.9%. Of 52 pheasants investigated, 25.9% were Campylobacter positive. Campylobacter jejuni was isolated from 86 (42.0%) poultry flock samples, 47 (43%) broiler samples and 15 (28%) wild pheasant samples. C. coli was found at a rate of 1.2% in poultry flocks, 13% in broilers and 21% in pheasants.     

Occurrence of heterocyclic amines in cooked meat products

Heterocyclic amines (HCAs), potent mutagens and a risk factor for human cancers, are produced in meats cooked at high temperature. The aim of this study was to determine the HCA content in cooked meat products (beef, chicken, pork, fish) prepared by various cooking methods (pan frying, oven broiling, and oven baking at 170 to 230 °C) that are preferred by U.S. meat consumers. The primary HCAs in these samples were PhIP (2-amino-1-methyl-6-phenylimidazo [4,5-b]pyridine) (1.49-10.89 ng/g), MeIQx (2-amino-3,8-dimethylimidazo [4,5-f]quinoxaline) (not detected-4.0 ng/g), and DiMeIQx (2-amino-3,4,8-trimethyl-imidazo [4,5-f]quinoxaline) (not detected-3.57 ng/g). Type and content of HCAs in cooked meat samples were highly dependent on cooking conditions. The total HCA content in well-done meat was 3.5 times higher than that of medium-rare meat. Fried pork (13.91 ng/g) had higher levels of total HCAs than fried beef (8.92 ng/g) and fried chicken (7.00 ng/g). Among the samples, fried bacon contained the highest total HCA content (17.59 ng/g).     

冷冻即食熟虾仁加工工艺

冷冻即食熟虾仁是采用南美白对虾为原料,经去头、蒸煮、去壳、调味、冻结等工艺而成,它具有口味鲜美、营养丰富、食用安全方便等特点,深受广大消费者的欢迎。     

Determination of bovine lactoferrin concentrations in cheese with specific monoclonal antibodies

In order to determine bovine lactoferrin concentration in cheese, bovine lactoferrin-specific monoclonal antibodies have been raised and an ELISA has been developed to determine lactoferrin concentrations in milk, whey and experimental soft, semi-hard and Swiss-type cheeses made with raw or pasteurised milk. The lactoferrin concentration in cheese was shown to depend on the cheese-making process, with higher values in Swiss-type and semi-hard cheeses than in soft cheeses. Furthermore, Western-blotting analysis of lactoferrin in cheese showed that this protein stayed intact throughout ripening in raw milk cheese, whereas it was partially hydrolysed in cheeses made with pasteurised milk. Based on these observations, we propose that cheese may constitute a natural dairy source of lactoferrin beneficial to health.