Rational monoclonal antibody development to emerging pathogens, biothreat agents and agents of foreign animal disease: The antigen scale

Many factors influence the choice of methods used to develop antibody to infectious agents. In this paper, we review the current status of the main technologies used to produce monoclonal antibodies (mAbs) from the B cells of antigen-sensitized animals. While companies are adopting advanced high-throughput methods, the major technologies used by veterinary and medical research laboratories are classical hybridoma fusion and recombinant library selection techniques. These methods have inherent advantages and limitations but have many common aspects when using immunized rodents. Laboratories with expertise in both methods of antibody development have a distinct advantage in their ability to advance mAb technology. New and re-emerging infectious threats in today's world emphasize the need for quality immunoreagents and the need to maintain expertise in mAb development. We provide examples of some common applications for mAb reagents used to identify pathogens such as the SARS-coronavirus (SARS-CoV), Bacillus anthracis, and foot-and-mouth disease (FMD) virus. We also outline a framework for investigators to make rational decisions concerning which method to use to develop mAbs based upon characteristics of the pathogen under study and the intended downstream application. Lastly, we provide parameters for the immunisation of mice and a classification system which describes the expected outcome for mAb development strategies when using classes of immunogens to generate mAbs with desired activities.     

Evaluation of four DNA extraction methods for the detection of Tritrichomonas foetus in feline stool specimens by polymerase chain reaction

Feces are increasingly valued as practical samples for molecular diagnosis of infectious disease. However, extraction of polymerase chain reaction (PCR) quality DNA from fecal samples can be challenging because of coextraction of PCR inhibitors. Because the type and quantity of PCR inhibitors is influenced by diet, endogenous flora, and concurrent disease, it is unlikely that extraction method performance with human feces can be directly extrapolated to that of domestic cats. In the present study, 4 commercially available DNA extraction methods were examined for their influence on the sensitivity of PCR for the detection of Tritrichomonas foetus in feline stool. DNA was extracted from serially diluted feline-origin T. foetus trophozoites in the absence or presence of feline feces. The ZR Fecal DNA kit was identified as affording the greatest analytical sensitivity and reproducibility and was able to detect >/=10 T. foetus organisms per 100 mg feces in 100% of PCR reactions. Further, the identified extraction method could be completed in the shortest time of all kits tested.     

Ethanol Addition for Enhancing Denitrification at the Uranium Mill Tailing Site in Monument Valley,AZ

Water, Air, & Soil Pollution - Past mining and processing of uranium ore at a former uranium mining site near Monument Valley, AZ has resulted in nitrate contamination of groundwater. The...     

From tides to mixing along the Hawaiian ridge

The cascade from tides to turbulence has been hypothesized to serve as a major energy pathway for ocean mixing. We investigated this cascade along the Hawaiian Ridge using observations and numerical models. A divergence of internal tidal energy flux observed at the ridge agrees with the predictions of internal tide models. Large internal tidal waves with peak-to-peak amplitudes of up to 300 meters occur on the ridge. Internal-wave energy is enhanced, and turbulent dissipation in the region near the ridge is 10 times larger than open-ocean values. Given these major elements in the tides-to-turbulence cascade, an energy budget approaches closure.     

Use of a commercially available culture system for diagnosis of Tritrichomonas foetus infection in cats

OBJECTIVE: To evaluate the efficacy of and optimize a commercially available culture system for sensitive and specific in-clinic culture of Tritrichomonas foetus from cat feces. DESIGN: Prospective study. SAMPLE POPULATION: Samples of freshly voided feces from 117 purebred cats and pure cultures of T. foetus obtained from a cat with chronic diarrhea. PROCEDURE: Optimal conditions for use of the culture system, such as quantity of fecal inoculum (0.025 to 0.2 g) and cultivation temperature (25 or 37 degrees C [98.6 or 77.0 degrees F]), were determined. Specificity of the system was examined by attempted culture of Giardia lamblia and Pentatrichomonas hominis. Sensitivity of the system to detect T. foetus was determined by inoculation of culture system pouches with serially diluted T. foetus suspensions with and without feces. RESULTS: Detection limit of the culture system was 1 and 1,000 T. foetus organisms without and with feces from cats, respectively. Optimal fecal inoculum was < 0.1 g of feces. At 37 degrees C, cultures yielded positive results in 24 hours; organisms remained viable for 1 to 6 days, and bacterial overgrowth was common. At 25 degrees C, cultures yielded positive results in 1 to 11 days; organisms were long-lived, and bacterial overgrowth was uncommon. Neither G. lamblia or P. hominis survived in the culture system. CONCLUSIONS AND CLINICAL RELEVANCE: The culture system was sensitive and specific for culture of T. foetus in feces of cats. Performance was optimal when test kits were inoculated with < or = 0.1 g of freshly voided feces and cultured at 25 degrees C.