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PubMed1986

Analysis of anti-HTLV-I antibody by strip radioimmunoassay--comparison with indirect immunofluorescence assay, enzyme-linked immunosorbent assay and membrane immunofluorescence assay.

Antibodies against human T-cell lymphotropic virus type I (HTLV-I) in the sera from 60 patients with adult T-cell leukemia and 21 carriers who were suspected of having HTLV-I infection were investigated by indirect immunofluorescence assay (IFA), enzyme-linked immunosorbent assay (ELISA), membrane immunofluorescence assay (MIA) and strip radio immunoassay based on the western blotting technique (SRIA). The sera of 2 of the carriers who were seropositive in IFA and ELISA were negative in MIA and did not react with virus-specific proteins by SRIA. Two sera were negative in IFA and ELISA. These sera were positive in MIA and reacted with only the envelope-related glycoprotein (gp46) and not with gag-related proteins (p28, p24, p19) by SRIA. These findings suggest that the main antigens defined by IFA and ELISA are gag-related proteins and some sera which do not contain anti-HTLV-I antibodies give false-positive results because of the reaction to unknown cellular components. Also some sera may have antibodies against only envelope glycoproteins, and these sera may give false-negative results in IFA and ELISA.

Antibodies, Viral↗

PubMed2004

Comparison of genotoxicity of textile dyestuffs in Salmonella mutagenicity assay, in vitro micronucleus assay, and single cell gel/comet assay.

The mutagenicity of textile dyes is an important consideration for the assurance of consumer protection and work safety. The mutagenicity testing of textile dyestuffs is crucial for accurately predicting health risks for consumers and workers exposed to dyes. Unfortunately, these data are often lacking. We studied the genotoxic activity of ten selected commercial textile dyestuffs, which are made up of mixtures of azo dyes and azo metal complex dyes as well as two anthraquinone dyestuffs. We used the Salmonella mutagenicity assay and cultured human keratinocytes (HaCaT cell line). In the S. typhimurium strain TA98, with and without S9, eight often dyestuffs investigated, and in strain TA 100, with and without S9, six often dyes caused frameshift mutations and base-pair substitutions in the dose range of 1-5000 microg/plate in a dose-related manner. All dyes, including those negative in the Salmonella mutagenicity assay, induced clastogenic effects in the in vitro micronucleus (MN) test in HaCaT cells as direct-acting mutagens in the concentration range of 5-150 microg/mL and with maximum MN frequencies between 1.1 and 7.2%, compared to negative controls that showed 0.2-0.4% MN cells. In the single cell gel/comet assay, all ten dyestuffs investigated caused DNA damage in HaCaT keratinocytes. The alkaline (pH >13) version used is capable of detecting DNA single strand breaks, alkali-labile sites, and DNA-DNA/DNA-protein cross-linking. Under the conditions of these screening tests, the textile dyes investigated are direct-acting genotoxic substances. The HaCaT cells testing protocol proposed has been shown to be an appropriate test system for evaluating mutagenicity of textile dyes on a base level.

Coloring Agents↗

PubMed2003

Laboratory diagnosis of two scrub typhus outbreaks at Camp Fuji, Japan in 2000 and 2001 by enzyme-linked immunosorbent assay, rapid flow assay, and Western blot assay using outer membrane 56-kD recombinant proteins.

Two scrub typhus outbreaks occurred among U.S. Marines training at Camp Fuji, Japan, between October 25 and November 3, 2000 and October 17 and November 30, 2001. Nine cases in approximately 800 Marines in 2000 and eight cases in approximately 900 Marines in 2001 (approximate attack rates = 1.1% and 0.9%, respectively) reported with signs and symptoms of fever, rash, headache, lymphadenopathy, myalgia, and eschar. Serologies and rapid response to doxycycline treatment indicated they had scrub typhus. Sixty-four convalescent serum samples (18 suspected cases and 46 negative controls) from U.S. Marines training at Camp Fuji during the outbreaks were assessed by enzyme-linked immunosorbent assay (ELISA), rapid flow assay (RFA), and Western blot assay for evidence of infection with Orientia tsutsugamushi, the causative agent of scrub typhus. All but one suspected case had serologic evidence of scrub typhus and all 46 control sera were non-reactive to O. tsutsugamushi antigens. The recombinant 56-kD antigen (r56) from the Karp, Kato and Gilliam strains of O. tsutsugamushi in an ELISA format provided better results than Karp r56 alone (ELISA and RFA) or whole cell antigen preparation from Karp, Kato and Gilliam (ELISA).

Adult↗

PubMed1993

Evaluation of viral membrane fusion assays. Comparison of the octadecylrhodamine dequenching assay with the pyrene excimer assay.

Membrane fusion, in particular the fusion of enveloped viruses, is often measured with an assay based on octadecylrhodamine (R18) fluorescence dequenching. We have studied the association of R18 with membranes and used the R18 assay to measure virus fusion in model systems and in cultured cells. The results were compared with those of an assay based on the decrease in excimer fluorescence of pyrene-labeled phospholipids. For liposomes made from premixed R18 and phosphatidylcholine (PC), R18 fluorescence quenching was proportional to the concentration of the probe up to about 4 mol %. No quenching was found at very low concentrations of R18. However, various artificial and biological membranes labeled by the addition of R18 from an ethanolic solution showed significant quenching at such low R18 concentrations. Thus, some of the R18 was not randomly distributed but likely was associated with the surface of the membranes in the form of highly quenched clusters or micelles. Moreover, in influenza virus membranes, R18 appeared highly quenched at very low concentrations, indicative of the probe interacting with viral proteins. In contrast, pyrene-labeled PC incorporated in either liposomes or reconstituted viral membranes (virosomes) showed an excimer/monomer fluorescence ratio proportional to the concentration of probe. When intracellular membrane fusion was investigated with R18-labeled influenza virus or Semliki Forest virus (SFV), fluorescence dequenching was observed in the absence of fusion, most likely due to spontaneous probe exchange.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed2004

Screening for antioxidant activity in edible plant products: comparison of low-density lipoprotein oxidation assay, DPPH radical scavenging assay, and Folin-Ciocalteu assay.

Oxidation of low-density lipoprotein (LDL) has been implicated in atherogenesis. Antioxidants that prevent LDL from oxidizing may reduce atherosclerosis. This study investigated LDL antioxidant activity in edible plant products for development of dietary supplementation to prevent atherosclerosis. Fifty-two kinds of edible plants were extracted using 70% aqueous ethanol solution, and the antioxidant activity of the extracts, which inhibit human LDL oxidation induced by copper ion, was determined on the basis of the oxidation lag time and represented as epigallocatechin 3-gallate equivalent. 1,1-Diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity and total phenolic content were also measured for comparisons with antioxidant activity in LDL. Plant products showing the greatest activity in LDL oxidation assay were akamegashiwa (Mallotus japonicus) leaf, Japanese privet (Ligustrum japonicum) leaf, green tea [Camellia sinensis (L.) O. Kuntze], and astringent persimmon (Diospyros kaki). The present study revealed high levels of LDL antioxidant activity in plant products for which such activity levels are underestimated in the DPPH radical scavenging assay and Folin-Ciocalteu assay.

Antioxidants↗

PubMed2003

[PTH assay: whole PTH assay (new IRMA assay)].

The common intact-PTH assay detects not only PTH (1-84) but also PTH (7-84) fragment. Recently, it is reported that PTH (7-84) fragment is the antagonist to PTH (1-84) biological action. Thus, conventional intact-PTH assay might mislead the overestimation of parathyroid function in uremic patients. Whole PTH assay, which detect only PTH (1-84) fragments may be useful for more precise evaluation of PTH activity in uremic patient.

English Abstract↗

PubMed2002

Evaluation of the genotoxic potential of some microbial volatile organic compounds (MVOC) with the comet assay, the micronucleus assay and the HPRT gene mutation assay.

Microbial volatile organic compounds (MVOC), metabolites of fungi detected in indoor moulds and in working places in compost facilities are considered as a potential health hazard. Their toxicological relevance, however, is largely unknown and data are rare. The aim of this study was to evaluate in vitro the genotoxic, clastogenic and mutagenic potential of same typical MVOC. For the study of DNA damage human lung carcinoma epithelial A549 cells, V79 Chinese hamster fibroblasts and human peripheral blood cells were exposed and subjected to the alkaline comet assay (single cell gel test). Taking the Chinese hamster V79 cell line as a target clastogenic effects were studied by the micronucleus test and mutagenic effects by the hypoxanthine-guanine-phosphoribosyl transferase gene mutation test (HPRT test). The cytogenic effects of MVOC were assessed by a clonogenic assay using the A549 cell line. The alkylating agent methyl methanesulfonate (MMS) was taken as a positive control. The results indicate that MVOC induced DNA damage is only seen in conditions in which also cytotoxic effects are observed. Clastogenic and mutagenic effects could not be detected.

Air Microbiology↗

PubMed1993

Comparison of vidas Clostridium difficile toxin-A assay and premier C. difficile toxin-A assay to cytotoxin-B tissue culture assay for the detection of toxins of C. difficile.

Damage to the intestinal mucosa by Clostridium difficile (CD) is toxin mediated. Two enzyme immunoassays (EIAs) for toxin-A detection, the automated Vitek immunodiagnostic assay system CDA (Vidas CDA), and the Premier toxin A (Premier) were tested for their ability to detect toxin A in 301 stool samples and compared with an in-house tissue culture assay for toxin B (TCA). Of these 301 samples, 49 were TCA positive and 252 were TCA negative. Agreement between Vidas CDA and TCA on the initial run was 85% (255 of 301) and increased to 94% (278 of 296) when discordant samples were retested from available frozen specimens. Corresponding levels of agreement for Premier were 91% (272 of 301) and 98% (284 of 288), respectively. If tissue culture positivity at any titer was used as the sole criterion for positivity of the specimen, agreement with positive TCA before and after repeat testing was 57% (26 of 49) and 74% (34 of 46) for Vidas CDA and 65% (32 of 49) and 95% (36 of 38) for Premier. Agreement with negative TCA titers was good: 90% for Vidas CDA and 95% for Premier, and 98% for Vidas CDA and 99% for Premier after repeat testing. Predictive values positive and negative after repeat testing were, respectively, 88% and 96% for Vidas CDA, and 95% and 99% for Premier. Results for the automated and manual EIA methods for detection of C. difficile toxin A were obtained in 2.5 h as compared with 36-48 h for tissue culture.

Bacterial Proteins↗

PubMed1996

Bicentric evaluation of Access Toxo immunoglobulin M (IgM) and IgG assays and IMx toxo IgM and IgG assays and comparison with Platelia Toxo IgM and IgG assays.

The recent Access immunoanalysis system (Sanofi Diagnostics Pasteur) for the serological diagnosis of toxoplasmosis was compared with the Abbott Toxo IMx EIA system, taking the Platelia Toxo immunoglobulin G (IgG) and Platelia Toxo IgM systems as references and using as confirmation methods an indirect fluorescence assay or a dye test for IgG and an immunosorbent agglutination assay (ISAGA) for IgM. A total of 1,461 serum samples were studied, of which 128 were collected from 42 recently seroconverted patients. Sensitivity and specificity rates of the Access system were 97.7 and 99.5%, respectively, for IgM and 98.6 and 100%, respectively, for IgG. Sensitivity and specificity rates of the Abbott IMx EIA system were 91 and 100%, respectively, for IgM and 92.5 and 100%, respectively, for IgG. The Access Toxo IgG and IgM EIA systems were found to be more sensitive than the Abbott Toxo IgG and IgM IMx EIA systems.

Animals↗

PubMed1984

Antibody assays for varicella-zoster virus: comparison of competitive enzyme-linked immunosorbent assay (ELISA), competitive radioimmunoassay (RIA), complement fixation, and indirect immunofluorescence assays.

The competitive ELISA test was compared with competitive RIA, complement fixation, and indirect fluorescent antibody test for the detection of antibody to varicella-zoster virus (VZV). ELISA and RIA were the most sensitive tests, being tenfold as sensitive as indirect immunofluorescence and 20 times as sensitive as complement fixation, which was surprisingly inadequate for detecting anti-VZV in sera from regular haemodialysis patients. The ELISA test was found to be the most suitable assay, employing a routine complement fixation test (CFT) grade viral antigen and not requiring radiolabelling and counting facilities, which are unavailable in most virology laboratories.

Adult↗

PubMed1993

[A comparative analysis of monoclonal assays for the determination of PSA: radioimmunometric assay (ELSA) vs microparticle immunoenzyme assay (MEIA)].

Comparative analysis of PSA values measured by MEIA and ELSA techniques in a group of 70 unselected patients. A good correlation was observed between PSA levels determined by ELSA-PSA immunoradiometric techniques and those obtained by MEIA-PSA (r = 0.93, p < 0.00001). However, ELSA-PSA values have been 1.73 +/- 0.1 times higher than those by MEIA-PSA. A mean-paired comparison indicates that PSA mean levels (0.48 +/- 0.07 and 0.29 +/- 0.05 for ELSA and MEIA, respectively) are significantly different and define two groups of nonhomogeneous values (p < 0.0001). The same results are obtained when patients with PSA values higher and lower than 4 ng/ml are analyzed separately. For patients with PSA lower than 1 ng/ml, the difference between mean ELSA-PSA and MEIA-PSA values disappears; 0.74 +/- 0.08 vs 0.62 +/- 0.05, respectively (p > 0.1). In this group, the results from both assays are statistically consistent. When considering the group of patients with PSA < 1 ng/ml, no difference between both techniques becomes apparent, which seems to indicate the absence of differences in sensitivity between both techniques when considering low levels of serum PSA. Nevertheless, it is clear that the results from these techniques can not overlap and are not comparable and so, to all practical effects, it is recommended that follow-up of any particular patient is made always with the same technique and even at the same laboratory.

Adult↗

PubMed1999

Comparison of the second-generation digene hybrid capture assay with the branched-DNA assay for measurement of hepatitis B virus DNA in serum.

The optimal hepatitis B virus (HBV) DNA quantitative assay for clinical use remains to be determined. We examined the sensitivity, linearity, and variability of a novel second-generation antibody capture solution hybridization assay, the Digene Hybrid Capture II assay (HCII), and compared it with another widely used solution hybridization assay, the branched-DNA (bDNA) assay (Quantiplex; Chiron Corp.). Our results showed similar and satisfactory assay linearity values, as well as interassay and intra-assay variability values, for both HCII and bDNA assays across different ranges of HBV DNA. Ninety-one percent of 102 serum samples from hepatitis B surface antigen-positive patients showed concordant results with the two assays. The HCII assay was more sensitive than the bDNA assay by 1 dilution, with the lowest reading being 0.9 pg/ml (3.8 pg/ml by bDNA assay). The HBV DNA seropositivity rates for the 102 samples were 58, 67, and 97% by bDNA, HCII, and nested PCR, respectively. While the relationship between results obtained with the bDNA assay and those with the HCII assay was nonlinear, with the bDNA assay yielding values 2.83 +/- 0.92-fold higher than those of the HCII assay, especially at high HBV DNA levels, a linear relationship was observed between the two sets of data after logarithmic conversion. The formula for interassay conversion of results was derived as follows: HBV DNA by HCII (picograms per milliliter) = 3.19 x [HBV DNA by bDNA (megaequivalents per milliliter)](0.866). The HCII assay was technically less complex and required a shorter assay time (4 h) than the bDNA assay (24 h). We conclude that the HCII assay compares favorably with the bDNA assay and offers the additional advantages of increased sensitivity and shorter assay time. The increased sensitivity should be particularly useful in monitoring the efficacy of antiviral therapies and detecting the emergence of drug-resistant HBV mutants.

Biological Assay↗

PubMed2004

Assay-specific decision limits for two new automated parathyroid hormone and 25-hydroxyvitamin D assays.

BACKGROUND: The recent development of nonradioactive automated assays for serum parathyroid hormone (PTH) and 25-hydroxyvitamin D (25OHD) has made measurement of these two hormones possible in many laboratories. In this study, we compared two new assays for PTH and 25OHD adapted on an automated analyzer, the LIAISON, with two manual immunoassays used worldwide. METHODS: We studied 228 osteoporotic patients, 927 healthy individuals, 38 patients with primary hyperparathyroidism, and 167 hemodialyzed patients. Serum PTH was measured with the Allegro and the LIAISON assays, and 25OHD was measured with DiaSorin RIA and the LIAISON assay. Regression analysis was used to calculate decision thresholds for the LIAISON assays that were equivalent to those of the Allegro PTH and DiaSorin 25OHD assays. RESULTS: The 25OHD concentrations obtained with the LIAISON assay and the RIA in osteoporotic patients were well correlated (r = 0.83; P <0.001). Regression and Bland-Altman analyses suggested that the LIAISON 25OHD assay reads lower than the DiaSorin RIA at low concentrations but higher at high concentrations. However, the cutoff (50 nmol/L) used in our laboratories to define vitamin D insufficiency with the DiaSorin RIA is applicable to the LIAISON 25OHD assay. In 927 healthy individuals, the 3rd-97th percentile intervals were 3-80 ng/L and 13-151 nmol/L for the LIAISON PTH and 25OHD concentrations, respectively. However, 506 individuals (54.6%) were vitamin D-insufficient; we therefore considered only the 421 individuals with a LIAISON 25OHD >50 nmol/L as eligible for the reference population for the LIAISON PTH assay. In this group, the 3rd-97th percentile interval for LIAISON PTH was 3-51 ng/L. Considering upper reference limits of 46 and 51 ng/L for the Allegro and LIAISON assays, respectively, the frequency of above-normal PTH concentrations in patients with primary hyperparathyroidism was similar in both assays. Regression analysis between serum PTH measured by the Allegro and LIAISON assays in 167 hemodialyzed patients and the corresponding Bland-Altman analysis of these data suggest that the LIAISON PTH assay tends to read higher than the Allegro assay at low concentrations but lower at high concentrations (>300 ng/L). CONCLUSIONS: Because clinical decision limits for both PTH and 25OHD should be assay specific, we propose equivalences between these assays and two manual assays used worldwide. These assay-specific decision limits should help potential users of the LIAISON PTH and 25OHD assays.

Autoanalysis↗

PubMed2004

Determination of estrogenic activity by LYES-assay (yeast estrogen screen-assay assisted by enzymatic digestion with lyticase).

In order to enhance the sensitivity and the speed of the yeast estrogen screen (YES)-assay, which has been established in many laboratories for the determination of estrogenic activity of compounds and environmental samples, the LYES-assay, a modified version of the YES-assay including a digestion step with the enzyme lyticase, was developed. With the LYES-assay the estrogenic activities of natural (17beta-estradiol E2 and estrone), synthetic (17alpha-ethinylestradiol EE2) and pharmaceutical estrogens (diethylstilbestrol DES) as well as xenoestrogens (4-nonylphenol NP and five parabens) were determined and compared with the results obtained by other in vitro-assays namely the conventional YES-assay, the E-Screen-assay (MCF-7 breast tumor cell proliferation) and a receptor binding-assay (RB) with human estrogen receptors hER-alpha and hER-beta. In the case of E2 the LYES-assay had a significantly lower limit of quantification (LOQ) than the conventional YES-assay and even two orders of magnitude lower than the RB-assay. Compared to the E-Screen-assay the LOQ of the LYES-assay was almost one order of magnitude higher. The time required to perform the LYES-assay was as little as seven hours compared to three to five days for the conventional YES-assay. Thus, the LYES-assay is a very good alternative to existing estrogenic in vitro-assays, since it has a good sensitivity, is cheap and much faster than the other assays.

Biological Assay↗

PubMed2002

Evaluation of the clinical usefulness of COBAS AMPLICOR HCV MONITOR assay (ver2.0): Comparison with AMPLICOR HCV MONITOR assay (ver1.0) and HCV core protein level.

The quantitation of serum levels of hepatitis C virus (HCV) RNA in chronic hepatitis C has been regarded as one of the most important indicators for the outcome of interferon (IFN) therapy. The AMPLICOR HCV MONITOR version 1.0 (AMPLICOR v1.0) assay is widely used for the evaluation of the HCV level. A new generation assay called the COBAS AMPLICOR HCV MONITOR version 2.0 (COBAS v2.0) assay, which is semiautomated and modified to amplify all genotypes equally, has been developed. The aim of this study was to evaluate the clinical relevance of the COBAS v2.0 assay in comparison with the AMPLICOR v1.0 assay and HCV core protein assay in patients with chronic hepatitis C before IFN therapy. HCV RNA was detectable in 230 cases (97.5%) and undetectable in 6 cases (2.5%) by the COBAS v2.0 assay. The RNA levels measured by the AMPLICOR v1.0 assay correlated significantly with those measured by the COBAS v2.0 assay, and the sensitivity of the new version 2.0 assay was better than that of version 1.0, especially in serotype 2. In relation to the outcome of IFN therapy, HCV RNA levels from virologically sustained responders by the AMPLICOR v1.0 assay were 82.3 +/- 22.9 kcopies/ml in serotype 1 and 36.9 +/- 13.4 kcopies/ml in serotype 2, and those from virologically nonsustained responders were 525.2 +/- 48.6 kcopies/ml in serotype 1 and 76.7 +/- 19.5 kcopies/ml in serotype 2. The rates of sustained response to <100 kcopies/ml were 34/63 (54.0%) in serotype 1 and 24/48 (50.0%) in serotype 2. A statistically significant virological response was seen in serotype 1 (P < 0.0001), but not in serotype 2. In contrast, the levels in virologically sustained responders by the COBAS v2.0 assay were 88.2 +/- 20.5 KIU/ml in serotype 1 and 136.8 +/- 40.1 KIU/ml in serotype 2, and those in virologically nonsustained responders were 608.8 +/- 48.4 KIU/ml in serotype 1 and 328.3 +/- 62.8 KIU/ml in serotype 2. The rates of sustained response to <100 KIU/ml were 33/60 (55.0%) in serotype 1 and 21/35 (60.0%) in serotype 2. Statistical significance in virological response was seen in both serotype 1 (P < 0.0001) and serotype 2 (P < 0.05). Although the sensitivity of the HCV core protein assay was lower than that with the COBAS v2.0 assay, the HCV core protein levels also correlated well with the results of the COBAS v2.0 assay. The HCV core protein levels of virologically sustained responders were 37.6 +/- 12.0 pg/ml in serotype 1, 81.3 +/- 37.0 pg/ml in serotype 2, and those of virologically nonsustained responders were 289.9 +/- 23.5 pg/ml in serotype 1, 191.4 +/- 32.1 pg/ml in serotype 2. This assay could predict the outcome of IFN therapy in both serotype 1 (P < 0.0001) and serotype 2 (P < 0.05). Thus, both the COBAS v2.0 assay and the HCV core protein assay showed that the viral load was an indicator of virologically sustained response in serotype 2 and in serotype 1.

Adolescent↗

PubMed2001

Dissection and weighing of accessory sex glands after formalin fixation, and a 5-day assay using young mature rats are reliable and feasible in the Hershberger assay.

The rodent Hershberger assay has been used predominantly by the pharmaceutical industry to evaluate androgenic and antiandrogenic chemicals for potential therapeutic use. However, this assay has not yet been formally validated and standardized for use in toxicology testing. There are many variations in the protocol used for this assay. The weight of androgen-dependent tissues is a definitive endpoint in the Hershberger assay. To find out the reliable assay protocol with feasibility, although many possible factors may affect assay reliability, the present study consist of a series of three separate experiments focused on method of dissection and weighing of accessory sex glands (ASGs: ventral and dorso-lateral prostate, seminal vesicles together with coagulating glands, and Cowper's glands), animal age and number of doses. Furthermore, male pubertal assay, an alternative to the Hershberger assay, was also examined its reliability. Experiment 1 explored whether reliably accurate ASG weights can be obtained after formalin fixation. The ASGs were collected from castrated male rats (11 weeks of age) treated daily with corn oil, or testosterone propionate (TP, 0.25 mg/kg/day, s.c.) and p,p'-DDE (0 or 100 mg/kg/day, p.o.) for 5 days. One day after the final treatment, the ASGs were removed carefully and weighed separately, and then fixed overnight in a 10% neutral-buffered formalin and weighed again. After that, the tissues were dried overnight in an oven and weighed again. A high correlation between fresh and fixed tissue weights, and a high correlation between fixed and dried tissue weights were noted. The changes in the tissue weight due to fixation were marginal and were proportional to the fresh weights of the individual tissue. Standard deviation of the fixed tissue weight in each group and the magnitude of responses to TP or p,p'-DDE in fixed tissues were equivalent to those in fresh or dried tissues. These findings indicate that formalin fixation does not interfere with interpretation of assay results, and this procedure was used in the subsequent experiments. Experiments 2 and 3 explored whether animal age or treatment duration altered assay sensitivity. In Experiment 2, antiandrogenic effect of p,p'-DDE (100 mg/kg/day) was detected after 5-and 10-day treatment irrespective of animal age (7 vs 11 weeks). In Experiment 3, antiandrogenic effects of flutamide (1 and 10 mg/kg/day) and p,p'-DDE (10 and 100 mg/kg/day) were compared between two different protocols, the 10-day assay using peripubertal rats and the 5-day assay using young mature rats. Results demonstrated that both protocols could significantly detect antiandrogenic effects of flutamide and p,p'-DDE. These findings demonstrate that (1) dissection and weighing of ASGs after formalin fixation is reliable in the Hershberger assay, (2) when this procedure is used, the 5-day Hershberger assay using young mature rats, expected to be more feasible assay than the 10-day assay using peripubertal rats, is also reliable as well as the 10-day assay using peripubertal rats. Furthermore, we confirmed that male pubertal assay with use of dissection and weighing of fixed tissues also reliable.

Androgens↗

PubMed2001

Comparison of a classical phagocytosis assay and a flow cytometry assay for assessment of the phagocytic capacity of sera from adults vaccinated with a pneumococcal conjugate vaccine.

Antibody- and complement-mediated phagocytosis is the main defense mechanism against Streptococcus pneumoniae. A standardized, easy to perform phagocytosis assay for pneumococci would be a great asset for the evaluation of the potential efficacy of (experimental) pneumococcal vaccines. Such an assay could replace the laborious phagocytosis assay of viable pneumococci (classical killing assay). Therefore, a newly developed phagocytosis assay based on flow cytometry (flow assay) was compared with the conventional killing assay and enzyme-linked immunosorbent assay (ELISA), using sera obtained from adults pre- and postvaccination with either a bivalent conjugate, a tetravalent conjugate, or the 23-valent polysaccharide vaccine. Highly significant correlations were observed between flow assay phagocytosis titers, killing assay phagocytosis titers, and ELISA antibody titers for serotype 6B and 23F as well. For serotype 19F, strong correlations were only observed between killing assay and ELISA titers. A potential drawback of the flow assay might be the low sensitivity compared with that of the killing assay. The choice of what assay to use, however, will depend on the objectives of the assay. When speed, easy performance, sample throughput, improved worker safety, absence of influence of antibiotics, and absence of false positives are the major criteria, the flow assay is the method of choice. When higher sensitivity is the major requirement, the classical killing assay should be used.

Adult↗

PubMed1999

Comparisons of a proposed five-seed assay method with the single-seed and batch assay methods for I-125 seeds in ultrasound-guided prostate implants.

A simple five-seed assay method was proposed and investigated. A commercial well ion chamber system with an NIST-traceable single-seed calibration constant was used for the single-seed assays. A batch seed holder was used for batch assays. For the five-seed assays, a second single-seed holder was modified such that all five seeds were loaded in a central region of the well ion chamber. Compared with the same seed in the standard single-seed holder, the relative chamber responses for the five seed positions were 0.993, 0.993, 1.000, 1.001, and 0.977, respectively, indicating little or no position-dependent chamber response and no self-attenuation among seeds. Subsequent comparison of assays with the single-seed and five-seed methods indicated only 0.4% difference in charge collection. The five-seed calibration constant was therefore taken to be the same as the single-seed calibration constant. The reproducibility of the five-seed assay method was found to be better than 0.8%. When a dummy seed replaced an active seed, a nearly 20% reduction in charge was found, indicating that the proposed five-seed assay method can detect a dead seed. Clinical comparison of all three assay methods showed that they produced qualitatively the same assay results when the batch assay method was performed with extra care. Compared with the single-seed assay method, the five-seed method is equally simple, rigid, and reproducible, but it demands much less assay time. Compared with the batch assay method, the five-seed method is much more reproducible and reliable because of its rigid assay geometry; it only demands a moderate amount of assay time and can detect dead seeds. The American Association of Physicists in Medicine Task Group 40 (AAPM TG40) states that, for brachytherapy, ideally every (i.e., 100%) loose seed should be calibrated. For procedures involving large number of loose seeds, it then recommends that 10% of seeds be calibrated. The proposed five-seed assay is very simple to implement. It will facilitate the compliance of the "10%" recommendation from the AAPM TG40; it will make the "ideally 100%" statement from AAPM TG40 a more realistic and practical QA procedure in seed assaying.

Brachytherapy↗