Drug-exposed infants and children: living with a lethal legacy.
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Child↗
Biomedical subjects
T J Green
Publications and source records attributed to T J Green.
Identification of common surface antigens among Babesia bigemina isolates by using monoclonal antibodies.
A group of monoclonal antibodies (MAb) was used to immunochemically characterize Babesia bigemina surface components. Two surface reactive MAb by an IFAT test were subsequently shown to bind to epitopes on the external surface of the parasite plasma membrane as evidenced by immunoelectron microscopy. Parasite components with relative sizes of 68, 62, 60, 58, 56, 54, 51, 49, 48, 47, 43, 36 kDa were identified with the group of MAb in at least 6 geographically different B. bigemina isolates. The antigenic components were demonstrated to be species-specific and apparently predominant. Both polyclonal antibodies (immune bovine serum) and highly diluted MAb specifically reacted with such bands producing a strong signal in Western blots.
Animals↗
Monoclonal IgM rheumatoid factor-like anti-globulins enhance the inhibitory effects of Plasmodium falciparum-specific monoclonal antibodies in vitro.
Monoclonal IgM rheumatoid factor-like anti-globulins were produced by in vitro stimulation of naive BALB/c spleen cells with lipopolysaccharide, and by hyperimmunization of mice with merozoites of Plasmodium falciparum, followed by fusion of the spleen cells to mouse myelomas. In vitro, these anti-globulins augmented the inhibitory effects of P. falciparum-specific polyclonal mouse sera and monoclonal IgG1 and IgG2b antibodies by binding to Fc fragments of IgG molecules attached to blood-stage parasites. In some instances, the presence of anti-globulins correlated with an increase in the number of schizonts which failed to disperse merozoites. In other cases, parasitaemia remained low in the absence of the schizont inhibition phenomenon, suggesting that anti-globulins contribute to host cell protection not only by agglutinating merozoites, but also by increasing the density of the antibody coat surrounding the parasites, thus interfering with parasite receptor-erythrocyte ligand interactions. The anti-globulins were not inhibitory when added to parasite cultures containing IgG not specific for P. falciparum. These results may help explain the function of IgM anti-globulins found at elevated serum levels in some patients with malaria or other chronic infectious diseases.
Animals↗
The oxygen dependence of mitochondrial oxidative phosphorylation measured by a new optical method for measuring oxygen concentration.
Oxygen-dependent quenching of phosphorescence has been used to measure the dependence of mitochondrial oxidative phosphorylation on oxygen concentration in suspensions of isolated rat liver mitochondria. An instrument has been designed which simultaneously monitors the phosphorescence lifetime of a fluorophor and the reduction of cytochrome c by dual wavelength spectrophotometry. The phosphorescence lifetime method gives very rapid (less than 100 ms) measure of the oxygen concentration (Vanderkooi, J. M., Maniara, G., Green, T. J., and Wilson, D. F. (1987) J. Biol. Chem. 262, 5476-5482) from concentrations characteristic of air-saturated media to as low as 2 x 10(-8) M. The results may be summarized as follows. For well coupled rat liver mitochondria at pH 7.0 and in the presence of ATP, as the oxygen concentration was lowered, increased cytochrome c reduction was observed to begin at oxygen concentrations greater than 20 microM. For mitochondria in the presence of uncoupler, cytochrome c reduction began at oxygen concentrations less than 1.0 microM. The oxygen dependence of reduction of cytochrome c in well coupled mitochondria treated with ATP was strongly dependent on the pH of the suspending medium. Reduction of cytochrome c began at higher oxygen concentrations as the pH was made more alkaline. The oxygen concentration for half-maximal respiratory rates was much larger for well coupled mitochondria treated with ATP (approximately 0.7 microM) than for mitochondria treated with uncoupler (less than 0.1 microM). It is concluded that the oxygen dependence of mitochondrial oxidative phosphorylation is such that mitochondria could function in their proposed role of tissue oxygen sensors for regulation of such diverse functions as local blood flow and electrical activity in the carotid body.
Adenosine Triphosphate↗
Phosphorimeters for analysis of decay profiles and real time monitoring of exponential decay and oxygen concentrations.
A phosphorimeter which can be assembled at low cost from mainly commercially available components and which has better time resolution, data acquisition rate, sensitivity, and flexibility than commercially available instruments is described. As a phosphorescence analyzer the instrument can measure phosphorescence lifetimes ranging from approximately 30 microseconds to seconds from samples with variable intensity, excitation, and emission spectra and which may follow complex decay behavior. Configured as a phosphorescence monitor it is designed for fast, repetitive calculation of phosphorescence lifetime, assuming single-exponential decay, and can be used to calculate oxygen concentration in biological samples in real time.
Luminescent Measurements↗
An optical method for measurement of dioxygen concentration based upon quenching of phosphorescence.
An optical method for measuring oxygen concentrations in aqueous solutions is described. This method is based upon the oxygen-dependent quenching of phosphorescence. Phosphorescence excitation and emission spectra and lifetimes of some of the probe molecules suitable for measurement of oxygen in aqueous solutions are given. The probes include fluorescein derivatives, 4'5'-diiodofluorescein, eosin Y, 5(and 6)-carboxyeosin, erythrosin, and 5(and 6)-carboxyerythrosin as well as the Zn(II), Y(III), Sn(IV), Lu(III), and Pd(II) derivatives of meso-tetra-(4-sulfonatophenyl)-porphine, meso-tetra-(N-methyl-4-pyridyl)-porphine and coproporphyrin. The phosphorescence lifetimes of the given probes were found to depend upon the oxygen concentration by a simple Stern-Volmer relationship with a quenching constant of approximately 10(9) M-1 S-1. Binding of the molecules to bovine serum albumin decreased the quenching constant for oxygen by approximately an order of magnitude and also inhibited probe self-quenching, indicating that at the protein binding site the probes are somewhat protected from collision with quenchers. The use of this optical method for measuring oxygen is demonstrated for reactions catalyzed by glucose oxidase and by cytochrome c oxidase. It is shown that, using this method oxygen concentrations can be measured from approximately 250 microM (air saturation) down to the nanomolar range.
Animals↗
A versatile and sensitive method for measuring oxygen.
Oxygen dependence of the lifetime of the excited triplet state of phosphorescent molecules can be used to measure the oxygen concentration in aqueous media. These measurements are insensitive to much of the optical interference that limits the usefulness of measurements based on the oxygen dependent quenching of luminescence intensity. The measurements also extend to significantly lower oxygen concentrations than are normally attainable using oxygen electrodes. The phosphorescence lifetimes can be accurately measured from a few microseconds to seconds, permitting a wide dynamic range of oxygen concentration measurements. With currently available probes, for example, it is possible to make continuous measurement of oxygen concentrations from 10(-4) M to 10(-8) M in a single experiment.
Animals↗
A sensitive urea-silver stain method for detecting trace quantities of separated proteins in polyacrylamide gels.
An ultrasensitive method using a urea-silver staining procedure to detect trace quantities of proteins in polyacrylamide gels (PAGE) is described. This technique is sensitive enough to detect picogram quantities of proteins resolved on sodium dodecyl sulfate-polyacrylamide gels. The major advantages of our method are that it provides a clear background and it is more sensitive than other techniques allowing it to substitute for radioisotopic techniques in some cases.
Electrophoresis, Polyacrylamide Gel↗
Reference values for peripheral blood lymphocytes from Aotus lemurinus ssp. griseimembra (owl monkey).
The reactivities of several monoclonal antibodies that define human lymphocyte cell-surface antigens have been tested with peripheral blood lymphocytes of Aotus lemurinus ssp. griseimembra. Based on reactivity patterns in humans, reactive MoAb were identified that mark pan-T, helper/inducer, suppressor/cytotoxic, pan-B, and natural killer cells. Reference values of these subsets in Aotus are presented. These MoAb should provide a useful tool for further phenotypic and functional dissection of the immune system in this simian model of human disease.
Animals↗
Erythrocytes and erythrocyte morphologies of healthy and colony-born owl monkeys (Aotus lemurinus griseimembra).
Inadequate availability of hematological reference data seriously restricts optimal utilization of the owl monkey (Aotus lemurinus griseimembra) as an experimental model. The current study investigated erythrocytic morphology in peripheral blood of healthy, colony-born owl monkeys. The blood of the subjects contained discoid erythrocytes, poikilocytes, and showed considerable anisocytosis. Also observed were nucleated erythrocytes, erythrocytes with Howell-Jolly bodies, and reticulocyte types I, II, and III. Heinz bodies were not detected.
Animals↗
Organization of Plasmodium falciparum genome: II. Sequence analysis of falci element.
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Plasmodium falciparum↗
Specific detection of Plasmodium falciparum malaria by a molecularly cloned DNA probe.
A highly repeated DNA sequence from Plasmodium falciparum was cloned and used as a probe in molecular hybridization to detect malaria. Our results indicate that the probe is specific to P. falciparum but not to other species of Plasmodium and is extremely sensitive. As little as a 20 pg parasite DNA, which is equivalent to about 1000 parasites can be detected. The cloned DNA can be used as a diagnostic tool to follow the course of infection of falciparum malaria.
Animals↗
Organization of Plasmodium falciparum genome: 1. Evidence for a highly repeated DNA sequence.
Plasmodium falciparum DNA, isolated from the merozoite stage, was cleaved with HindIII and cloned in pBR322 and lambda L47.1 vectors. Plasmid clones containing 13.4, 7.0, 4.3, 4.1 and 1.5 kb inserts were characterized in some detail. The inserts contain several repeating units of smaller size. Nucleic acid hybridization studies showed that the repeat element is present in the Plasmodium DNA at a very high copy number and appears to be distributed widely throughout the genome.
Animals↗
Cloning and characterization of Plasmodium falciparum FCR-3/FMG strain.
The FCR-3/FMG African strain of Plasmodium falciparum was cloned by the limiting dilution technique in in vitro culture to obtain parasite populations derived from a single cell. The basic in vitro culture technique used was that of Trager and Jensen, performed in microtiter culture plates. The cloning sequence was repeated serially three times, and three parasite clones with a higher than 99% probability of single cell derivation were isolated. Two of these clones were determined by electron microscopic examination to possess the K+ and one the K- trait. The clones were found to be equally sensitive to chloroquine in vitro but varied in their in vivo pathogenicity for Aotus, the K- clone being non-pathogenic.
Animals↗
In vitro cultivation of Babesia bigemina.
A strain of Babesia bigemina was isolated from an infected calf and propagated in vitro. Culture conditions included washing of infected and normal bovine erythrocytes in a special solution, and the use of a 5% to 10% (v/v) erythrocyte suspension in medium 199 (with 20% to 50% fresh normal bovine serum) at a depth of 4 mm in a 5% CO2, 2% O2, 93% N2 atmosphere. After 36 days in vitro and 9 subcultures, the cultured organism was inoculated into a susceptible calf. This calf developed clinical signs of disease and recovered when treated with 1% trypan blue solution. The strain was also reisolated from the second calf. The original isolate had been maintained in continuous in vitro cultivation for more than 99 days.
Animals↗
Effect of secretin on histamine-induced duodenal ulceration in guinea-pigs.
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Animals↗
In vitro propagation of Plasmodium falciparum for merozoite antigens.
A system with continuous flow of medium that uses disposable tissue culture flasks for the in vitro cultivation of P. falciparum is described. This scaled-up modification of the Trager and Jensen method gives merozoite yields sufficient for experimental vaccine studies in animals. A method of increasing the yield of merozoites by decreasing the pH of the medium is presented.
Animals↗
Demonstration of the role of cytophilic antibody in resistance to malaria parasites (Plasmodium berghei) in rats.
This paper reports the results of a study of the nature of the immune response against Plasmodium berghei parasites by inbred rats. A macrophage-cytophilic antibody specific for malarial antigens was identified and characterized. Detection of the antibody on the macrophage surface was accomplished by the parasite adherence tests and by the indirect fluorescent antibody technique. Isolation and purification of the macrophage-cytophilic and opsonic antibodies from hyperimmune rat serum was accomplished by QAE-Sephadez A-50 elution chromatography, and of the macrophage-cytophilic antibody by adsorption with and elution from syngeneic macrophages as well. Characterization of the cytophilic antibody as immunoglobulin G1 was done by immunoelectrophoresis and by Ouchterlony-type double diffusion in gel. Passive protection tests in weanling inbred rats have demonstrated that the opsonizing antibody conferred some protection against P. berghei. The macrophage-cytophilic antibody, on the other hand, was not protective alone but acted synergistically with the opsonizing antibody.
Animals↗