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Biomedical subjects

R G Hamilton

Publications and source records attributed to R G Hamilton.

At least 127 records · Page 7Linked to original sources
PubMed1984

Immunoradiometric assay for quantitation of Dirofilaria immitis antigen in dogs with heartworm infections.

An immunoradiometric assay (IRMA) was developed, optimized, and validated for detection of parasite-specific antigen in sera from hosts with filarial infections, using Dirofilaria immitis in dogs as a model. The precision, reproducibility, and parallelism of the IRMA were examined, using precision profile analysis. The IRMA had acceptable precision and reproducibility [less than 15% intra-assay coefficient of variation (CV)] over a working range of 10 to 2,000 ng of D immitis-antigen (AG)/ml. The IRMA parallelism (agreement between dilutions) was acceptable (less than 10% interdilutional CV) with laboratory-spiked D immitis AG sera containing no D immitis-antibody (AB). However, it was not acceptable (greater than 20% interdilutional CV) for analysis of sera from naturally infected dogs containing D immitis AB, probably due to dissociation of immune-complexed AG with increasing serum dilution. Nonparallelism limited the accuracy of binding data interpolation from the standard curve. Specificity of the IRMA was enhanced by preabsorption of the radiolabeled detection antibody with Toxocara canis AG before use. Varying amounts of D immitis AG (22 to 1,000 ng/ml) were detected in 42% (20/48) of microfilaremic dogs. The presence of AG-specific AB at concentrations as low as 1 microgram/ml reduced the ability of the IRMA to detect D immitis AG. Factors that influence the accuracy and sensitivity of immunoassays for circulating filarial antigens are discussed.

Animals↗
PubMed1984

[Automated recording of precision profiles for the radioimmunological determination of T4 using a pocket calculator of the HP-41 CV type].

Precision profiles as useful tool for assurance of assay quality in 10 independent T4RIAs have been automatically obtained using a small programmable pocket calculator HP-41 CV. For each T4 assay batch, the within-assay coefficient of variation varied from 7 to 11% in the hormone concentration range of 2 to 20 micrograms/dl. The difference in coefficient of variation for all the 10 successive assay batches of T4, never exceeded 1% in the same hormone concentrations regions. All the above findings demonstrate that the precision profile can be used as a powerful tool for assessing the assay quality and consistency of overall random error between successive assay batches.

Computers↗
PubMed1983

Dirofilaria immitis: performance and standardization of specific antibody immunoassays for filariasis.

The factors associated with the development, optimization, and validation of immunoassays for the detection of parasite-specific antibody in filariasis infections were investigated using the dog heartworm, Dirofilaria immitis as a model. We examined two assays, the Protein A solid-phase radioimmunoassay (SPRIA) and enzyme-linked immunosorbent assay (ELISA), for quantitation of specific antibody to the parasite in canine serum. Precision, reproducibility, and parallelism were examined using response-error relationships and precision profile analyses. A staphylococcal Protein A saturation analysis was applied to the standardization of IgG anti-parasite antibody reference sera in weight per volume units (microgram/ml). Using the mean minimal detectable dose + 3 SD and an intraassay precision profile less than 10% coefficient of variation (CV) as criteria for assay sensitivity, the SPRIA and ELISA displayed comparable positive thresholds of 1 microgram/ml IgG anti-parasite antibody/ml of serum. Both assays demonstrated good reproducibility (less than 15% interassay CV) and parallelism (less than 20% interdilutional CV) over their working ranges (SPRIA: 1-40 micrograms/ml; ELISA: 1-5 micrograms/ml). Specificity of each assay was enhanced by preadsorption of cross-reacting antibodies in canine serum (i.e., specific for Toxocara canis antigens) with solid-phase antigen prior to assay. Methods for comparing different immunoassay designs are considered in relation to the variables that influence the assays' performance characteristics.

Adsorption↗
PubMed1983

Assessment of venom-specific IgG antibody in patients treated for hymenoptera allergy.

The IgG antibody (Ab) response achieved with specific venom immunotherapy was explored in 32 patients with Hymenoptera hypersensitivity. Venom-specific IgG Ab was quantitated before and after 1 year of immunotherapy using two solid phase radioimmunoassay (SPRIA) methods. An agarose-based test using 125I-Staphylococcus aureus Protein A (SPRIA) was used to determine specific IgG for five Hymenoptera species: yellow jacket (YJ), honeybee (HB), yellow-faced hornet (YH), white-faced hornet (WFH), and Polistes (POL). A cellulose disk test using 125I-anti-IgG (IgG RAST) was available only for YJ and HB venoms. Acceptable agreement (90% concordance) was observed with IgG anti-HB levels measured in the two assays. For the YJ-IgG, however, 17/69 (25%) of sera positive in the SPRIA were negative in the IgG RAST, whereas the converse was not observed. This result suggests that the IgG RAST is insufficiently sensitive to detect YJ-IgG responses in all patients on maintenance level immunotherapy. Using the Protein A SPRIA, there was excellent agreement between the venom used for immunotherapy and the specificity of the IgG Ab response. In 31 patients treated with a total of 90 venom species, 90/90 venom IgG levels were increased or maintained at high pretreatment levels in response to immunotherapy. In the same patients venom IgG levels obtained for venom species not included in therapy were undetectable or declined in 55/60 cases; in 4 cases treatment with YJ venom stimulated a WFH and/or YH IgG response, the remaining case, YJ venom stimulated a small rise in POL IgG. These apparent discrepancies can be explained by variable cross-reactivity among vespid and POL venoms. Among 32 patients with a combined total of 87 positive venom skin tests, 1 year of specific immunotherapy resulted in greater than 5 micrograms/ml of venom-specific IgG in 61 instances. In 25 instances, the level of venom IgG was detectable but less than 5 micrograms/ml, and in 1 case venom IgG could not be detected. Based on recent analyses by Golden et al., some or all of these latter 26 cases may represent suboptimal therapy despite a standard immunotherapy regimen. We conclude that venom IgG measurements can provide a specific and quantitative assessment of the immunologic response to venom therapy, and that such assessment may be clinically useful in detecting instances of suboptimal immunotherapy.

Antibody Specificity↗
PubMed1982

Activated charcoal filter counting for radioiodine effluent concentration determination in protein iodinations.

Regulatory agencies have recently placed emphasis upon quantification of 125I released to the environment during protein iodinations at radioiodination facilities. This necessitates air sampling in order to determine the concentration of 125I in the effluent. Air sample trapping mechanisms generally employed are activated charcoal filters. Difficulty arises in quantifying the activity of 125I trapped because of the attenuation of the 125I decay photons by the charcoal. Evaluation of the activity incident upon commercially available filters using a scintillation detector and large detector source separation is considered here. It is demonstrated that the activity in the filter may be treated as an exponential distribution within an attentuating matrix. This treatment essentially adds a constant correction factor to the counting efficiency of a given geometry for a filter-affluent flow rate combination. Finally, it is shown that an approximation assuming a uniform distribution of activity produces a large error in correction factor to the counting efficiency for the filters examined.

Air Pollution, Radioactive↗
PubMed1981

The quantitation of parasite-specific human IgG and IgE in sera: evaluation of solid-phase RIA and ELISA methodology.

We have developed a non-competitive solid-phase radioimmunoassay (SPRIA) to quantitate both human IgE and IgG antibodies against soluble adult antigens of Brugia malayi (B.m.), a filarial parasite causing extensive infection throughout the tropics. Previously enzyme-linked immunosorbent assays (ELISA) had been used to detect microgram/ml levels of IgG anti-B.m., but IgE antibodies were difficult to detect in this system. Since the SPRIA successfully quantitates both IgG and IgE anti-B.m., we sought to examine the reasons for the SPRIA's apparent superiority in detecting IgE anti-B.m. by extracting specific IgG from sera with high levels of IgE and IgG anti-B.m. antibodies. IgE anti-B.m. was then quantitated in these sera using both the SPRIA and ELISA methods. Results indicate that IgG anti-B.m. does not interfere with detection of specific IgE antibody in the SPRIA but does interfere in the ELISA. While ELISA permits detection of IgE anti-B.m. in the absence of competing IgG anti-B.m., as levels of specific IgG increase, the IgE is no longer detectable. These differences between SPRIA and ELISA can be explained by the SPRIA's antigen excess conditions which assure that there are sufficient antigens both to detect all anti-B.m. antibodies present in the serum and to adequately represent all antigen specificities in the crude B.m. extract. Our findings commend the use of SPRIA methods over ELISA in assessment of B.m.-specific IgE antibody in filariasis and indicate a potential role for SPRIA methods in absolute quantitation of specific serum antibodies.

Animals↗
PubMed1981

Hyperlabile diabetes accompanied by insulin resistance.

It is generally held that high insulin antibody concentrations, by "buffering" abrupt swings in free insulin concentrations after injections of exogenous insulin, tend to stabilize blood glucose variations in diabetic patients. However, we encountered a patient with extremely labile diabetes coexisting with insulin resistance. This patient's injections were switched to pure porcine insulin from his usual mixed bovine/porcine insulin, in an effort to decrease his insulin requirement. This treatment was successful, and, as his insulin dosage decreased, his diabetic lability diminished substantially. His diabetes was eventually considered stabilized on about 22 units of porcine insulin daily. The serial decrease in his insulin antibody concentrations, monitored by use of solid-phase radioimmunoassay, paralleled the disappearance of his diabetic lability as well as the decrease in his insulin requirement.

Adult↗
PubMed1981

IgE responses in human filariasis. I. Quantitation of filaria-specific IgE.

We have developed a noncompetitive solid phase radioimmunoassay to quantitate human IgE antibodies against soluble adult antigens of Brugia malayi (B.m.), a filarial parasite, in sera of patients with various forms of clinical filariasis in Madras, India. A single reference serum was shown to contain 23 micrograms/ml of B.m.-specific IgE by depletion analysis and was used as a standard serum throughout the study. The levels of specific IgE ranged in the patients sera from 2 to 23,000 ng/ml. When these individuals were divided into clinical groups, the individuals with tropical pulmonary eosinophilia had the highest levels (mean = 8630 ng/ml) and were significantly higher than all the other groups (p less than 0.001). The lowest levels were seen in patients with circulating microfilariae (mean = 30.5 ng/ml). Patients exhibiting lymphatic obstruction (i.e., chronic pathology group) had levels slightly higher than microfilaremics (mean = 68 ng/ml) but were not significantly different (p less than 0.1). Surprisingly, individuals living in endemic areas but who had no clinical signs of filariasis also showed appreciable levels of B.m.-specific IgE (mean = 55 ng/ml). The B.m.-specific IgE represented 0.1 to 48% of the total IgE. High percentages of specific IgE may be responsible for evoking allergic symptomatology in patients with tropical pulmonary eosinophilia.

Adolescent↗
PubMed1980

Protein radioiodination in a radioassay laboratory: evaluation of commercial Na125I reagents and related biohazards.

Three commercial Na125I solutions (Amersham, New England Nuclear, and Union Carbide) have been examined with respect to multiple parameters affecting their use in the radioiodination of three representative peptides (insulin, growth hormone, and gastrin): % of radioiodine incorporation in protein; immunoreactivity and non-specific binding properties of the radiolabeled proteins; pH, volatility, and radionuclidic purity of radioiodine solutions; and vial construction with respect to multidose use. All three commercial Na125I produced radioiodinated proteins of good quality for use in radioligand assays. The radioiodines differed with respect to the amount of iodine released during initial vial opening as a consequence of different pH levels: 15 nCi/mCi (pH 12.5) to 1.0 microCi/mCi (pH 7.5). Two of the three products were shipped in vials with poor construction with respect to multidose use. Selection of a radioiodine was therefore reduced to the secondary considerations of iodine volatility and vial construction. The volatilized radioiodine observed during the spill of millicuries quantities of unbuffered pH 7.5 Na125I was 14 microcuries per millicurie within the first 30 minutes. One thickness of rubber gloves reduced potential skin contamination from an accidental spill to insignificant levels: 20-30 picocuries per microcurie. Common good housekeeping procedures: i.e. rubber gloves, laboratory coat and a fume hood were found to be sufficient protection to eliminate most radioiodine volatility and contamination hazards associated with protein radiolabeling procedures.

Binding Sites↗
PubMed1979

Solid phase radioimmunoassay for quantitation of antigen-specific IgG in human sera with 125I-protein A from Staphylococcus aureus.

Radiolabeled protein A from Staphylococcus aureus (Staph A) has been used to develop a solid phase, noncompetitive radioimmunoassay for quantitation of specific IgG antibody. The assay involves two incubations: First, agarose-insolubilized antigen is mixed with serum samples for 1 to 4 hr during which specific antibody is bound; second, after a washing procedure, the solid phase immune complexes are incubated for 4 to 18 hr with 125I-Staph A, during which the radiolabeled detection protein binds to the insolubilized specific IgG antibody. In a comparative study of the IgG antiphospholipase A antibody content of 23 human sera drawn from honeybee venom-sensitive patients, resulted of the Staph A assay correlated highly (r = 0.981, p less than 0.001, N = 23) with those obtained from a liquid phase, competitive radioimmunoprecipitation (double antibody) assay. The two assays demonstrated comparable precision, sensitivity, and reproducibility. In contrast, the use of 125I-Staph A in the solid phase radioimmunoassay was superior to 125I rabbit anti-human IgG because of lower negative serum (blank) values, shorter time required to reach equilibrium binding, and greater precision and reproducibility. In principle, the 125I Staph A assay may be applied ot IgG quantitation for crude allergen extracts as well as purified antigens. Furthermore, the sera of a number of mammalian species may be studied without further modification.

Animals↗
PubMed1977

Technetium-99m phytate as a bone-marrow imaging agent: biodistribution studies in animals: concise communication.

Technetium-99m phytate has been suggested as a bone-marrow imaging agent. This article compares the biodistribution of Tc-99m labeled "bone marrow" phytate, sulfur colloid, and diphosphonate in young rats and rabbits. Autoclaved bone marrow phytate revealed significant long-base depositon, but 96% of this activity was associated with compact bone and only 4% with bone marrow. This distribution is similar to that of diphosphonate, but significantly different from that of sulfur colloid. Technetium-99m-phytate is not recommended as a bone-marrow imaging agent.

Animals↗
PubMed1977

A comparative evaluation of techniques for rapid and efficient in vivo labeling of red cells with [99mTc] pertechnetate.

Red blood cells (RBCs) labeled in vivo with 99mTcO4- have recently been recommended for blood-pool imaging, but the optimum conditions for in vivo labeling of RBCs have not been clearly defined. We therefore evaluated several stannous-ion preparations and stannous-ion concentrations to determine which provided the best labeling. The effect of the time interval between the Sn(II) and 99mTcO4- injections and the effect of carrier technetium on labeling efficiency were also studied. Maximal in vivo labeling efficiency was obtained using an intravenous dose of 10 microgram Sn(II)/kg followed 5-30 min later by an injection of 99mTcO4-. Neither the chelated form of stannous ion used in these studies nor the amount of carrier present had a significant effect on labeling efficiency. The biologic half-time of Tc-99m RBCs labeled in vivo was similar to that of Tc-99m RBCs labeled in vitro. In vivo labeling is a rapid and efficient method for the preparation of Tc-99m RBCs.

Animals↗