Determination of carazolol residues in porcine tissue by radioreceptor assay

A radioreceptor assay was developed for the determination of the p-blocker carazolol in porcine muscle and kidney. The method involves a simple alkaline extraction procedure using diethyl ether followed by a competitive assay between carazolol residues and [H-3]-dihydroalprenolol ([H-3]-DHA) using solubilised beta2-adrenoceptors isolated from a transfected cell line. The limit of detection (LOD) was determined using 20 reference blank samples of pig kidney and pig muscle. LODs for muscle (0.93 mug kg(-1)) and kidney (1.47 mug kg(-1)) were well below their respective European community maximum residue limits, (MRLs 5 and 25 mug kg(-1), respectively). The assay was used to investigate if carazolol residues persisted in pig tissues for up to 30 h post-intramuscular injection at the recommended dose rate (10 mug carazolol/kg body weight). The highest mean +/- S.D. concentrations were detected at 1 h post-injection in kidney (10.84 +/- 1.3 mug kg(-1)) and muscle (3.59 +/- 0.2 mug kg(-1)) which were less than the respective MRLs. It is concluded that this method offers a robust and rapid alternative to other methods for the screening of carazolol residues in pig meat. (C) 2002 Elsevier Science B.V. All rights reserved.

A microtitre plate assay for the detection of antibiotics in porcine urine

Techniques for screening porcine samples for antimicrobial residues in the EU usually involve analysis of samples taken post slaughter, and are either time consuming or expensive. Some of the positive test results at this screening stage could be avoided by allowing the animal sufficient withdrawal time following drug treatment. A method is described that can detect the presence of five major antibiotics in porcine urine at concentrations below 1 mu g ml(-1) for each of the compounds. The test uses Bacillus subtilis, which is already widely employed in antimicrobial inhibition assays, and when combined with a colorimetric substrate, p-nitrophenyl-beta-D-glucopyranoside, can detect inhibitory substances within an assay time of four and a half hours. The method, which uses microtitre plate technology, could be developed into a convenient test kit for use at farm level to determine whether animals were still excreting antimicrobials in their urine prior to their submission for slaughter.

Production, characterisation and applications of monoclonal antibodies to two novel porcine bocaviruses from swine in Northern Ireland

The production, preliminary characterisation and applications of monoclonal antibodies (mAbs) against two novel swine bocaviruses isolated in cell culture from swine in Northern Ireland are described. Of the 17 stable final clones produced, four were characterised. All were of the IgG2a isotype and showed no cross-reactivity with either bocavirus strain. Partial neutralisation was observed with PBoV4 mAbs and homologous virus. The two mAbs selected for use in antigen-detecting ELISAs were successful in highlighting those fractions containing infectious virus within sucrose gradients. This is the first report of the production of specific reagents that will prove useful in the study of the biology of these viruses and swine bocavirus-associated diseases.

Evaluation of Mycoplasma hyopneumoniae bacterins for porcine torque teno virus DNAs

Objective-TO determine whether commercial Mycoplasma hyopneumoniae bacterins sold for use in swine contain porcine torque teno virus (TTV).

Depletion of four nitrofuran antibiotics and their tissue-bound metabolites in porcine tissues and determination using LC-MS/MS and HPLC-UV

Depletion of the nitrofuran antibiotics furazolidone, furaltadone, nitrofurantoin and nitrofurazone and their tissue-bound metabolites AOZ, AMOZ, AHD and SEM from pig muscle, liver and kidney tissues is described. Groups of pigs were given feed medicated with one of the nitrofuran drugs at a therapeutic concentration (400 mg kg(-1)) for ten days. Animals were slaughtered at intervals and tissue samples collected for analysis for six weeks following withdrawal of medicated feed. These samples were analysed both for parent nitrofurans (using LC-MS/MS and HPLC-UV), and for tissue-bound metabolites (using LC-MS/MS). The parent drugs were detectable only sporadically and only in pigs subjected to no withdrawal period whatsoever. This confirms the instability of the four major nitrofuran antibiotics in edible tissues. In contrast, the metabolites accumulated to high concentrations in tissues (ppm levels) and had depletion half lives of between 5.5 and 15.5 days. The metabolites of all four drugs were still readily detectable in tissues six weeks after cessation of treatment. This emphasizes the benefits of monitoring for the stable metabolites of the nitrofurans.

Development of calcium phosphate cement for the augmentation of traumatically fractured porcine specimens using vertebroplasty

The study aim was to develop and apply an experimental technique to determine the biomechanical effect of polymethylmethacrylate (PMMA) and calcium phosphate (CaP) cement on the stiffness and strength of augmented vertebrae following traumatic fracture. Twelve burst type fractures were generated in porcine three-vertebra segments. The specimens were randomly split into two groups (n=6), imaged using microCT and tested under axial loading. The two groups of fractured specimens underwent a vertebroplasty procedure, one group was augmented with CaP cement designed and developed at Queen's University Belfast. The other group was augmented with PMMA cement (WHW Plastics, Hull, UK). The specimens were imaged and re-tested . An intact single vertebra specimen group (n=12) was also imaged and tested under axial loading. A significant decrease (p<0.01) was found between the stiffness of the fractured and intact groups, demonstrating that the fractures generated were sufficiently severe, to adversely affect mechanical behaviour. Significant increase (p<0.01) in failure load was found for the specimen group augmented with the PMMA cement compared to the pre-augmentation group, conversely, no significant increase (p<0.01) was found in the failure load of the specimens augmented with CaP cement, this is attributed to the significantly (p<0.05) lower volume of CaP cement that was successfully injected into the fracture, compared to the PMMA cement. The effect of the percentage of cement fracture fill, cement modulus on the specimen stiffness and ultimate failure load could be investigated further by using the methods developed within this study to test a more injectable CaP cement.

Multi sulfonamide screening in porcine muscle using a surface plasmon resonance biosensor

A binding protein displaying broad-spectrum cross-reactivity within the sulfonamide group was used in conjunction with a sulfonamide specific sensor chip and a surface plasmon resonance biosensor to develop a rapid broad spectrum screening assay for sulfonamides in porcine muscle. Results for 40 samples were available in just over 5 h after the completion of a simple sample preparation protocol. Twenty sulfonamide compounds were detected. Acetylated metabolites were not recognised by the binding protein. Limit of detection (mean-three times standard deviation value when n = 20) was calculated to be 16.9 ng g(-1) in tissue samples. Intra-assay precision (n = 10) was calculated at 4.3 %CV for a sample spiked at 50 ng g(-1) with sulfamethazine, 3.6 %CV for a sample spiked at 100 ng g(-1) with sulfamethazine, 7.2 %CV for a sample spiked at 50 ng g(-1) with sulfadiazine and 3.1 %CV for a sample spiked at 100 ng g-1 with sulfadiazine. Inter-assay precision (n = 3) was calculated at 9.7 %CV for a sample spiked at 50 ng g-1 with sulfamethazine, 3.8 %CV for a sample spiked at 100 ng g(-1) with sulfamethazine, 3.5 %CV for a sample spiked at 50 ng g(-1) with sulfadiazine and 2.8 %CV for a sample spiked at 100 ng g(-1) with sulfadiazine. (C) 2004 Elsevier B.V. All rights reserved.

Evaluation of Microcollimated Pars Plana External Beam Radiation in the Porcine Eye

Purpose: To evaluate the clinical and histological side effects of a prototype stereotactic radiotherapy system delivering microcollimated external beam radiation through pars plana in porcine eyes.

Methods: Five Yucatan mini-swine (10 eyes) were randomized to five treatment groups. Eight eyes were dosed with X-ray radiation on Day 1, and two eyes served as untreated controls. Treated eyes received doses up to 60 Gy to the retina and up to 130 Gy to the sclera using single or overlapping beams. The treatment beams were highly collimated such that the diameter was approximately 2.5 mm on the sclera and 3 mm on the retinal surface. Fundus photography, fluorescein angiography (FA), and spectral domain optical coherence tomography (SD-OCT) were obtained on days 7, 30, 60, and 110. Images were examined by a masked grader and evaluated for abnormalities. Animals were sacrificed on day 111 and gross and histopathological analysis was conducted.

Results: Histological and gross changes to eye structures including conjunctiva and lens were minimal at all doses. Fundus, FA, and SD-OCT of the targeted region failed to disclose any abnormality in the control or 21 Gy treated animals. In the 42 and 60 Gy animals, hypopigmented spots were noted after treatment on clinical exam, and corresponding hyperfluorescent staining was seen in late frames. No evidence of choroidal hypoperfusion was seen. The histological specimens from the 60 Gy animals showed photoreceptor loss and displacement of cone nuclei.

Conclusion: Transcleral stereotactic radiation dosing in porcine eyes can be accomplished with no significant adverse events as doses less than 42 Gy.

Enrichment of low-abundance proteins from bovine and porcine serum samples for proteomic studies

One of the main applications of serum proteomics is the identification of new biomarkers for animal disease or animal production. However, potential obstacles to these studies are the poor performance of affinity serum depletion methods based on human antigens when using animal samples, and loss of minor serum components bound to albumin and other proteins. In the present study, we have analyzed the efficiency and reproducibility of the ProteoMiner® beads with bovine and porcine serum samples, and compared to a traditional immunoaffinity-based albumin and IgG depletion system specific for human samples. The ProteoMiner kit is based on the use of a combinatorial peptide binding library and intends to enrich low-abundance proteins.