Cloning of canine gamma-tubulin (TUBG1) cDNA and mapping to CFA9.
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Animals↗
Biomedical subjects
E Ostrander
Publications and source records attributed to E Ostrander.
Interpreting epidemiological research: blinded comparison of methods used to estimate the prevalence of inherited mutations in BRCA1.
While sequence analysis is considered by many to be the most sensitive method of detecting unknown mutations in large genes such as BRCA1, most published estimates of the prevalence of mutations in this gene have been derived from studies that have used other methods of gene analysis. In order to determine the relative sensitivity of techniques that are widely used in research on BRCA1, a set of blinded samples containing 58 distinct mutations were analysed by four separate laboratories. Each used one of the following methods: single strand conformational polymorphism analysis (SSCP), conformation sensitive gel electrophoresis (CSGE), two dimensional gene scanning (TDGS), and denaturing high performance liquid chromatography (DHPLC). Only the laboratory using DHPLC correctly identified each of the mutations. The laboratory using TDGS correctly identified 91% of the mutations but produced three apparent false positive results. The laboratories using SSCP and CSGE detected abnormal migration for 72% and 76% of the mutations, respectively, but subsequently confirmed and reported only 65% and 60% of mutations, respectively. False negatives therefore resulted not only from failure of the techniques to distinguish wild type from mutant, but also from failure to confirm the mutation by sequence analysis as well as from human errors leading to misreporting of results. These findings characterise sources of error in commonly used methods of mutation detection that should be addressed by laboratories using these methods. Based upon sources of error identified in this comparison, it is likely that mutations in BRCA1 and BRCA2 are more prevalent than some studies have previously reported. The findings of this comparison provide a basis for interpreting studies of mutations in susceptibility genes across many inherited cancer syndromes.
Chromatography, High Pressure Liquid↗
Noninvasive quantification of muscle oxygen in subjects with and without claudication.
The disabling pain of intermittent claudication (IC) arises from oxygen deprivation in the lower limbs during walking. Measurement of the oxygen deficiency within the limb tissue now appears possible with recently expanded understanding of the photon transport through tissue for photons in the visible and near infrared range. Noninvasive measurement consists of preferentially measuring photons that have traveled more deeply into limb tissues and that, therefore, may reach locations of ischemic tissue. Oxygen measurements appear to be possible up to a depth approaching 1.5 cm beneath the surface of the skin. The present study reports on data acquired from the limbs of 11 subjects with IC and 12 subjects without IC. The subjects with IC are patients with clinical findings of claudication based upon segmental Doppler pressure profiles and subjective reports by the patient of pain during exercise. The subjects without IC are individuals with no prior history of ischemic vascular disease. The results consist of photon reflectance measurements at red and infrared wavelengths (approximately 660 nm and 880 nm respectively) taken before, during, and after exercise. Infrared reflectance indices are plotted as well as oxygenation indices generated from combining red and infrared reflectances. A compilation of exercise data shows responses that are generally consistent with the expected physiological responses to mild exercise in subjects with and without IC. We anticipate that the findings of this study may lead to an objective noninvasive testing procedure for measuring the ischemic and exercise-induced changes in muscle oxygenation in the presence of claudication. If the testing of ischemic hypoxia continues to show consistency and accuracy in determining the disability of the subjects with IC, future studies can more effectively test modes of conservative management, such as cessation of smoking, alternative exercise regimens, weight loss, and alternative pharmacological agents.
Exercise↗
Use of (CA)n polymorphisms to determine the origin of blood cells after allogeneic canine marrow grafting.
We have used a polymerase chain reaction-based assay measuring polymorphic (CA)n repeats, a class of simple sequence repeats, to assess the success of allogeneic canine marrow transplants. Results were compared with those obtained with karyotype analysis of dividing cells in recipients that were sex mismatched with their marrow donors. Twenty recipients were conditioned for transplantation of genotypically DLA-identical littermate marrow by 450 cGy of total-body irradiation. In 2 recipients, results could not be compared, since either only cytogenetic or dinucleotide (CA)n marker data existed. Both dogs had autologous marrow recovery. In 15 of the remaining 18 recipients, complete agreement was found between the results obtained with dinucleotide (CA)n markers, cytogenetic studies, and granulocyte changes after transplantation. Seven of the 15 showed eventual autologous recovery, 6 displayed mixtures of host and donor cells, and 2 showed donor-type hematopoiesis. Two of the 18 dogs showed mixed chimerism with (CA)n markers and autologous recovery by cytogenetics, findings that may be related to differences in cells analyzed by the two techniques--i.e., all nucleated cells by (CA)n markers versus dividing cells by cytogenetics. In one additional recipient, results of marrow cytogenetics, granulocyte changes, and (CA)n markers were consistent with a successful allograft, while peripheral blood cytogenetics suggested autologous recovery, possibly the result of erroneous blood sampling. Polymerase chain reaction-based testing for dinucleotide repeat (CA)n polymorphisms, originally developed for genetic mapping in the dog, is useful and reliable when compared with cytogenetic studies, in assessing the success of allogeneic marrow transplants in dogs.
Animals↗