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PubMedDate not supplied

Assessment of DNA damage in germ cells of male rabbits treated with isoniazid and procarbazine.

The 2 hydrazine derivatives isoniazid (INH) and procarbazine hydrochloride (P) were injected intravenously into rabbits. Radioactive thymidine was injected into both testicles. Rabbits were ejaculated repeatedly, sperms were counted and incorporation of [3H] thymidine into sperm head DNA was determined by liquid scintillation counting. In P-treated rabbits (5 and 50 mg/kg) radioactivity was significantly increased in sperms that were in late phases of spermatocyte and early phases of spermatid maturation at the time of treatment. This indicates that DNA repair synthesis, (unscheduled DNA synthesis, UDS) occurred following drug-induced DNA damage in these germ cells. Normal DNA synthesis in spermatogonia was inhibited by the high dose only. INH (50 and 125 mg/kg) did not cause UDS in spermatocytes and spermatids and did not affect normal DNA synthesis in spermatogonia. The results are in agreement with literature data indicating that P is a potent mutagen and carcinogen. INH, on the other hand, has weak mutagenic and carcinogenic activities that are most apparent in mice.

Animals↗
PubMedDate not supplied

Ageing and testicular function in Octodon degus.

The present study analyses cell loss and proliferation which account for the decrease in the number of germ cell populations in the senile male Octodon degus. This is a good model to study ageing in wild animals, since it has recently been incorporated as a laboratory animal but still has a high degree of genetic heterogeneity, thus representing a situation found in natural systems. The cell loss from pachytene spermatocytes to round spermatids is estimated by cell counts in the cross section of seminiferous tubules. DNA testicular synthesis is measured by scintillation counting and the index of labelling of spermatogonia by radioautography of testes comparing sexually mature young animals and senile animals. Other determinations in both groups are testis weight, thickness of the albuginea and tubular wall, daily sperm production, percentage of depleted seminiferous tubules and nuclear cell diameters of germ cells. The results suggest a decrease in the number of cell population in the senile animals resulting from an increase in physiological cell loss coupled with a decreased proliferative spermatogonial activity. There is also a decreased yield of meiosis in terms of round spermatid production. Lowered testosterone levels both in plasma and testicular parenchymal fluid are found in senile animals. All these senescent changes reflect an altered remodelling activity of the seminiferous epithelium and presumably also of Leydig cells.

Aging↗
PubMedDate not supplied

Receptor binding assay for paralytic shellfish poisoning toxins: optimization and interlaboratory comparison.

A receptor binding assay (RBA) for detection of paralytic shellfish poisoning (PSP) toxins was formatted for use in a high throughput detection system using microplate scintillation counting. The RBA technology was transferred from the National Ocean Service, which uses a Wallac TriLux 1450 MicroBeta microplate scintillation counter, to the California Department of Health Services, which uses a Packard TopCount scintillation counter. Due to differences in the detector arrangement between these 2 counters, markedly different counting efficiencies were exhibited, requiring optimization of the RBA protocol for the TopCount instrument. Precision, accuracy, and sensitivity [limit of detection = 0.2 microg saxitoxin (STX) equiv/100 g shellfish tissue] of the modified protocol were equivalent to those of the original protocol. The RBA robustness and adaptability were demonstrated by an interlaboratory study, in which STX concentrations in shellfish generated by the TopCount were consistent with MicroBeta-derived values. Comparison of STX reference standards obtained from the U.S. Food and Drug Administration and the National Research Council, Canada, showed no observable differences. This study confirms the RBA's value as a rapid, high throughput screen prior to testing by the conventional mouse bioassay (MBA) and its suitability for providing an early warning of increasing PSP toxicity when toxin levels are below the MBA limit of detection.

Amphibian Proteins↗
PubMed1987

Comparison of cyclosporine determinations in whole blood by three different methods. HPLC, 125I RIA and 3H RIA.

The authors have analyzed and compared the cyclosporine concentrations in whole blood specimens from pediatric renal transplant patients using three different methods: high-performance liquid chromatography (HPLC) (5u C18 reverse-phase column), 3H radioimmunoassay (RIA) (Sandoz, E. Hanover, NJ), and 125I RIA (substituted 3H-tracer in Sandoz Kit with 125I tracer made by Immuno Nuclear Corporation, Stillwater, MN). Results obtained by the 125I RIA correlated well with results obtained by the 3H RIA. Both RIA methods had similar correlation with the HPLC method. The 125I RIA method showed higher sensitivity and greater precision than the 3H RIA method. The authors conclude that the 125I RIA method can be used for cyclosporine determination in whole blood specimens. The use of the 125I RIA provides a simple and rapid method with higher counting efficiency and less background quenching than the 3H RIA method, which requires cumbersome liquid scintillation counting procedures.

Chromatography, High Pressure Liquid↗
PubMedDate not supplied

[Radiologic follow-up and prognosis of efficiency of systemic Sr-89 Cl-2 therapy of bone metastases in prostate cancer].

We evaluated possibilities of bone scintigraphy with 99mTc-methylendiphosphonate (99mTc-MDP) and magnetic resonance imaging (MRI) in follow-up and prediction of effect in patients with extensive bone metastatic disease treated with betha-emitter 89SrCl2. 24 patients with prostate cancer and extensive metastatic involvement of skeleton were referred for the study. 89SrCl2 was injected as single injection of 150 MBq (4 mCi), in eighteen from Amersham plc., England, as Metastron, in six--from Medradiopreparat, Russia). In all patients bone scintigraphy with 99mTc-MDP and MRI study of metastatic regions were performed before and in 3 months after 89SrCl2 injection. Patients treated with Metsatron were also studied in 6 months after injection. Quantitative analysis of data comprised count and anatomic dimensions of metastatic areas and calculation of indices [metastasis/intact bone] both for scintillation count of 99mTc-MDP bone scans and signal intensity of T1-weighted MRI scan. Henceforth, we conclude the data of bone scanning with 99mTc-MDP and of MRI give evidencies for significant regress of bone metastases in patients treated with 89SrCl2 besides symptomatic suppression of pain syndrome. 99mTc-MDP bone scanning is also of predictive value for the prognosis of therapeutic effect of systemic radiotherapy with 89SrCl2 in prostate cancer.

Aged↗
PubMed1986

Measurement of Glucose Utilization by Pseudomonas fluorescens That Are Free-Living and That Are Attached to Surfaces.

The assimilation and respiration of glucose by attached and free-living Pseudomonas fluorescens were compared. The attachment surfaces were polyvinylidene fluoride, polyethylene, and glass. Specific uptake of [C]glucose was determined after bacterial biomass was measured by (i) microscopic counts or (ii) prelabeling of cells by providing [H]leucine as substrate, followed by dual-labeling scintillation counting. The glucose concentration was 1.4, 3.5, 5.5, 7.6, or 9.7 muM. Glucose assimilation by cells which became detached from the surfaces during incubation with glucose was also measured after the detached cells were collected by filtration. The composition of the substratum had no effect on the amount of glucose assimilated by attached cells. Glucose assimilation by attached cells exceeded that by free-living cells by a factor of between 2 and 5 or more, and respiration of glucose by surface-associated cells was greater than that by free-living bacteria. Glucose assimilation by detached cells was greater than that by attached bacteria. Measurements of biomass by microscopic counts gave more consistent results that those obtained with dual-labeling, but in general, results obtained by both methods were corroborative.

Journal Article↗
PubMed2001

Simultaneous determination of 226Ra and 210Pb in groundwater and soil samples by using the liquid scintillation counter-suspension gel method.

A method for the simultaneous determination of 226Ra and 210Pb in groundwater and soil samples by liquid scintillation counting was developed. Radium and lead were separated together from the samples as Ba(Ra) x PbSO4 co-precipitate, which was centrifuged and dissolved with 0.1 M EDTA solution (pH 9.0). Radium was separated as Ba(Ra)SO4 co-precipitate by adding ammonium sulfate and adjusting the pH of the solution to 4.2. Lead remaining in the solution was separated as PbSO4 precipitate by adding 9 M sulfuric acid. These Ba(Ra)SO4 and PbSO4 precipitates were purified with EDTA solution and used for measurement. To save time and to make counting samples simpler, direct counting of Ba(Ra)SO4 and PbSO4 precipitates instead of the phosphoric acid fusion method was attempted. Ba(Ra)SO4 and PbSO4 precipitates were suspended in the scintillation gel, and measured. Two liquid scintillation cocktails, Instagel XF and UltimaGold AB were used to prepare the counting samples. A mixture of water (40%), Instagel XF (40%) and UltimaGold AB (20%) formed a stable gel. Activities of 226Ra and 210Pb were calculated from the alpha spectrum of Ba(Ra)SO4 and beta spectrum of PbSO4, respectively. The long-term stability of the suspension gel was good. The analytical results of 226Ra and 210Pb in spiked groundwater samples were in good agreement with the known concentrations of 226Ra and 210Pb. The analytical values of 226Ra and 210Pb in the soil reference samples were within 11.5 and 1.6% of the relative error from the reference values, respectively.

Journal Article↗
PubMed2002

An evaluation of two sample preparation methods for measuring 3H and 14C in incinerator ash and spent lime.

This paper compares the performance of two methodologies, oxidation and gel suspension, for measuring the 3H and 14C specific activity in incinerator ash and lime. The two methods were judged on the basis of the count time needed to achieve a specified minimum detectable concentration. Both methods involve pretreatment of the sample matrix followed by liquid scintillation counting. Both methods perform adequately for measuring 3H in spent lime, but only oxidation delivers adequate sensitivity for the more demanding 14C case. In addition, the gel suspension method fails to detect over 95% of the 3H seen by the oxidation method in incinerator ash.

Carbon Radioisotopes↗
PubMed2002

Sample preparation for quantitation of tritium by accelerator mass spectrometry.

The capability to prepare samples accurately and reproducibly for analysis of tritium (3H) content by accelerator mass spectrometry (AMS) greatly facilitates isotopic tracer studies in which attomole levels of 3H can be measured in milligram-sized samples. A method has been developed to convert the hydrogen of organic samples to a solid, titanium hydride, which can be analyzed by AMS. Using a two-step process, the sample is first oxidized to carbon dioxide and water. In the second step, the water is transferred within a heated manifold into a quartz tube, reduced to hydrogen gas using zinc, and reacted with titanium powder. The 3H/1H ratio of the titanium hydride is measured by AMS and normalized to standards whose ratios were determined by decay counting to calculate the amount of 3H in the original sample. Water, organic compounds, and biological samples with 3H activities measured by liquid scintillation counting were utilized to develop and validate the method. The 3H/1H ratios were quantified in samples that spanned 5 orders of magnitude, from 10(-10) to 10(-15), with a detection limit of 3.0 x 10(-15), which is equivalent to 0.02 dpm tritium/mg of material. Samples smaller than 2 mg were analyzed following addition of 2 mg of a tritium-free-hydrogen carrier. Preparation of organic standards containing both 14C and 3H in 2-mg organic samples demonstrated that this sample preparation methodology can also be applied to quantify both of these isotopes from a single sample.

DNA↗
PubMed1996

A new method for quantitative determination of tritium-labeled nucleoside kinase products adsorbed on DEAE-cellulose.

The counts of tritiated compounds--adsorbed to paper disks, paper chromatograms, electrophoretograms or TLC plates--can be strongly affected by extended self-absorption of tritium beta-particles on the matrix, due to their low energy. Therefore fully quantitative results can be obtained only by elution of the substances or decomposition of the matrix and subsequent counting in homogeneous solution. In this study we describe a new method for fast and proper decomposition of cellulose matrices by cellulase digestion prior to scintillation counting. This new approach yields up to 98% recovery. For method validation recombinant herpes simplex type 1 thymidine kinase was characterised kinetically. The Km of 0.2 microM remained the same as expected but Vmax was considerably higher yielding 1050 pmol/microgram/min.

Adsorption↗
PubMed2000

Measurements of linear absorption coefficients of liquid scintillators using synchrotron radiation

The use of liquid scintillation counting as a fundamental radionuclide standardisation method requires a correct description of the physical phenomena induced by the interaction of the ionising radiation with the liquid scintillator. In particular the standardisation of radionuclides decaying by electron capture as 55Fe and other electron-capture nuclides, requires the knowledge of total linear absorption coefficients of the liquid scintillator used for absolute activity measurement. Total linear absorption coefficient measurements have been undertaken using synchrotron radiation. Linear absorption coefficients of two widely used commercial liquid scintillators were measured in the 5.5-23 keV energy range. Small discrepancies were noted between theoretical and measured values of the Ultima-Gold mass attenuation coefficients for low energy photons.

Journal Article↗
PubMed1996

The effect of lovastatin on [3H]thymidine uptake in HTC-4 and LLC-L1 tumor cells.

Lovastatin, a 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor, has been shown to inhibit the in vitro and in vivo growth of a number of different cell lines. However, the mechanism by which lovastatin exerts its effect is not clear. In this experiment, we investigated the effect of lovastatin on the incorporation of [3H]thymidine by Hepatoma Tissue Culture-4 (HTC-4) and Lewis Lung Carcinoma L-1 (LLC-L1) tumor cells. Tumor cells were grown under standard conditions and treated with four different concentrations of lovastatin. Cell growth was evaluated by daily hemacytometer cell counts. On Day 4, the plates were pulsed with 10 microCi [3H]thymidine. After 24 hr, the plates were harvested and [3H]thymidine incorporation was measured by scintillation counting. Lovastatin inhibited both HTC-4 and LLC-L1 cell growth in a dose-dependent manner. At the highest lovastatin dose, both LLC-L1 and HTC-4 cell growth was slowed to less than 15% of control. Remarkably, however, both cell lines showed a paradoxical, dose related, increase in [3H]thymidine uptake. Cell cycle analysis using flow cytometry was performed on Day 5 in the LLC-L1 cell line. As the lovastatin concentration increased, a lower percentage of cells was found in the G1 phase of the cell cycle and a higher percentage of cells was found in the S and G2 phases. These findings suggest that these tumor cells are undergoing brisk DNA repair or that lovastatin is more effectively blocking cell division than cellular DNA replication.

Animals↗
PubMed2000

Measurement of 49V half-life

The accepted value for the 49V half-life is either 330+/-15 days, based on a few old measurements with large dispersion, or 338+/-5 days, from a more recent measurement. Methods of study have included proportional counting and Nal X-ray detection. A new direct measurement of the 49V half-life has been carried out for over 2.5 half lives using liquid scintillation counting, as an additional contribution due to the scarce measurements of this radionuclide. A half-life value of 328.6+/-3.3 days was determined on the basis of an overall uncertainty analysis of 0.99%.

Journal Article↗
PubMed2001

Effects of human tissue plasminogen gene transfer on allograft coronary atherosclerosis.

BACKGROUND: Transplant coronary atherosclerosis is a major limiting factor to successful long-term cardiac transplantation. The depletion of tissue plasminogen activator (tPA) in the arteriolar smooth muscle cells has been associated with a higher incidence of accelerated graft atherosclerosis. In vivo overexpression of tPA may inhibit accelerated graft atherosclerosis and improve the long-term results of heart transplantation. We evaluated the feasibility, distribution, and effects of intracoronary transfer of the human tPA (htPA) gene in a rabbit heterotopic cardiac transplant model, using a novel cationic liposome compound designed for improved delivery to vascular endothelium. METHODS: Human tPA cDNA under the control of the SV40 promoter (100 microg) was complexed with the novel cationic liposome (+/-)-N-(3-aminopropyl)-N,N-dimethyl-2,3-bis(dodecyloxy)-1-propanaminium bromide (GAP: DLRIE) (50 microg), and delivered ex vivo to the donor heart by slow intracoronary infusion. Control hearts received an "empty" liposome preparation. Grafts were then implanted into recipient rabbits in the heterotopic cervical position. For the analysis of gene expression, beating donor hearts were collected at 4 days. To examine the effects of htPA expression on graft atherosclerosis, animals received a 0.5% cholesterol diet for 30 days posttransplant, as well as 10 mg/kg cyclosporine A daily. Beating hearts were collected at 30 days posttransplant and analyzed for the development of transplant atherosclerosis by image analysis. RESULTS: Northern blot analysis for the htPA messenger RNA (mRNA) transcripts showed significantly higher counts in hearts receiving the htPA gene as compared to controls. The distribution of these transcripts favored the left ventricle (LV) and septal regions over the right ventricle (RV). Scintillation analysis of specimens stained by immunoflourescence showed expression of htPA throughout the perivascular myocardium that was significantly higher in grafts transduced with the htPA gene than in control or native hearts. Expression in the vascular wall was also significantly enhanced. Scintillation counts per x 200 field were 262 +/- 145 in htPA-transduced hearts and 20 +/- 27 in controls (p = 0.001), and mean luminescence was 83.7 +/- 12.5 in htPA-transduced hearts and 62.9 +/- 12.8 in controls (p = 0.01). Intimal hyperplasia was assessed by mean percent luminal stenosis in small- and medium-sized arteries and was 31.12 +/- 23.5% in htPA-transduced hearts and 86.59 +/- 17.5% in control hearts (p < 0.0001). These results demonstrate that expression of the htPA gene can be induced by ex vivo intracoronary gene transfer at the time of allograft preservation. Liposome-mediated delivery of the htPA gene at the time of transplantation results in significant early transgene expression, and significantly inhibits the development of graft coronary atherosclerosis.

Animals↗
PubMed1995

Determination of waterborne 222Rn concentrations using AC canisters.

A method for measuring 222Rn concentration in water using charcoal canisters is presented. 222Rn is transferred within a few minutes from water contained in a 0.720 L bottle to a charcoal canister using a portable degassing unit. In the laboratory, gamma counting is performed at least 10 h after sampling to determine waterborne 222Rn concentration. The results obtained with charcoal canisters are compared to measurements made with Marinelli beakers calibrated in a comparison with liquid scintillation counting. The efficiency of transferring dissolved 222Rn in water to activated charcoal is 99%, based on measurements made using Marinelli beakers. The lower limit of detection at the 95% confidence level is approximately 1 kBq m-3 for a 15 min gamma count. The system was used to measure radon concentration in mineral groundwater near Rome.

Radon↗
PubMed1986

Age-dependent variation of lymphocyte function in the postoperative child.

Circulating lymphocyte profiles and reactivity normally vary with age. Operation results in depression of both lymphocyte counts and blastogenesis but the relationship of age to these alterations has not been previously evaluated in the pediatric surgical patient. This report analyzes the relationship of age to lymphocyte alteration in the postoperative child. Thirty-five healthy children (age range 1 mo to 12 yr), admitted for elective herniorrhaphy, underwent perioperative lymphocyte assay. Anesthesia consisted of halothane and NO2, and operative time averaged 54 minutes. Three milliliters of heparinized whole blood was obtained at induction of anesthesia and at 2 hours postoperatively. Lymphocytes were separated by Histopaque gradient and assayed for absolute count, total T-cells, and total B-cells. Lymphocyte reactivity was determined by 3H-Thymidine incorporation during incubation with the mitogens phytohemagglutinin (PHA), pokeweed (PWM), and/or concanavalin A (Con A) and results expressed as the logarithm of scintillation counts per minute. Differences in preoperative and postoperative values were analyzed for significance by paired T-test. The same differences were evaluated for relationship to age by regression analysis. Operation resulted in significant decreases in all lymphocyte counts (absolute, T- and B-cells), and the operative-induced alteration in both absolute and total B-cell counts were significantly correlated with age (P less than .03, P less than .007, respectively). Similarly, operation resulted in significant depression of lymphocyte reactivity and alteration in B-cell response (PWM) was significantly correlated with age (P less than .008). In all instances of significant correlation, lymphocyte alterations were inversely related to age.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗