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PubMed1983

Simultaneous determination of total cholesterol concentration and radioactivity in plasma.

Total plasma cholesterol concentration and radioactivity were measured simultaneously using a gas chromatograph equipped with a flame ionization detector and an effluent splitter. More than 99% of the recovered radioactivity was in the cholesterol peak. Specific activities were highly correlated with the amounts of labeled cholesterol present in plasma. The recovery of label was quantitative over a wide range of carrier cholesterol concentrations. The method is highly reproducible, accurate, rapid and specific.

Acetates↗
PubMed1981

A method for rapid determination of IgG containing circulating immune complexes using polyethyleneglycol and radioactively labeled protein A.

A simple method for rapid determination of IgG containing circulating immune complexes by commercially available reagents was developed. In this method, serum is incubated with 2.5% polyethyleneglycol. After washing, the precipitate is incubated with radioactively labeled protein A, which binds to IgG in the immune complex. After a further washing, the radioactivity bound is measured. Artificially formed complexes of heat-aggregated human IgG are diluted and a reference curve is constructed. This method is compared to the solid phase C1q-binding method. Our method is slightly more sensitive than the C1q-binding method.

Adolescent↗
PubMed1983

Extension of the ELISA method to the measurement of the specific radioactivity of viruses in crude cellular extracts.

The double-antibody sandwich method of ELISA, which allows accurate quantitative determination of plant viruses, was extended to a radiochemical procedure which permits direct measurement of the specific radioactivity of virus labelled in vivo and present in very crude plant homogenates. Evidence is presented showing that 20 to 50% of the virus introduced in the polystyrene wells during the antigen incubation step could be trapped in the sandwich. The percentage of virus bound increased with the concentration of the coating antibody and was almost proportional to the concentration of the antigen and to the incubation time of the antigens. Complete dissociation of the double-antibody sandwich was achieved by incubation with 0.2 M KOH or NaOH (pH 13.3), and the label carried by the virus was measured by scintillation counting of the solubilization fluid. The ratio infected/healthy was much higher for the radiochemical procedure than for the immunosorbent assay itself since binding of the virus to the coating antibody was not accompanied by any nonspecific trapping of radioactive contaminants in the double-antibody sandwich. The procedure was highly sensitive since the background corresponded to the scintillation counting background. The detection of label carried by tobacco mosaic virus was possible when the tobacco samples contained at least 5 ng of virus carrying a label as low as 40 dpm 3H or 20 dpm 14C.

Enzyme-Linked Immunosorbent Assay↗
PubMed1985

A new physical assay method for tobacco mosaic virus using a radioactive virus recovery standard and the first derivative of the ultraviolet absorption spectrum.

A new physical assay method for tobacco mosaic virus is described which incorporates two improvements on previous procedures. Losses of up to 75% of virus during extraction are corrected by adding a trace of radioactive virus to leaf samples before homogenization, and determining percentage recovery of radioactivity in final virus preparations. Estimation of virus concentration in partially purified preparations is from the first derivative (dA/d lambda) of the ultraviolet absorption spectrum, using a pronounced signal from the tryptophan fine-structure absorption band at 285-293 nm. This method is highly insensitive to ultraviolet-absorbing contaminants, which cause errors and increase variation between replicates, when estimation of virus concentration is by normal measurement of ultraviolet absorption (A260). The method can be applied to at least some other viruses.

RNA, Viral↗
PubMed1991

Comparison of four non-radioactive and 35S-based methods for the detection of human papillomavirus DNA by in situ hybridization.

Human papillomavirus DNA was detected in 40 condylomatous lesions of various sites (vulva, cervix, larynx, penis and anus) by in situ hybridization using 35S-labelled probes and four non-radioactive probes to compare the various sensitivities of these techniques on the same material (formalin-fixed and paraffin-embedded sections). Radioactive probes yielded 28 positive results out of 40 (70%). Sulphonated probes (HybriCyte kit) also gave 28 positive results with a fine pattern of hybridization grains and equal sensitivity to 35S-labelled probes. Biotinylated and digoxigenin-labelled probes gave analogous results (25 positive reactions with the PathoGene kit, 26 with the Viratype kit, and 25 with digoxigenin-labelled probes) but are slightly less sensitive than radiolabelled and sulphonated probes especially when the signal is weak.

Adolescent↗
PubMed1995

Value of a new rapid non-radioactive sequencing method for analysis of the cytomegalovirus UL97 gene in ganciclovir-resistant strains.

Various DNA changes located within a restricted region of the UL97 open reading frame were shown to be associated with the resistance of cytomegalovirus strains to ganciclovir (GCV). In order to analyse this UL97 region in sensitive and GCV-resistant strains, a non-radioactive sequencing assay (Promega, Madison, WI, USA) which combines the dideoxy visualisation by silver-staining of the gel was used. Using this assay, polymerase chain reaction products from results were obtained within 1 day. Point mutations modifying the amino acid sequence of the putative UL97 catalytic site were detected in three isolates. These led to an alanine to valine substitution in residue 594 in one strain with reduced GCV sensitivity, and to a cysteine to glycine substitution in residue 592 in two GCV-resistant isolates. These mutations were different from the DNA changes previously mapped in GCV-resistant laboratory or field strains. No amino acid substitution in the UL97 catalytic site was found in GCV-sensitive isolates. Transfer marker experiments are in progress in order to test the significance of these DNA changes for GCV resistance. This rapid non-radioactive sequencing protocol could be a useful tool for analysing the UL97 region encoding the putative UL97 catalytic site of clinical isolates.

Amino Acid Sequence↗
PubMed1996

Non-radioactive assay of natural killer cell-mediated cytotoxicity against cytomegalovirus-infected fibroblasts by DNA fragmentation ELISA.

Cell-mediated cytotoxicity against cytomegalovirus (CMV)-infected fibroblasts (FS-4 cells) was investigated by a non-radioactive assay, and by DNA fragmentation ELISA and LDH release assay and the assays were compared to the standard chromium release assay. Fragmentation of DNA and LDH activity were detected in the supernatant of CMV-infected FS-4 cells cultured with non-adherent peripheral blood mononuclear cells (PBMC). The DNA fragmentation ELISA was most sensitive to cytotoxicity against CMV-infected FS-4 cells and showed excellent correlation with the standard chromium release assay. DNA fragmentation of CMV-infected FS-4 cells by non-adherent PBMC was reduced markedly by treatment with anti-leu 11b plus complement. Thus, the present DNA fragmentation ELISA is non-radioactive, highly sensitive and a useful method for detecting natural killer cell-mediated cytotoxicity against CMV-infected fibroblasts.

Adult↗
PubMed1996

Detection and variant identification of HHV-6 by a non-radioactive hybridization microplate assay for amplimers detection.

A non-radioactive hybridization microtiter plate assay was developed and evaluated for detection of the HHV-6 genome and to identify HHV-6 variants A and B. The viral DNA is amplified by the polymerase chain reaction using a 5'-end-biotinylated primer. The biotinylated amplimers are captured on avidin-coated microtiter plates, denaturated with sodium hydroxide and hybridized to a 3'-end-digoxigenin-labelled probe. Subsequently, anti-digoxigenin Fab fragments conjugated with alkaline phosphatase are used for the revelation of the hybridized probe. The result is obtained by measuring the intensity of light emitted with a spectrophotometer. This new assay was compared to the standard analysis of amplified products by Southern hybridization consisting of gel electrophoresis of the amplimers, transfer onto a nylon membrane, and hybridization with a 32P-labelled oligomeric probe. Both methods exhibited the same sensitivity and specificity. Thus, a non-radioactive hybridization microtiter plate assay may be a suitable alternative to isotopic techniques.

Biotin↗
PubMed1983

Effect of hycanthone administered in vivo upon the incorporation of radioactive precursors into macromolecules of Schistosoma mansoni.

Mice infected with Schistosoma mansoni were treated with hycanthone or with 8-chloro-2[2-(diethylamino)ethyl]-2H-[1]benzothiopirano-[4,3, 2-cd]-indazole-5-methanesulphonate (IA-4). Schistosomes were obtained by perfusion at various times after drug administration and tested for their ability to incorporate radioactive precursors of DNA, RNA and protein. In adult worms, male or female, the incorporation of radioactive thymidine was severely and irreversibly inhibited after treatment with either drug. Uridine and leucine incorporations were also inhibited, though to a lesser extent. On the contrary, the synthetic activities of immature worms were unaffected by hycanthone and only partially or temporarily depressed by IA-4. Hycanthone-resistant schistosomes, when tested between 1 and 7 days after treatment, showed a pattern of precursor incorporation which was virtually identical to that of untreated worms. These results are consistent with the hypothesis that hycanthone and IA-4 may kill schistosomes by interfering with their nucleic acid synthesis.

Animals↗
PubMed1995

Combined non-radioactive detection of peptide hormones and their mRNAs in stomach somatostatin cells.

Non-radioactive in situ hybridization (ISH) and immunocytochemistry (ICC) have been used to detect somatostatin (SS) messenger RNA (mRNA) and peptide in antropyloric mucosa of the stomach in the rats. We have applied a method of non-radioactive in situ hybridization histochemistry using digoxigenin labelled oligonucleotide probes to detect somatostatin gene expression in the stomach. In prehybridization stage we used proteinase K (PK) in various concentrations (from 1 to 10 micrograms/ml) and periods (from 10 min to 1 h) but we maintained high background. However it was possible to detect the somatostatin mRNAs in the stomach mucosa making use of either background preventing solutions during the prehybridization, or of levamisole (20 microliters/mg) added into the hybridization buffer or of pepsin. Somatostatin mRNA and peptide signals were scattered all through the mucosa especially localized particularly at the base of the pyloric glands. SS peptide shown by ICC and SS mRNA shown by ISH were observed in different cells.

Animals↗
PubMed1993

Stereotactic treatment of brain tumors with radioactive implants or external photon beams: radiobiophysical aspects.

We perform calculations, based on the linear-quadratic model, to assess the biologically effective doses (BED) of tumor and normal tissue in the stereotactic irradiation of brain tumors with either radioactive implants or radiosurgery techniques. Treatment protocols for radiosurgery and radioactive implants, as obtained from the literature, are reviewed and compared. A figure of merit is defined to be the ratio of tumor to normal tissue BED, expressed in units of Gy10/Gy3. These comparisons indicate a clear radiobiological advantage for brachytherapy, unless the radiosurgery is to be delivered in a large number of fractions. The differences in dose uniformity, and in the volume of normal tissue encompassed by the high dose regions, are factors that may also influence clinical results.

Brachytherapy↗
PubMed1990

Radioactive gold ring dermatitis.

A superficial squamous cell carcinoma developed in a woman who wore a radioactive gold ring for more than 30 years. Only part of the ring was radioactive. Radiation dose measurements indicated that the dose to basal skin layer was 2.4 Gy (240 rad) per week. If it is assumed that the woman continually wore her wedding ring for 37 years since purchase, she would have received a maximum dose of approximately 4600 Gy.

Carcinoma, Squamous Cell↗
PubMed1986

Fate and distribution of radioactive sodium diethyldithiocarbamate (Imuthiol) in the mouse.

The distribution of 35S in mouse tissues has been investigated by radioactivity counts and by autoradiography after the intravenous injection of 35S-labeled imuthiol (sodium diethyldithiocarbamate). Radioactive Imuthiol is selectively localized on liver, thymus and brain neocortex, most likely as the methyl ester, within minutes after dosing. Lung or white brain matter did not fix the labeled thiol. Blood, kidney and guts show rapid elimination patterns, and can be considered as passage organs which did not fix Imuthiol. The findings are consistent with the immunopharmacological data which demonstrate that Imuthiol exerts its T-cell recruiting and activating influence through a multi-step pathway involving the brain neocortex, the thymus and the liver.

Animals↗
PubMed1993

The occurrence and bioavailability of radioactive 137Cs in small forest lakes in Southern Finland.

Following the Chernobyl accident in late April 1986, central Finland was subjected to considerable radioactive fallout. The radioactive isotope of caesium, (137)Cs, was potentially the most harmful isotope in the fallout because of its long half-life (30 years). (137)Cs activities remained unexpectedly low within the biota (algae, zooplankton, aquatic moss, fish) of some humic forest lakes in southern Finland compared to the clear water lakes within the same district. This observation suggested that humic substances, by binding (137)Cs chemically, may reduce its bioavailability in lake water. Our laboratory experiments (throughflow systems; gel chromatography) demonstrated that binding of (137)Cs by humic substances was negligible in untreated lake waters; only when most of the alkali metals had been removed with an ion exchange resin was any binding of (137)Cs by low molecular humic fractions apparent. Instead, the concentrations of cations (especially potassium) were of overwhelming importance for the bioavailability of (137)Cs within the lacustrine ecosystems. The concentrations of (137)Cs within food chains are expected to stay at a high level for many years especially in lakes with long water renewal times.

Journal Article↗
PubMed1990

Excretion and blood radioactivity levels following [14C]senecionine administration in the rat.

Macrocyclic pyrrolizidine alkaloids (PAs) are a mixed group of phytotoxins with similar chemical structures and varying biological effects. A commonly studied member of this group is senecionine (SEN) which causes hepatotoxicity. We have undertaken metabolism and excretion studies of SEN in rats to provide data for comparison between PAs and to evaluate the potential role of metabolism and excretion in toxicity. Following intravenous administration of [14C]SEN (60 mg/kg, 10 microCi/kg), bile, urine and blood were collected over a 7-h period. Of the total administered radioactivity, 44% and 43% were excreted in the bile and urine, respectively. Using mass spectroscopy, senecionine N-oxide (SENNOX) was identified as the major metabolite in bile (52% of 44%) and urine (30% of 43%). For the total 7 h, less than 5% in bile and 18% in urine was excreted as parent alkaloid. The plasma concentration of Senecionine-equivalents/g (SEN-EQ/g) decreased from 107 to 12 nmol, while red blood cell (RBC) concentrations declined from 109 to 26 nmol/g. Without bile collection, the plasma levels of SEN-EQ were similar, while the final RBC level was almost double (47 vs. 26 nmol/g) and total radioactivity excreted in the urine was increased (59% vs. 43%). Biliary pyrrolic metabolites were estimated to be 1.43 mg, using a dehydroretronectin standard.

Animals↗
PubMed1981

Radioactive 2-deoxy-D-glucose incorporation into the prefrontal and premotor cortex of the monkey performing a forelimb movement.

Radioactive 2-deoxy-D-glucose (2-DG) incorporation into the monkey prefrontal and premotor cortex was studied in relation to extention-flexion movement at the wrist joint in two experimental and two control monkeys. With 2-DG injection and 45 min' intensive task performance thereafter, the following areas showed increased accumulations of radioactive glucose: the dorsomedial and dorsolateral prefrontal areas, including the lateral and medial banks of the principle sulcus; the ventral prefrontal and orbitofrontal areas; the cingulate gyrus, and the premotor cortex. In these areas, patch- or strip-like patterns were observed in the accumulation of 2-DG.

Animals↗
PubMed1991

Detection of mRNA encoding crustacean hyperglycemic hormone (CHH) in the eyestalk of the crayfish Orconectes limosus using non-radioactive in situ hybridization.

A non-radioactive in situ hybridization procedure for the localization of the mRNA encoding the crustacean hyperglycemic hormone (CHH) in the eyestalk of the crayfish Orconectes limosus has been developed. Based on the partial amino acid sequence of CHH, polymerase chain reactions were performed to generate complementary DNA (cDNA) clones encoding CHH. Non-radioactively labelled probes derived from the cDNA sequence were used to establish suitable conditions in terms of tissue fixation and pretreatment for detection of the CHH-encoding mRNA in combination with an immunocytochemical staining using a polyclonal antibody for CHH. Localization of the mRNA in the CHH perikarya was obtained with a complementary RNA probe in combination with pepsin/HCl treated Bouin-fixed eyestalks. The immunocytochemical staining confirmed that this cRNA probe specifically hybridized with mRNA of cell somata belonging to the CHH-producing cell system in the eyestalk of Orconectes limosus.

Amino Acid Sequence↗
PubMed1988

Adenocarcinoma of the prostate: radioactive gold seed implant plus external irradiation.

A total of 119 patients with a diagnosis of adenocarcinoma of the prostate (Stage B and C) were treated at St. Joseph Hospital Houston, Texas from 1971 through 1984 using a combination of radioactive gold seed implant and external irradiation. The prognostic significance of tumor grading and pelvic node involvement was analyzed. Five and 10-year survival for Stage B was 100% and 85% respectively; for Stage C it was 68% and 43% respectively. The cumulative 94% local control rate for Stage B and C cases obtained in this report suggests more effectiveness to control the disease locally with acceptable rate of complications, when radioactive gold seed implant is added.

Adenocarcinoma↗