Biomedical subjects

A Dasgupta

Publications and source records attributed to A Dasgupta.

At least 109 records · Page 6Linked to original sources

PubMed1998

Gas chromatography-electron ionization and chemical ionization mass spectrometric analysis of urinary phenmetrazine after derivatization with 4-carbethoxyhexafluorobutyryl chloride--a new derivative.

Phenmetrazine is a central nervous system stimulant currently used as an anorectic agent. The drug is abused and is reported to cause death from overdose. We describe a new derivatization method for phenmetrazine using 4-carbethoxyhexafluorobutyryl chloride. Quantitation of urinary phenmetrazine can be easily achieved by using N-ethyl amphetamine as an internal standard. The electron ionization mass spectrum of 4-carbethoxyhexafluorobutyryl derivative of phenmetrazine showed a molecular ion at m/z 427 and a base peak at m/z 70. In the methane chemical ionization mass spectrum, the base peak was observed at m/z 428 (protonated molecular ion). In the electron ionization mass spectrum of 4-carbethoxyhexafluorobutyryl derivative of the internal standard, N-ethyl amphetamine we did not observe a molecular ion. However, in the chemical ionization mass spectrum, the protonated molecular ion at m/z 414 was the base peak. The retention time of derivatized phenmetrazine (8.4 min) was substantially longer than the retention time of the underivatized molecule. Moreover, underivatized phenmetrazine showed poor peak shape (substantial tailing) while derivatized phenmetrazine had excellent chromatographic properties. The within-run and between-run precisions of the assay were 2.6% and 3.1% respectively at a urinary phenmetrazine concentration of 10 micrograms/mL. The assay was linear for urinary phenmetrazine concentration of 1 to 100 micrograms/mL with a detection limit of 0.2 microgram/mL.

Amphetamines↗

PubMed1997

Cleavage of transcriptional activator Oct-1 by poliovirus encoded protease 3Cpro.

In HeLa cells, RNA polymerase II mediated transcription is severely inhibited by poliovirus infection. Both basal and activated transcription are affected to bring about a complete shutoff of host cell transcription. We demonstrate here that the octamer binding transcription factor, Oct-1, is cleaved in HeLa cells infected with poliovirus. Incubation of Oct-1 with the purified, recombinant 3Cpro results in the generation of the cleaved Oct-1 product seen in virus infected cells. Poliovirus infection leads to the formation of altered Oct-1 DNA complexes that can also be generated by incubation of Oct-1 with purified 3Cpro. We also show that Oct-1 cleaved by 3Cpro loses its ability to inhibit transcriptional activation by the SV40 B enhancer. These results suggest that cleavage of Oct-1 in poliovirus infected cells leads to the loss of its activity.

3C Viral Proteases↗

PubMed1997

Lamotrigine analysis in plasma by gas chromatography-mass spectrometry after conversion to a tert.-butyldimethylsilyl derivative.

Lamotrigine (lamictal) is a new anticonvulsant drug recently approved by the FDA for clinical use. Therapeutic monitoring of lamotrigine is useful for patient management (therapeutic range 1-4 microg/ml). Here we describe a gas chromatography-mass spectrometric identification and quantitation of lamotrigine after extraction from human serum and derivatization. Lamotrigine was extracted from alkaline serum with chloroform and derivatized with N-methyl-N-(tert.-butyldimethysilyl) trifluoroacetamide containing 2% tert.-butyldimethylchlorosilane. Oxazepam-d5 was used as an internal standard. The tert.-butyldimethylsilyl derivative of lamotrigine showed distinct molecular ions at m/z 483 and 485 as well as other peaks at m/z 426, 370 and 334 for unambiguous identification. The base peak was observed at m/z 199. Similarly, the tert.-butyldimethysilyl derivative of oxazepam-d5 showed molecular ions at m/z 519 and 521 along with other characteristic peaks at m/z 462, 376 and 318. For the analysis of lamotrigine, the mass spectrometer was operated in the selective ion monitoring mode. The within-run and between-run precisions were 4.3% (mean=3.01, S.D.=0.13 microg/ml) and 5.1% (mean=2.93, S.D.=0.15 microg/ml), respectively at a serum lamotrigine concentration of 3.0 microg/ml. The within-run and between-run precisions were 8.2% (mean=0.49, S.D.=0.04 microg/ml) and 10.6% (mean=0.47, S.D.=0.05 microg/ml), respectively at a serum lamotrigine concentration of 0.5 microg/ml. The assay was linear for serum lamotrigine concentrations of 0.5-20 microg/ml. The detection limit was 0.25 microg/ml. The assay was free from interferences from common tricyclic antidepressants, benzodiazepines, other common anticonvulsants, salicylate and acetaminophen.

Anticonvulsants↗

PubMed1997

Gas chromatographic-mass spectrometric identification and quantitation of urinary phenols after derivatization with 4-carbethoxyhexafluorobutyryl chloride, a novel derivative.

Urinary phenol is analyzed widely to determine benzene exposure in humans. Most methods utilize direct measurements of phenols after extraction from urine using gas chromatography or high-performance liquid chromatography. We describe a novel derivatization of urinary phenols using 4-carbethoxyhexafluorobutyryl chloride after extraction from urine and subsequent analysis by gas chromatography-mass spectrometry. The derivative elutes at significantly higher temperature than phenol and the method is free from interferences from more volatile components in urine. We also observed excellent chromatographic properties of these derivatives. In addition, we observed strong molecular ions for the 4-carbethoxyhexafluorobutyryl derivative of phenol (m/z 344), p-cresol (m/z 358) and the internal standard 3,4-dimethylphenol (m/z 372) and other characteristic ions in the electron ionization, thus aiding in unambiguous identification of these compounds. The protonated molecular ions (m/z 373 for derivatized phenol, m/z 359 for derivatized p-cresol and m/z 373 for the internal standard) were the base peaks (relative abundance 100%) in the chemical ionization, although other secondary peaks were less abundant. The assay is linear for phenol concentration of 1-100 mg/l. The within-run and between-run precisions were 4.8% (mean = 52.4, S.D. = 2.5 mg/l) and 8.1% (mean = 53.0, S.D. = 4.3 mg/l) respectively, and the detection limit was 0.5 mg/l.

Acids↗

PubMed1997

Decreased total antioxidant capacity but normal lipid hydroperoxide concentrations in sera of critically ill patients.

Oxidative stress (when free radical generation exceeds antioxidant defense) occurs in many human diseases and makes significant contributions to their pathogenesis. We automated assays estimating the antioxidant status of serum, and lipid hydroperoxide concentrations using the Monarch analyzer. In this assay, the antioxidant status of serum is measured by its ability to inhibit generation of free radicals from 2,2'-amino-di-[3-ethylbenzthiazole sulphonate] by metmyoglobin and hydrogen peroxide. The assay for measuring lipid peroxidation products in serum utilizes the ability of lipid hydroperoxides to generate methylene blue from 10N-methylcarbamyl 3,7-dimethylamino 10H-phenothiazine. We detected no lipid hydroperoxide in sera obtained from 24 controls. The mean antioxidant status was 1.69 mmol/l (SD; 0.20 mmol/l) in controls. The antioxidant capacity of serum was significantly reduced in sera of critically ill patients in the medical intensive care unit (mean = 1.05, SD = 0.26 mmol/l) with p value less than 0.05 by independent t-test, two tailed. Lipid peroxidation products were not significantly elevated in critically ill patients, however lipid peroxidation products were significantly elevated in hemodialysis patients (mean = 1.40, SD = 0.47 mumol/l) and in some kidney transplant recipients. We conclude that antioxidant capacity of sera in critically ill patients is significantly reduced.

Antioxidants↗

PubMed1997

Decreased total antioxidant capacity and elevated lipid hydroperoxide concentrations in sera of epileptic patients receiving phenytoin.

Oxidative stress (free radical generation exceeds antioxidant defense) occurs in many diseases and after exposure to certain chemicals. We automated assays estimating the antioxidant status of serum and lipid hydroperoxide concentrations using the Syva-30R automated analyzer. In this assay, the antioxidant status of serum is measured by its ability to inhibit the generation of free radicals from 2,2'-amino-di-[3-ethylbenzthiazole sulphonate] by metmyoglobin and hydrogen peroxide. The assay for measuring lipid hydroperoxide concentration in serum utilizes the ability of lipid hydroperoxide to generate methylene blue from 10N-methylcarbamyl 3,7-dimethylamino 10H-phenothiazine. Phenytoin is known to initiate oxidative damage to proteins and lipids in murine maternal hepatic and embryonic tissue organelles. Therefore, we measured lipid hydroperoxide concentrations in sera of epileptic patients receiving phenytoin. We observed significantly elevated lipid hydroperoxide concentrations in epileptic patients receiving phenytoin compared to controls. The total antioxidant capacity of sera of epileptic patients was lower than the antioxidant capacity of control sera. We observed poor correlation between the serum lipid hydroperoxide concentration and triglyceride or cholesterol concentration in epileptic patients. We conclude that lipid hydroperoxide concentrations were elevated in sera of epileptic patients.

Antioxidants↗

PubMed1997

Chromatographic separation of DNA dependent RNA polymerases and molecular properties of RNA polymerase II from a Leishmania Spp.

Multiple forms of DNA-dependent RNA polymerases have been isolated and characterized from Leishmania strain UR6 promastigotes. RNA polymerases from this organism fail to resolve into multiple forms by conventional chromatography on DEAE-Sephadex A25, but could be separated by a modification of the method using CM-Sephadex C25. The CM-Sephadex bound enzyme is resistant to alpha-amanitin even up to a concentration of 250 micrograms/ml. The activity which flows through CM-Sephadex further resolves into two forms upon chromatography on DEAE-Sephadex A25. These forms are sensitive to alpha-amanitin to different extent. Enzyme activity in peak I is 50% inhibited by 3 micrograms/ml and in peak II by 50 micrograms/ml of the drug respectively. The enzyme in peak I has been further purified by heparin agarose and fast performance liquid chromatography (FPLC) on MonoQ. The enzyme has Stoke's radius of 70 A, a sedimentation coefficient of 17.6S and an f/fo of 1.35. Analysis of ammonium sulfate and metal ion optima of the enzyme in peak I, relative activities with Mn+2 versus Mg+2 and template specificities gave results similar to those reported for other type II RNA polymerases in eukaryotes. The MonoQ purified enzyme resolves into 16 polypeptides on denaturing polyacrylamide gel and densitometric analysis suggests that 9 major bands are present in the stoichiometry expected of RNA polymerase subunits having molecular weights: 154000; 104000; 77000; 64000; 52000; 48000; 46000; 45000 and 39000 respectively.

Animals↗

PubMed1997

Rapid detection of oleander poisoning using fluorescence polarization immunoassay for digitoxin. Effect of treatment with digoxin-specific Fab antibody fragment (ovine).

Poisoning from the oleander plant is common. Taking advantage of the high cross-reactivity of oleandrin, the major cardiac glycoside found in the oleander plant, we demonstrated that the serum digitoxin assay can be successfully used for the rapid diagnosis of oleander poisoning. Digitoxin is rarely used for treatment of cardiac disorders in the United States and has a therapeutic range of 19.7 to 39.3 nmol/L. In a typical oleander poisoning, serum oleandrin concentrations may reach 174 mmol/L or more. A serum specimen supplemented with 174 mmol/L of oleandrin containing no digitoxin showed an apparent digitoxin concentration of 1,272.1 nmol/L, a very high value compared with the range of the serum digitoxin assay, which is 2.6 to 104.8 nmol/L. Moreover, the response of the serum digitoxin assay with serum specimens containing various concentrations of oleandrin (and no digitoxin) is linear. Therefore, the oleandrin concentration in serum can be calculated from the apparent digitoxin concentration to access the severity of poisoning. Recently, the usefulness of the digoxin-specific Fab antibody fragment in the treatment of oleander poisoning has been described; however, no laboratory test was performed to demonstrate the progress of therapy. We demonstrated that the digoxin-specific Fab antibody can bind oleandrin in vitro, thus reducing the pharmacologically active free oleandrin. Because Fab and oleandrin bound to Fab are absent in the protein-free ultrafiltrates, monitoring the activity of free oleandrin in the ultrafiltrates can be used for monitoring the effectiveness of therapy.

Cardenolides↗

PubMed1997

Rickets and protein malnutrition in northern Nigeria.

The aim of the study was to explore the relationship between protein nutritional status and the development of rickets in children living in northern Nigeria. The diagnosis of rickets in 16 children between the ages of 10 months and 7 years was confirmed using established, and recently developed clinical and biochemical parameters. Twenty-seven children devoid of skeletal stigmata were age- and sex-matched to the rachitic patients. A battery of clinical laboratory and anthropometric measurements designed to assess calcium homeostasis, skeletal growth, the extent of bone remodeling or resorption, and protein nutritional status were performed on all subjects. Our central finding was that although the rachitic children were moderately malnourished, their protein nutritional status was significantly better as measured by the serum prealbumin concentration (15.4 v. 12.5 mg/dl, P = 0.0012) when compared with the severely malnourished children who were devoid of any indication of rickets. This may be due, in part, to the fact that actively growing children are more likely to develop rickets than are children whose linear growth is impeded. Unexpectedly, we found that the mean concentrations of serum 1,25-dihydroxyvitamin D in both the rachitic and control group were higher than any values for the active vitamin D metabolite previously reported in the literature.

Albumins↗

PubMed1997

Postmortem lipid levels for the analysis of risk factors of sudden death: usefulness of the Ektachem and Monarch analyzers.

Elevated serum cholesterol, triglyceride, and free fatty acid levels have been identified as risk factors for sudden death from cardiovascular disease and increased risk for myocardial ischemia or arrhythmias; therefore, correlation of antemortem and postmortem lipid levels may be useful in establishing the cause, pathophysiology, or familial risk factors of sudden death. In the present study, antemortem (within 72 h) and postmortem (within 24 h) cholesterol, triglyceride, free fatty acid, and albumin levels were analyzed in seven autopsied hospitalized patients from the University of New Mexico Hospital in Albuquerque, New Mexico. The cholesterol, triglyceride, and albumin levels were measured by dry-slide technology on an Ektachem 700 analyzer, and the free fatty acid levels were measured on a Monarch analyzer with a commercially available kit from Wako Chemical. Postmortem cholesterol levels averaged 13% lower than antemortem levels, postmortem triglyceride levels averaged 38% higher than antemortem levels, postmortem free fatty acid levels averaged 23% lower than antemortem levels, and postmortem albumin levels were essentially unchanged (<0.01% higher) from antemortem levels. Whether the antemortem and postmortem differences in lipid levels were the result of postmortem degradation products, a general phenomenon (such as variable enzyme degradation), or an idiosyncracy of the Ektachem or Monarch systems could not be definitely established. These preliminary results suggest that caution should be exercised when interpreting postmortem cholesterol, triglyceride, and free fatty acid levels analyzed on the Ektachem or Monarch systems.

Adult↗

PubMed1997

Elevated free fatty acid concentrations in lipemic sera reduce protein binding of valproic acid significantly more than phenytoin.

Higher concentrations of free valproic acid and phenytoin have been reported in patients with uremia and liver disease. Free fatty acids also displace valproic acid and phenytoin. This is a study of the magnitude of displacement of valproic acid and phenytoin from protein binding by free fatty acid in lipemic sera. Higher concentrations of free fatty acids in lipemic sera affected protein binding of valproic acid significantly more than that of phenytoin. Supplementing normal sera with free fatty acids also increased the free concentrations of both valproic acid and phenytoin as expected, but the observed effect was several times higher in magnitude with valproic acid. There was an increased free fraction of valproic acid in patients who received valproic acid and had hypertriglyceridemia. In a patient with uremia, there was also a significant increase in free valproic acid concentration after routine hemodialysis caused by an increase in free fatty acid concentration secondary to hemodialysis. Increased protein binding of valproic acid in sera was observed after treatment with activated charcoal because charcoal can remove free fatty acid. Because higher free fatty acid concentration significantly affects protein binding of valproic acid, careful monitoring of free valproic acid in patients with lipid disorder may be beneficial.

Bilirubin↗

PubMed1997

Effects of digoxinlike immunoreactive substances and digoxin FAB antibodies on the new digoxin microparticle enzyme immunoassay.

Digoxin-like immunoreactive substance (DLIS) is known to interfere with fluorescence polarization immunoassay (FPIA) (Digoxin II, Abbott Laboratories) and falsely elevates the total digoxin concentrations. Digoxin FAB antibody (Digibind) is also known to affect digoxin results by FPIA assay. The authors studied the effects of DLIS and Digibind on a new microparticle enzyme immunoassay (MEIA) for digoxin recently introduced by Abbott Laboratories, compared with the standard FPIA method and chemiluminescence assay (ACS-digoxin, Ciba-Corning). They studied 30 volume-expanded patients (term pregnancy, liver and renal disease) for the presence of DLIS. None of these patients received digoxin. They observed measurable DLIS concentrations in 12 of 30 patients by the FPIA assay and in only 1 patient by both MEIA and ACS assays. The concentration of DLIS in that patient was 0.31 ng/ml of digoxin equivalent by the MEIA assay, 0.36 ng/ml by the ACS assay, and 1.15 ng/ml by the FPIA assay. When they supplemented serum containing digoxin with low to high concentrations of digibind (0.5, 1.0, 2.0 and 4.0, 10, and 20 micrograms/ml), and measured digoxin concentrations by FPIA, MEIA, and ACS assays, they observed lower than expected values of total digoxin. However, when they supplemented serum containing no digoxin with high concentration of digibind (5.0, 10.0 and 20.0 micrograms/ml) and supplemented protein-free ultrafiltrates with digoxin, they observed expected digoxin concentrations in the ultrafiltrates by all three assays, indicating that the ultrafiltrates are essentially free of digibind.

Antibodies↗

PubMed1997

Analytical performance of a new chemiluminescent phenytoin (ACS:180) assay.

The authors, as a beta testing site, evaluated the ACS:180-phenytoin chemiluminescent assay (Ciba-Corning Diagnostics Corp., Medfield, MA, U.S.A.) by comparing its performance with a widely used fluorescence assay for phenytoin (Abbott Laboratories, Abbott Park, IL, U.S.A.). The ACS:180-phenytoin assays were run on a ACS-180 analyzer and fluorescence polarization assays on a TDx analyzer. The within-run precision for ACS-phenytoin assay was determined using controls obtained from Ciba-Corning. The CVs were 2.9% for low control (mean = 5.5, SD = 0.16 microgram/ml, n = 10), 2.8% for the medium control (mean = 13.4, SD = 0.37 microgram/ml, n = 10), and 2.7% for the high control (mean = 24.6, SD = 0.66 microgram/ml, n = 10). The corresponding between run precisions were 4.1% for the low control (mean = 5.4, SD = 0.22 mg/ml, n = 10), 3.1% for the medium control (mean = 13.8, SD = 0.43 mg/ml, n = 10), and 2.9% (mean = 24.5, SD = 0.70 mg/ml, n = 10) for the high control. The assay was linear from 0.5 to 40 micrograms/ml of serum phenytoin concentrations with a detection limit of 0.24 microgram/ml. The recoveries were 93-97% for concentrations of phenytoin of 5-30 micrograms/ml. They also compared 111 serum specimens collected from patients receiving phenytoin. The concentrations of phenytoin ranged from none detected to 32.4 micrograms/ml. Using fluorescence polarization assay as x-axis (reference method) and ACS:180-phenytoin assay as y-axis, they obtained the following regression line: y = 1.0x - 0.26, r = 0.993. They conclude that the ACS-phenytoin assay has a good precision and that the results correlate well with the fluorescence polarization assay.

Fluorescence Polarization↗

PubMed1997

Digitoxinlike and digoxinlike immunoreactivitis in sera of patients with uremia and liver disease as measured by fluorescence polarization immunoassays: poor correlation between digitoxinlike and digoxinlike immunoreactivities.

Digoxinlike immunoreactive substances (DLIS) cross-react with antidigoxin antibodies and falsely elevante total digoxin levels. Cross-reactivity of DLIS with various immunoassays for digoxin has been extensively studied in the past. The digitoxin molecule differs from digoxin by having one extra hydroxyl group in the aglycone ring. Therefore, DLIS may also falsely elevate total digitoxin concentrations. However, in the past, limited studies have shown the presence of digitoxinlike immunoreactive substances (DTLIS) in cord blood and sera of neonates. We compared DLIS and DTLIS concentrations in sera of patients with uremia, liver disease, and hypoalbuminemia. We found measurable DLIS concentrations in 11 of 45 patients by the fluorescence polarization immunoassay (FPIA) for digoxin (range, 0.20-0.89 ng/ml digoxin equivalent). These patients did not receive any digoxin or digitoxin. Using the FPIA, we also found elevated DTLIS concentrations in 10 of these 45 patients (range, 2.88-21.24 ng/ml digitoxin equivalent). Given the narrow therapeutic range of digitoxin (15-30 ng/ml), this cross-reactivity is significant. Elevated concentrations of DTLIS in sera falsely elevated the measured concentrations of digitoxin (positive interference) when known amounts of digitoxin were added to such sera. Interestingly, we found a poor correlation between DLIS and DTLIS concentrations in sera of patients with liver disease and uremia (r = 0.58), with some patients having no measurable DLIS activity but measurable DTLIS activity and vice versa. Digoxin showed only 4-8% cross-reactivity against digitoxin antibody with a wide range of concentration. Some proposed DLIS compounds (nonesterified fatty acids, cholic acid, lysophospholipid, and DHEA-sulfate) did not show any cross-reactivity with the digitoxin assay at a concentrations much higher than the physiologic range. We conclude that DTLIS activity is present in patients with liver disease and uremia, and the correlation between DLIS activity and DTLIS activity is poor. Moreover, some proposed DLIS do not explain the DTLIS activity detected in serum.

Digitoxin↗

PubMed1997

Valproic acid-ketoconazole interaction in normal, hypoalbuminemic, and uremic sera: lack of interaction in uremic serum caused by the presence of inhibitor.

Ketoconazole is an antifungal agent widely used in the management of patients with fungal infection, especially in patients with acute acquired immuno-deficiency syndrome (AIDS). Ketoconazole is 99% bound to serum albumin and may interact with valproic acid, an anticonvulsant with 90% to 95% binding to serum albumin. The interaction may be more significant in hypoalbuminemia, a common finding in patients with AIDS. However, valproic acid-ketoconazole interaction has not been reported. The authors prepared two serum pools from patients receiving valproic acid with normal serum albumin and another pool from patients with hypoalbuminemia. Another serum pool was prepared from uremic patients not receiving valproic acid. The aliquots of serum pool were supplemented with various concentrations of ketoconazole, representing therapeutic and slightly higher therapeutic concentrations. The concentrations of free valproic acid were determined in protein-free ultrafiltrates (prepared by centrifuging specimens at 25 degrees C with the Centrifree Micropartition System at 1500 g for 20 minutes) using fluorescence polarization immunoassay. In the serum pool with normal albumin concentration, the authors observed statistically significant displacement of valproic acid only at higher ketoconazole concentrations (10 and 20 micrograms/ml) whereas, in the serum pool with hypoalbuminemia, they observed statistically significant displacement of valproic acid by ketoconazole with lower and higher concentrations of ketoconazole. The magnitude of displacement was more significant at high valproic acid concentrations (95 and 150 mg/ml, respectively) probably because of the concentration-dependent binding of valproic acid to serum albumin. The authors observed no displacement of valproic acid by ketoconazole in the uremic serum pool. On the other hand, the free valproic acid concentrations were decreased in the presence of ketoconazole in the uremic serum pool.

Anticonvulsants↗

PubMed1997

A rapid cost-effective high-performance liquid chromatographic (HPLC) assay of serum lamotrigine after liquid-liquid extraction and using HPLC conditions routinely used for analysis of barbiturates.

Lamotrigine (lamictal) is a new anticonvulsant drug approved by the FDA for clinical use. Therapeutic monitoring of lamotrigine is useful for patient management and avoidance of toxicity. The suggested therapeutic range is 1 to 4 micrograms/ml. The authors describe a simple high-performance liquid chromatographic (HPLC) method for analysis of lamotrigine from serum. Serum (0.5 ml) was alkalinized with borate buffer (pH 9.8). Lamotrigine and the internal standard thiopental were extracted with 10 ml of chloroform. After evaporation of the extract, the residue was reconstituted in the mobile phase (prepared by mixing 750 ml of potassium dihydrogen phosphate, 550 ml of deionized water, 430 ml of methanol, and 100 microliters of triethylamine as an ion pairing reagent) and injected into an LC-18 column (15 cm x 4.6 mm). The authors use this HPLC system routinely in their laboratory for the analysis of barbiturates. They demonstrated that the same system can be used for the analysis of lamotrigine. The within-run and between-run precisions of the lamotrigine assay were 1.63% (mean = 3.05, SD = 0.05 microgram/ml, n = 6) and 3.7% (mean = 2.97 micrograms/ml, SD = 0.11, n = 8). The assay was linear for serum lamotrigine concentrations of 0.5 microgram/ml to 20 micrograms/ml with a detection limit of 0.5 microgram/ml. The authors observed excellent correlation between serum lamotrigine concentrations measured by their assay and a reference laboratory in six patients receiving lamotrigine. Their assay is free from interferences from common tricyclic antidepressants, benzodiazepines, other common anticonvulsants, salicylate, and acetaminophen.

Anticonvulsants↗

PubMed1997

Interference of oleandrin and oleandrigenin in digitoxin immunoassays: minimal cross reactivity with a new monoclonal chemiluminescent assay and high cross reactivity with the fluorescence polarization assay.

Toxicity from ingestion of the oleander plant is common. Oleandrin, the oleander glycoside, has structural similarity to cardiac glycoside digoxin and is known to cross react with various digoxin immunoassays. The authors studied the cross reactivity of oleandrin and its deglycosylated congener oleandrigenin with a fluorescence polarization immunoassay for digitoxin and compared their results with a new chemiluminescent assay for digitoxin on the Automated Chemiluminescent System (ACS:180 Plus) from Chiron Diagnostics. Even though the chemiluminescent assay has been reported to be comparable with the fluorescence polarization assay among normal patient population, oleandrin and oleandrigenin showed very high cross reactivities with the fluorescence polarization immunoassay and minimal cross reactivity with the new chemiluminescent assay. When the authors supplemented a serum specimen containing no digitoxin with 50 micrograms/ml of oleandrin, the fluorescence polarization assay recorded a value of 535.7 ng/ml of digitoxin equivalent, whereas the new chemiluminescent assay recorded a value of 10.3 ng/ml of digitoxin equivalent. The cross reactivity of oleandrigenin with the fluorescence polarization immunoassay for digitoxin was significantly lower than oleandrin. The presence of oleandrin also falsely elevated total digitoxin level in a specimen supplemented with digitoxin and oleandrin. The authors also measured free digitoxin concentration by the fluorescence polarization immunoassay in the ultrafiltrate of serum supplemented with digitoxin and oleandrin. Because digitoxin and oleandrin are bound strongly to protein, monitoring free digitoxin concentration by the fluorescence polarization immunoassay instead of total digitoxin concentration does not eliminate oleandrin interference. The authors conclude that fluorescence polarization immunoassay for digitoxin has a high cross reactivity with oleandrin and can falsely elevate digitoxin concentration in the presence of oleandrin, whereas the new chemiluminescent assay for digitoxin is almost free from interferences from oleandrin.

Anti-Arrhythmia Agents↗

PubMed1997

Poliovirus-encoded 2C polypeptide specifically binds to the 3'-terminal sequences of viral negative-strand RNA.

The poliovirus-encoded, membrane-associated polypeptide 2C is believed to be required for initiation and elongation of RNA synthesis. We have expressed and purified recombinant, histidine-tagged 2C and examined its ability to bind to the first 100 nucleotides of the poliovirus 5' untranslated region of the positive strand and its complementary 3'-terminal negative-strand RNA sequences. Results presented here demonstrate that the 2C polypeptide specifically binds to the 3'-terminal sequences of poliovirus negative-strand RNA. Since this region is believed to form a stable cloverleaf structure, a number of mutations were constructed to examine which nucleotides and/or structures within the cloverleaf are essential for 2C binding. Binding of 2C to the 3'-terminal cloverleaf of the negative-strand RNA is greatly affected when the conserved sequence, UGUUUU, in stem a of the cloverleaf is altered. Mutational studies suggest that interaction of 2C with the 3'-terminal cloverleaf of negative-strand RNA is facilitated when the sequence UGUUUU is present in the context of a double-stranded structure. The implication of 2C binding to negative-strand RNA in viral replication is discussed.

Base Sequence↗