Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Reversibility”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 559 records · Page 31Linked to original sources
PubMed2004

Discriminations, reversals, and extra-dimensional shifts in the Göttingen minipig.

Göttingen minipigs were trained on a set-shifting procedure involving discriminations, reversals, and extra-dimensional shifts. The discriminations used were black-white discriminations and right-left discriminations. The initial visual and spatial discrimination seemed equally difficult, and only for the visual modality was reversal found to be more difficult than the initial discrimination. Visual reversal was more difficult than spatial reversal, and a larger number of perseverative sessions were found for visual reversal compared to spatial reversal. The acquisition of the extra-dimensional shift from the visual to the spatial dimension was not inferior to the learning of spatial reversal. Neither was the acquisition of the extra-dimensional shift from the spatial to the visual dimension inferior to the learning of visual reversal. Thus, no evidence was found for attention to stimulus dimensions in discrimination learning of the pigs.

Animals↗
PubMed2004

A group II intron-encoded maturase functions preferentially in cis and requires both the reverse transcriptase and X domains to promote RNA splicing.

Mobile group II introns encode proteins with both reverse transcriptase activity, which functions in intron mobility, and maturase activity, which promotes RNA splicing by stabilizing the catalytically active structure of the intron RNA. Previous studies with the Lactococcus lactis Ll.LtrB intron suggested a model in which the intron-encoded protein binds first to a high-affinity binding site in intron subdomain DIVa, an idiosyncratic structure at the beginning of its own coding region, and then makes additional contacts with conserved catalytic core regions to stabilize the active RNA structure. Here, we developed an Escherichia coli genetic assay that links the splicing of the Ll.LtrB intron to the expression of green fluorescent protein and used it to study the in vivo splicing of wild-type and mutant introns and to delineate regions of the maturase required for splicing. Our results show that the maturase functions most efficiently when expressed in cis from the same transcript as the intron RNA. In agreement with previous in vitro assays, we find that the high-affinity binding site in DIVa is required for efficient splicing of the Ll.LtrB intron in vivo, but in the absence of DIVa, 6-10% residual splicing occurs by the direct binding of the maturase to the catalytic core. Critical regions of the maturase were identified by statistically analyzing ratios of missense to silent mutations in functional LtrA variants isolated from a library generated by mutagenic PCR ("unigenic evolution"). This analysis shows that both the reverse transcriptase domain and domain X, which likely corresponds to the reverse transcriptase thumb, are required for RNA splicing, while the C-terminal DNA-binding and DNA endonuclease domains are not required. Within the reverse transcriptase domain, the most critical regions for maturase activity include parts of the fingers and palm that function in template and primer binding in HIV-1 reverse transcriptase, but the integrity of the reverse transcriptase active site is not required. Biochemical analysis of LtrA mutants indicates that the N terminus of the reverse transcriptase domain is required for high-affinity binding of the intron RNA, possibly via direct interaction with DIVa, while parts of domain X interact with conserved regions of the catalytic core. Our results support the hypothesis that the intron-encoded protein adapted to function in splicing by using, at least in part, interactions used initially to recognize the intron RNA as a template for reverse transcription.

Base Sequence↗
PubMed1998

Intravasal contraception with styrene maleic anhydride and its noninvasive reversal in langur monkeys (Presbytis entellus entellus).

In male langurs with azoospermia induced by vas occlusion with styrene maleic anhydride (SMA), the exploratory feasibility of azoospermia reversal by a new noninvasive reversal procedure has been assessed. Palpation, percutaneous electrical stimulation of the vas deferens, forced vibratory movement, suprapubic percussion, and per rectal digital massage of the vas deferens are the components of the multimodal noninvasive reversal procedure. The exploratory investigation reveals that a single application of the procedure leads to reversal of azoospermia. Normospermia with normal motility and viability appears after third ejaculation onwards after reversal manipulations. Ultrastructure of the spermatozoa, using scanning and transmission electron microscopy, revealed that the spermatozoa attained normalcy and sperm functional tests (i.e., hypo-osmotic swelling test, slide test for acrosome intactness, and test for sperm mitochondrial activity index) further confirmed the normalcy of the spermatozoa toward their fertilizing ability. Semen biochemistry appeared normal throughout the course of investigation. The morphology of the vas deferens, which showed exfoliation of the epithelium, was in the process of regaining normalcy after 90 days of reversal manipulations. The results suggest that noninvasive reversal technique offers the possibility of the functional azoospermia reversal within a short period of time.

Animals↗
PubMed1999

The thumb domain of the P51-subunit is essential for activation of HIV reverse transcriptase.

The biologically relevant and active form of human immunodeficiency virus reverse transcriptase is a heterodimer produced in a two-step dimerization process. Dimerization involves first the rapid association of the two subunits, followed by a slow conformational change yielding a fully active form. In the present study, we demonstrate that the interaction between the thumb domain of p51 and the RNase-H domain of p66 plays a major role in an essential conformational change required for proper folding of the primer/template and the tRNA-binding site, for maturation and for activation of heterodimeric reverse transcriptase. A synthetic peptide derived from the sequence within the thumb domain of p51, which forms the interface with the RNase-H domains of p66, binds heterodimeric reverse transcriptase with an apparent dissociation constant in the nanomolar range and selectively inhibits activation of heterodimeric reverse transcriptase with an inhibition constant of 1.2 microM. A detailed study of the mechanism of inhibition reveals that this peptide does not require dissociation of heterodimeric RT for efficient inhibition and does not affect subunit association, but interferes with the conformational change required for activation of heterodimeric reverse transcriptase, resulting in a decrease in the affinity of reverse transcriptase for the tRNA and an increase in the stability of the primer/template/reverse transcriptase complex. We have previously proposed that the dimeric nature of reverse transcriptase represents an interesting target for the design of antiviral agents. On the basis of this work, we propose that the conformational changes involved in the activation of reverse transcriptase similarly represent an important target for the design of novel antiviral compounds.

Amino Acid Sequence↗
PubMed2005

Serotonergic modulation of prefrontal cortex during negative feedback in probabilistic reversal learning.

This study used functional magnetic resonance imaging to examine the effects of acute tryptophan (TRP) depletion (ATD), a well-recognized method for inducing transient cerebral serotonin depletion, on brain activity during probabilistic reversal learning. Twelve healthy male volunteers received a TRP-depleting drink or a balanced amino-acid drink (placebo) in a double-blind crossover design. At 5 h after drink ingestion, subjects were scanned while performing a probabilistic reversal learning task and while viewing a flashing checkerboard. The probabilistic reversal learning task enabled the separate examination of the effects of ATD on behavioral reversal following negative feedback and negative feedback per se that was not followed by behavioral adaptation. Consistent with previous findings, behavioral reversal was accompanied by significant signal change in the right ventrolateral prefrontal cortex (PFC) and the dorsomedial prefrontal cortex. ATD enhanced reversal-related signal change in the dorsomedial PFC, but did not modulate the ventrolateral PFC response. The ATD-induced signal change in the dorsomedial PFC during behavioral reversal learning extended to trials where subjects received negative feedback but did not change their behavior. These data suggest that ATD affects reversal learning and the processing of aversive signals by modulation of the dorsomedial PFC.

Adolescent↗
PubMed1997

Efficient in vitro inhibition of HIV-1 gag reverse transcription by peptide nucleic acid (PNA) at minimal ratios of PNA/RNA.

We have tested the inhibitory potential of peptide nucleic acid (PNA) on in vitro reverse transcription of the HIV-1 gag gene. PNA was designed to target different regions of the HIV-1 gag gene and the effect on reverse transcription by HIV-1, MMLV and AMV reverse transcriptases (RTs) was investigated. We found that a bis-PNA (parallel antisense 10mer linked to antiparallel antisense 10mer) was superior to both the parallel antisense 10mer and antiparallel antisense 10mer in inhibiting reverse transcription of the gene, thus indicating triplex formation at the target sequence. A complete arrest of reverse transcription was obtained at approximately 6-fold molar excess of the bis-PNA with respect to the gag RNA. At this molar ratio we found no effect on in vitro translation of gag RNA. A 15mer duplex-forming PNA was also found to inhibit reverse transcription at very low molar ratios of PNA/ gag RNA. Specificity of the inhibition of reverse transcription by PNA was confirmed by RNA sequencing, which revealed that all tested RTs were stopped by the PNA/RNA complex at the predicted site. We propose that the effect of PNA is exclusively due to steric hindrance, as we found no signs of RNA degradation that would indicate PNA-mediated RNase H activation of the tested RTs. In conclusion, PNA appears to have a potential to become a specific and efficient inhibitor of reverse transcription in vivo , provided sufficient intracellular levels are achievable.

Base Sequence↗
PubMed2006

Regulation of hepadnavirus reverse transcription by dynamic nucleocapsid phosphorylation.

Reverse transcription, an essential step in the life cycle of all retroelements, is a complex, multistep process whose regulation is not yet clearly understood. We have recently shown that reverse transcription in the pararetrovirus duck hepatitis B virus is associated with complete dephosphorylation of the viral core protein, which forms the nucleocapsid wherein reverse transcription takes place. Here we present a genetic study of the role of this dynamic nucleocapsid phosphorylation in regulating viral reverse transcription. Detailed analyses of the reverse transcription products synthesized within nucleocapsids composed of core phosphorylation site mutants revealed that alanine substitutions, mimicking the nonphosphorylated state, completely blocked reverse transcription at a very early stage. In contrast, aspartate substitutions, mimicking the phosphorylated state, allowed complete first-strand DNA synthesis but were severely defective in accumulating mature double-stranded DNA. The latter defect was due to a combination of mutant nucleocapsid instability during maturation and a block in mature second-strand DNA synthesis. Thus, the reversible phosphorylation of the nucleocapsids regulates the ordered progression of reverse transcription.

Amino Acid Sequence↗
PubMed2000

Sequences in the 5' and 3' R elements of human immunodeficiency virus type 1 critical for efficient reverse transcription.

The genome of human immunodeficiency virus type 1 (HIV-1) contains two direct repeats (R) of 97 nucleotides at each end. These elements are of critical importance during the first-strand transfer of reverse transcription, during which the minus-strand strong-stop DNA (-sssDNA) is transferred from the 5' end to the 3' end of the genomic RNA. This transfer is critical for the synthesis of the full-length minus-strand cDNA. These repeats also contain a variety of other functional domains involved in many aspects of the viral life cycle. In this study, we have introduced a series of mutations into the 5', the 3', or both R sequences designed to avoid these other functional domains. Using a single-round infectivity assay, we determined the ability of these mutants to undergo the various steps of reverse transcription utilizing a semiquantitative PCR analysis. We find that mutations within the first 10 nucleotides of either the 5' or the 3' R sequence resulted in virions that were markedly defective for reverse transcription in infected cells. These mutations potentially introduce mismatches between the full-length -sssDNA and 3' acceptor R. Even mutations that would create relatively small mismatches, as little as 3 bp, resulted in inefficient reverse transcription. In contrast, virions containing identically mutated R elements were not defective for reverse transcription or infectivity. Using an endogenous reverse transcription assay with disrupted virus, we show that virions harboring the 5' or the 3' R mutations were not intrinsically defective for DNA synthesis. Similarly sized mismatches slightly further downstream in either the 5', the 3', or both R sequences were not detrimental to continued reverse transcription in infected cells. These data are consistent with the idea that certain mismatches within 10 nucleotides downstream of the U3-R junction in HIV-1 cause defects in the stability of the cDNA before or during the first-strand transfer of reverse transcription leading to the rapid disappearance of the -sssDNA in infected cells. These data also suggest that the great majority of first-strand transfers in HIV-1 occur after the copying of virtually the entire 5' R.

Animals↗
PubMedDate not supplied

Clinical implications of HIV drug resistance to nucleoside and nucleotide reverse transcriptase inhibitors.

This review focuses on issues pertinent to the epidemiology and clinical interpretation of resistance to nucleos(t)ide reverse transcriptase inhibitors. Nucleoside reverse transcriptase inhibitor resistance mutations, and especially thymidine analog mutations, remain the most common form of resistance detected in drug-naive patients. At the same time, improved treatment strategies, changes in prescribing policies, and prompt management of treatment failure are changing the prevalence and patterns of nucleoside reverse transcriptase inhibitor resistance in treatment-experienced patients. The clinical interpretation of nucleoside reverse transcriptase inhibitor resistance is being increasingly refined, helped by improved understanding of resistance pathways, development of sophisticated methods of analysis of genotypic resistance patterns, and introduction of clinically relevant cutoffs. Correlation of genotypic and phenotypic resistance data to clinical outcomes is essential to allow appropriate interpretation. In some cases, phenotypic data, either obtained directly by phenotypic tests or extrapolated from genotypic results, provide the most immediate predictors of virologic response. In other cases, genotypic analyses identify mutations that impact on responses without showing a marked effect on the phenotype, by either acting as "sentinel" markers for the presence of resistance undetectable by standard methods, or by lowering the genetic barrier to the evolution of resistance. The potential benefits of nucleoside reverse transcriptase inhibitor resistance, through hypersusceptibility and fitness effects, are also increasingly understood and exploited in clinical practice. Although progress has been significant, there remain many challenges. It is often questioned whether genotypic scores and clinical cutoffs obtained by various methods and from frequently small datasets can be reliably extrapolated to the general population of treated patients. At the same time, there is a need to define the role of reverse transcriptase mutations that are identified by statistical analyses as being associated with nucleoside reverse transcriptase inhibitor exposure, but have unknown effects on virus phenotype and clinical outcome. Novel mechanisms have also been proposed to play a role in nucleoside reverse transcriptase inhibitor resistance, including changes in RNaseH that are not targeted by routine testing at present. One additional, currently unresolved issue is the clinical relevance of minority resistant species and the feasibility of introducing ultra-sensitive resistance tests in routine diagnostic settings. The most appropriate viral-load cutoff for performing resistance tests and the reliability of results obtained at low copy numbers are similarly controversial. In spite of these limitations, resistance testing with appropriate interpretation provides an important guide to successful treatment outcomes and necessary support to the introduction of new treatment strategies.

Drug Resistance, Multiple, Viral↗
PubMed1995

Long-term morbidity following jejunoileal bypass. The continuing potential need for surgical reversal.

OBJECTIVE: To review the late sequelae of jejunoileal bypass (JIB) and the potential role of late surgical reversal in ameliorating morbidity and mortality following JIB. DESIGN: Patients who underwent JIB between 1965 and 1977 were contacted and pertinent health-event information was gathered. Early sequelae were defined as disorders occurring within the first 2 years after JIB; late sequelae were those occurring after 2 years. Health events occurring between 0 and 23 years after JIB were documented. SETTING: A private, tertiary referral center. PATIENTS: Patients underwent JIB for morbid obesity that had failed medical and/or psychiatric interventions. MAIN OUTCOME MEASURES: Body mass index (BMI) (weight kilograms divided by the square of the height in meters), diarrhea, electrolyte imbalance, acute, and chronic liver disease, renal disease, JIB reversal, reason for JIB reversal, death, and cause of death. RESULTS: A total of 453 morbidity obese patients underwent JIB. By 2 years following JIB, the mean (+/- SD) BMI dropped from 49.3 +/- 8.1 to 31.1 +/- 0.8 and remained at this level until year 15, after which weight gradually increased (BMI, 35.4 +/- 3.1). The most severe early complication was acute liver failure, which occurred in 7% of patients and caused seven deaths. At 15 years, the actuarial probability of the most common serious late complications related to JIB were renal disease (37%), with two deaths; diarrhea (29%); and liver disease (10%), with three deaths. One hundred thirty-eight patients (31%) had a bypass reversal. The most common indications for reversal were diarrhea and electrolyte disturbance (29%), renal disease (19%), and liver disease (17%). Fifty-six patients died more than 30 days after JIB: 64% before JIB reversal, 13% at the time of reversal, and 23% subsequently. CONCLUSIONS: Jejunoileal bypass is associated with progressive accrual of serious, sometimes life-threatening complications. Lifelong follow-up for early diagnosis and surgical reversal before life is threatened should reduce the morbidity and mortality associated with this procedure.

Acid-Base Imbalance↗
PubMed2004

Activation of lignin peroxidase in organic media by reversed micelles.

Activation of lignin peroxidase (LIP) in an organic solvent by reversed micelles was investigated. Bis(2-ethylhexyl)sulfosuccinate sodium salt (AOT) was used as a surfactant to form a reversed micelle. Lyophilized LIP from an optimized aqueous solution exhibited no enzymatic activity in any organic solvents examined in this study; however, LIP was catalytically active by being entrapped in the AOT reversed micellar solution. LIP activity in the reversed micelle was enhanced by optimizing either the preparation or the operation conditions, such as water content and pH in water pools of the reversed micelle and the reaction temperature. Stable activity was obtained in isooctane because of the stability of the reversed micelle. The optimal pH was 5 in the reversed micellar system, which shifted from pH 3 in the aqueous solution. The degradation reaction of several environmental pollutants was attempted using LIP hosted in the AOT reversed micelle. Degradation achieved after a 1-h reaction reached 81%, 50%, and 22% for p-nonylphenol, bisphenol A, and 2,4-dichlorophenol, respectively. This is the first report on the utilization of LIP in organic media.

Colloids↗
PubMed1987

Prospectively randomized clinical trial to compare in situ and reversed saphenous vein grafts for femoropopliteal bypass.

Two hundred and fifteen femoropopliteal bypass procedures using autologous saphenous vein grafts were randomly allocated to either the reversed or in situ technique. Eleven veins (5 per cent) were rejected at operation on the basis of their small size, nine in the reversed group and two in the in situ group, and there were two (2 per cent) perioperative deaths in each group, leaving 102 reversed and 98 in situ grafts for further study. The cumulative patency at 3 years of the reversed grafts was 77 per cent and that of the in situ grafts was 68 per cent (n.s.). The patency of all grafts was affected adversely by small veins (P less than 0.005), long grafts (P less than 0.05), low volume of blood flow in the grafts (P less than 0.001) and poor run-off (P less than 0.05). These factors influenced the outcome of the in situ and reversed operations to a similar degree and there was no statistically significant difference between them within any subgroup. The mean compliance of the in situ grafts measured 3 months or more after operation with an ultrasonic echo-tracking system was 0.024 +/- 0.01 per cent/mmHg (+/- s.d.) compared with 0.017 +/- 0.01 per cent/mmHg for the reversed grafts (t = 2.43, P less than 0.02). The incidence of fibrous stricture formation as shown by intravenous digital subtraction angiography was 29 per cent in both the reversed and the in situ grafts. The results of the study to date indicate that reversed and in situ vein grafts are equally effective for femoropopliteal bypass.

Adult↗
PubMed1993

Prospective randomized comparison of in situ and reversed infrapopliteal vein grafts.

A three-centre prospective randomized trial was undertaken to compare the efficacy of in situ and reversed saphenous vein grafts for long bypasses to tibial and peroneal arteries. Of 162 patients entered into the study, 82 received an in situ graft and 80 a reversed vein graft. All operations were for limb salvage and the two groups of patients were well matched for age, sex, incidence of diabetes, smoking habits and coronary artery disease. At a maximum follow-up of 3 years there were 48 primary graft failures: 19 in the in situ group and 29 in the reversed vein group. Of these, three in situ grafts and seven reversed grafts were salvaged by secondary intervention. Secondary cumulative patency rates calculated at 3 years after operation were 68 and 66 per cent respectively for in situ and reversed grafts (P not significant). Cumulative limb salvage rates were 78 per cent for in situ grafts and 87 per cent for reversed grafts (P not significant). Separate analysis of a subgroup with small veins (< or = 3.5 mm minimum diameter) showed cumulative patency rates at 2 years of 74 per cent for in situ grafts and 60 per cent for reversed grafts (P not significant). These results indicate that for veins > 3.5 mm in diameter the in situ and reversed techniques for operation are equally effective. Some doubt remains about the best way of using smaller veins; a large number of such veins need to be studied to resolve this question.

Aged↗
PubMed2006

Degradation patterns of tetracycline antibiotics in reverse micelles and water.

The objective of this study was to determine the chemical stability of tetracycline and oxytetracycline hydro-chlorides in reverse micelles. Their reverse micellar solutions were prepared using cetyltrimethylammonium bromide, water and ethyl formate. The aqueous solutions of the tetracycline antibiotics were also prepared for comparison. The reverse micellar and aqueous solutions were stored at 37 degrees C. Samples were analyzed by high performance liquid chromatography. When evaluation was performed on an aqueous tetracycline HCl solution, its half-life was estimated to be 329 h. Its chemical stability was not improved after being dissolved in the reverse micelles, and a similar half-life of 330 h was observed. However, there were noticeable differences between the two systems in terms of degradation kinetics and degradation byproducts. On the other hand, oxytetracycline HCl was unstable in water so that its half-life was only 34 h. Very interestingly, pronounced improvement in stability was attained with the reverse micellar system: upon dissolving in the reverse micelles, its half-life was increased to 2402 h. There were also marked differences in degradation patterns and mechanisms of oxytetracycline HCl in water and the reverse micelles. Our study indicates that the reverse micellar system has potential applications in solubilizing and stabilizing oxytetracycline HCl, thereby contributing to the development of its dosage forms.

Cetrimonium↗
PubMed2005

Effects of prolonged acclimation to intermediate photoperiod and photo-schedule reversal in photosensitive golden hamsters.

We investigated the effect of prolonged acclimation to 12 hr of light and photo-schedule reversal during the time of photosensitivity in golden hamsters (Mesocricetus auratus). Before the experiments, animals were housed under natural photoperiod and then transferred to 12L:12D (light 12 hr:dark 12 hr) in autumn for 12 weeks. After 4 weeks of acclimation, photo-schedule was reversed (12D:12L). First experiments were done after 4 weeks of acclimation to an ambient temperature (T(a)) of 23 degrees C and a 12L:12D photo-schedule. We examined the daily variations in brown adipose tissue (BAT) capacity for nonshivering thermogenesis (NST). Noradrenaline (NA) injections were given every 4 hr while BAT temperature (T(BAT)) and preferred ambient temperature (PT(a)) were monitored continuously and simultaneously in a thermal gradient system. Then, we investigated the effect of light-dark cycle reversal on a daily rhythm of NST. The hamsters were acclimated to the photo-schedule reversed by 12 hr and the same T(a). After 4 and 8 weeks of acclimation to a reversed photo-schedule, the experiments were repeated. We found that the daily rhythm of the response to NA was entrained to the new light-dark cycle after 4 weeks of acclimation to a reversed photo-schedule. Maximum effect of NA was always recorded during the light phase and in the latter part of the dark phase of the day. NA-induced increase in T(BAT) was correlated with the decrease in PT(a), and was also inversely correlated with pre-injection T(BAT). These data imply that the daily rhythm of the capacity for NST opposes the daily rhythm of body temperature (T(b)). After 8 weeks of acclimation to the reversed photo-schedule, the rhythmicity of the response to NA disappeared, and the daily fluctuations in T(BAT) were the smallest. This lack of rhythm may be a physiological adaptation to winter conditions when the daily amplitude of T(b) rhythm is markedly reduced and, as a consequence, NST capacity does not vary within the day. Moreover, after 8 weeks of acclimation to reversed photo-schedule, NST capacity decreased while response to saline increased. During the experiments, hamsters were photosensitive and were changing to their winter status. However, because of the lack of cold during acclimation, the capacity for NST did not increase. Increased responsiveness to saline, indicating an increase in stress-induced thermogenesis, might be advantageous for "fight or flight" reaction.

Acclimatization↗
PubMed1996

Antigen--antibody interactions in the reverse micellar system: quenching of the fluorescence of fluorescein-labeled atrazine by antibodies against atrazine.

This work presents a new method for performing homogeneous fluoroimmunoassay in apolar organic media, quenching fluoroimmunoassay (QFIA). This method is based on utilization of the reverse micellar system of Aerosol OT (AOT) in n-octane as a medium for the analysis of compounds with low water solubility. It is shown using the system for determination of a hydrophobic pesticide atrazine as an example. The conjugate of atrazine with fluorescein (FA) serves as a label for fluorescence detection of antigen-antibody interaction in the reverse micellar system. The fluorescence quantum yield of this compound drastically depends on the micro-environment of the label in the reverse micelle system. Specifically, the binding of this conjugate with the antibodies solubilized in the reverse micelles results in fluorescence quenching. We found that quenching efficiency depends on the properties of the reverse micellar system (surfactant concentration, hydration degree w0, w0 = [water]/[surfactant], etc.). The optimal conditions for quenching of FA fluorescence by antibodies in reverse micelles of AOT in n-octane are low surfactant concentration and hydration degree, allowing one to get large reversed micelles (w0 = 15-20) capable of retaining solubilized antibodies. Addition of free atrazine results in displacement of the conjugate and restoration of its fluorescence. The sensitivity of the analysis to atrazine is only 10 times less than that of the commonly used method of homogeneous immunoassay, polarization fluoroimmunoassay, in aqueous solution using the same antibodies and conjugate. The advantage of QFIA in reverse micelles is that the analyte can be added when dissolved in nonpolar organic solvent.

Animals↗
PubMed2000

Effects of Nucleoside Analogs on Native and Site-Directed Mutants of HTLV Type 1 Reverse Transcriptase.

A bacterial assay was developed for testing HTLV-1 reverse transcriptase sensitivity to common nucleoside analog inhibitors in an Escherichia coli strain characterized by a temperature sensitive PolI/RecA deletion phenotype. This genetic complementation assay exploits the ability of HTLV-1 reverse transcriptase to functionally replace these missing activities at nonpermissive temperatures. The four inhibitors tested, dideoxyinosine, dideoxyadenosine, deoxythymidine, and didehydrodeoxythymidine are well-known inhibitors of HIV reverse transcriptase. All except dideoxyadenosine showed a strong activity against HTLV-1 reverse transcriptase with IC(50); in the nanomolar range. Sequence alignments were used to identify amino acid residues in HTLV-1 reverse transcriptase, which correspond to those identified as important for drug-resistance in HIV reverse transcriptase. Mutations of some of these HTLV-1 residues altered the IC(50) for the inhibitors as expected, which suggests that these amino acids have a function in HTLV-1 reverse transcriptase similar to that of their homologs in HIV reverse transcriptase. Copyright 2000 Academic Press.

Journal Article↗
PubMed2000

Reversible cross-linking and CO treatment as an approach in red cell stabilization.

We explored the use of the reversible cross-linking reagent dimethyl 3,3-dithiobispropionimidate (DTBP) in combination with CO treatment as an approach to stabilizing erythrocyte structure and function. Erythrocytes were cross-linked with different concentrations of DTBP for different times. DTBP increased erythrocyte osmotic stability, blocked lysolecithin-induced echinocytosis, and decreased erythrocyte deformability in a concentration- and time-dependent manner. Reversal of the cross-linking with the reducing agent dithioerythritol (DTE) restored osmotic fragility and response to lysolecithin as well as deformability. Complete reversal, however, is a function of the DTBP concentration and the time of cross-linking. The effects of cross-linking with 5 mM DTBP for 1 h were completely reversible after treatment with 10 mM DTE for 20 min. Longer incubation times or higher concentrations of DTBP resulted in partial reversal by DTE of the effects produced by DTBP. Cross-linking and reversal only slightly reduced the ATP content. The hemoglobin contained in the cross-linked and reversed cells could still undergo reversible oxygenation and deoxygenation. Erythrocytes were pretreated with CO, cross-linked with 5 mM DTBP for 1 or 3 h, loaded with a solution containing 500 mM glucose for 24 h, and freeze-dried in a medium containing 15% (w/v) albumin. Rehydration followed in distilled water. Complete recovery, measured as the percentage of free hemoglobin, was achieved for cells cross-linked with 5 mM DTBP for 3 h and freeze-dried to a final water content of 10-15%. Non-cross-linked cells lysed 100% on rehydration in distilled water. No methemoglobin (MetHb) formation as a result of freeze-drying was detected in CO-treated cells. In non-CO-treated cells 20% of the Hb was converted to MetHb.

Adenosine Triphosphate↗