Lymphatic filariasis: possible pathophysiological nexus with oxidative stress.

Wuchereria bancrofti-mediated lymphatic filariasis is widely prevalent. Diversity in immune response presumably may lead to myriad clinical presentations, such as overt chronic filariasis, occult filariasis with atypical systemic manifestation and asymptomatic microfilariae carrier state. Anticipated oxidative stress during inflammatory response to infective conditions might complicate the immune response and thus might alter the disease outcome. The present study was carried out to assess the status of oxidative stress in different clinical presentations of bancroftian filariasis. Twenty-five microfilariae carriers and 30 cases each of chronic filariasis and occult filariasis were compared to 30 endemic normal individuals. Serum malondialdehyde level and superoxide dismutase enzyme activity were measured by spectrophotometric methods and levels of filarial antigen were measured by ELISA. In the filarial cases, the levels of these parameters were assayed again after treatment with diethylcarbamazine citrate (DEC). Results showed significant (P<0.05) association of oxidative stress with chronic and occult filariasis but not with microfilarial carriers. DEC therapy in both clinical cases and carriers resulted in a significant reduction of oxidative stress associated with decreased antigen level (P<0.01). These findings suggest the possible involvement of oxidative stress in filarial disease pathology.

The effect of temperature and oligonucleotide primer length on the specificity and efficiency of amplification by the polymerase chain reaction.

The polymerase chain reaction (PCR) is most effectively performed using a thermostable DNA polymerase such as that isolated from Thermus aquaticus. Since temperature and oligonucleotide length are known to control the specificity of oligonucleotide hybridization, we have investigated the effect of oligonucleotide length, base composition, and the annealing temperature on the specificity and efficiency of amplification by the PCR. Generally, the specificity of PCR is controlled by the length of the oligonucleotide and/or the temperature of annealing of the primer to the template. An empirical relationship between oligonucleotide length and ability to support amplification was determined. This relationship allows for the design of specific oligonucleotide primers. A model is proposed which helps explain the observed dependence of PCR on annealing temperature and length of the primer.

Nonextractive spectrofluorimetric determination of aluminium in real, environmental and biological samples using chromotropic acid.

An ultra-sensitive and highly selective nonextractive fluorimetric method is presented for the rapid determination of aluminium at nano-trace levels using chromotropic acid as a fluorimetric reagent [lambda(ex) = 360 nm and lambda(em) = 390 nm] in the pH range of 4.1-4.7. The fluorescence intensity of the metal chelate (2:3 complex) reaches a constant value within 1/2 hr and remains unchanged for over 48 hr. The fluorescence intensity aluminium concentration calibration curve is collinear between 1 and 300 ng/ml of Al. A constant fluorescence intensity is obtained over a wide range (1:50-1:1500) of Al:reagent molar concentrations. Large excesses of over 60 cations, anions and complexing agents (like tartrate, oxalate, phosphate, thio-urea, SCN(-), etc.) do not interfere in the Al determination. The developed method was successfully used in assaying aluminium in several standard reference materials (Al-bronze, brass, stainless steel) as well as in some environmental and biological samples. The method is very precise and accurate (S.D = +/-0.001 on 10 ng/ml; 11 determinations).

Sensitive spectrofluorimetric determination of ruthenium at nanotrace levels using 2-(alpha-pyridyl) thioquinaldinamide [PTQA].

A new spectrofluorimetric method for the determination of ruthenium with nonfluorescent 2-(alpha-pyridyl) thioquinaldinamide (PTQA) is described. The oxidative reaction of Ru(III) upon PTQA gives oxidised fluorescent product (lambda(ex(max))=347 nm; lambda(em(max))=486 nm). The sensitivity of the fluorescence reaction between ruthenium and PTQA is greatly increased in the presence of Fe (III). The reaction is carried out in the acidity range 0.01-0.075 M H(2)SO(4). The influence of reaction variables is discussed. The range of linearity is 1-400 microg l(-1) Ru(III). The standard deviation and relative standard deviation of the developed method are +/-1.210 microg l(-1) Ru (III) and 2.4%, respectively (for 11 replicate determinations of 50 microg l(-1) Ru (III)). The effect of interferences from other metal ions, anions and complexing agents was studied; the masking action is discussed. The developed method has been successfully tested over synthetic mixtures of various base metals and platinum group metals, synthetic mixtures corresponding to osmiridium, certified reference materials in spiked conditions and rock samples.

Spectrophotometric determination of vanadium(v) and its application to vanadium steels containing chromium, molybdenum, tungsten and manganese.

A newly synthesized reagent, N-o-toluoyl-N-o-tolylhydroxylamine is used in a sensitive and selective spectrophotometric method for determination of vanadium(V). The method has been successfully applied to vanadium determination in Mn-Mo-Cr-V steels. The system in 2-6M hydrochloric acid medium obeys Beer's law at 510 nm in the range of vanadium concentration from 0.5 to 10.0 mug ml .

Direct gravimetric determination of molybdenum(VI) in presence of other ions and its application to alloy steel.

N-o-Toluoyl-N-o-tolylhydroxylamine has been synthesized and employed successfully as a gravimetric reagent for direct determination of molybdenum(VI). The dried complex (110-120 degrees ) has definite composition, MoO(2)(C(15)H(14)O(2)N)(2). The method is quite simple, rapid, sensitive and gives reproducible results. The precipitation is quantitative at room temperature over a wide range of acidity. The reagent is highly selective and the method is satisfactory for analysing alloy steels.

Spectrofluorimetric determination of molybdenum in some real and environmental samples.

A very simple, highly-sensitive and selective quenchofluorimetric method for the rapid determination of molybdenum(VI) in aqueous media is described. The method is based on the instantaneous quenching action by the metal-ion upon the native fluorescence of bathophenanthrolinedisulphonate (4,7-diphenyl-1,10-phenanthrolinedisulphonate) solution [lambda(ex) (max) 288 nm; lambda(em) (max) 444.8 nm] in the optimum pH-range of 3.0-3.7 at room temperature (25 +/- 5 degrees ). The fluorescence quenching is co-linear in the range of 0.01-1.0 ppm molybdenum. Large excesses of over 50 cations, anions and some common complexing agents were found to have no interference. Cu, Ni, Co, Fe and V can be tolerated only up to the corresponding amount of molybdenum. Interference from greater amounts can however be removed by a one-step ion-exchange separation process. The developed method was successfully tested over several standard alloys, synthetic mixtures of various compositions, factory effluents and in spiked environmental waters.

Selective gravimetric method for direct determination of tin(IV) with N-o-toluoyl-N-o-tolylhydroxylamine and its application to brass and bronze analyses.

A selective gravimetric method for determination of tin(IV) with N-o-toluoyl-N-o-tolylhydroxylamine by direct weighing as Sn(C(15)H(14)NO(2))(2)Cl(2) is recommended. The metal complex, dried at 110-120 degrees , gives reproducible results. The method is simple, rapid and quantitative over a wide range of acidity (1-4N sulphuric acid) and is superior to existing gravimetric methods. The method works satisfactorily for determining tin in brass and bronze.

Liquid-liquid extraction of osmium thiocyanate with hexamethylphosphoramide and direct spectrophotometric determination in the organic phase.

The use of hexamethylphosphoramide (HMPA) in the osmium-thiocyanate system makes the method more sensitive and selective. The blue colour formed by osmium(VIII) with thiocyanate in 1.5M hydrochloric acid is intensified in the presence of HMPA and the complex becomes readily extractable into chloroform. The colour system has its absorption maximum at 595 nm and obeys Beer's law over the range 0.5-16 mug of Os per ml. The optimum range is 2-10 mug/ml. The molar absorptivity is 2.09 x 10(4)l.mole(-1).cm(-1). The method is simple, sensitive and free from interference from many metal ions, including ruthenium and other platinum metals.

A rapid and selective method for direct gravimetric determination of vanadium(V) and its application to alloy steel and some rocks.

N-o-Iodobenzoyl-o-tolyhydroxylamine has been synthesized and used as gravimetric reagent for direct determination of vanadium(V). The complex dried at 105-120 degrees has the definite composition VO(C(14)H(11)O(2)NI)(2)Cl. The method is simple, rapid, sensitive and selective and gives results reproducible within 0.1%. Precipitation is quantitative at room temperature over a wide range of acidity. The method is satisfactory for alloy steels and some rocks. The very low conversion factor (0.0632) is favourable for determining very small amounts of vanadium.

Visual acuity testing in cataract--an insight (cataract classification density based).

The senile cataracts have been graded on the basis of density objectively. The letter visual acuity, laser interferometric visual acuity and pin hole visual acuity were compared in various grades of cataracts and controls (phakic and aphakic) in 140 eyes. It was found that good correlation exists in all eyes except when cataract density is grade III or IV. The laser interferometry has good prognostic value when the predictability is assessed in early stages of cataract (Grade I & II).

Phosphoproteins: structural components of oncornaviruses.

Oncornaviruses, which contain a virion-associated protein kinase, were found to possess phosphoproteins as virion structural components. One major phosphoprotein common to strains of laboratory and wild mouse oncornaviruses and a strain of feline leukemia virus was shown to be a polypeptide of about 12, 000 mol wt. In addition to this, the Kirsten strain of murine sarcoma virus contained a second major phosphoprotein of about 10, 000 mol wt, and mouse erythroblastosis virus contained a second major phosphoprotein that was either identical to or comigrated with the virion glycoprotein of about 74, 000 mol wt. The major phosphoprotein of RD-114 virus was found to be of about 16, 000 mol wt. The major phosphoamino acid of the 12, 000-mol wt polypeptide of the mouse erythroblastosis virus was identified as phosphoserine, and that of the 16, 000-mol wt polypeptide of the RD-114 virus was identified as phosphothreonine.

Binding characteristics of wild mouse type C virus to mouse spinal cord and spleen cells.

Binding characteristics of mouse spinal cord and spleen cells to naturally occurring, ecotropic (paralytogenic and lymphomagenic 1504M virus) and amphotropic (lymphomagenic 1504A virus) retroviruses of wild mice were investigated. 125I-labeled ecotropic (N-tropic) virus bound efficiently to both spinal cord and spleen cells from SWR/J mice (Fv-1n), but labeled amphotropic (N-tropic) virus bound efficiently only to spleen cells. The extent of binding of 1504M virus to the spinal cord cells was related to the time of incubation and to the amount of labeled input virus. 1504M virus bound to both glial and neuronal subpopulations of the spinal cord with almost equal efficiency. Binding of 125I-labeled 1504M virus to SWR/J mouse spinal cord cells was competitively inhibited by unlabeled homologous virus, whereas an excess of unlabeled 1504A virus inhibited only 10% of the ecotropic virus binding. Unlabeled 1504M virus almost completely inhibited the low-level binding of 125I-labeled 1504A virus to spinal cord cells. The different extent of binding of 1504M virus to spinal cord cells from different strains of mice (SWR/J, NIH Swiss, BALB/c-Jax, Lake Casitas wild, and CD-1) correlated with the susceptibility to paralysis in these strains of mice after inoculation with 1504M virus. Although the spinal cord cells of BALB/c mice contained a moderate amount of 1504M virus receptor sites, these mice were resistant to virus-induced paralysis because of their Fv-1b genotype. Our results indicate that the receptor sites for wild mouse ecotropic retrovirus are strain and organ specific and suggest that the presence of such receptors in central nervous system tissue may be a prerequisite for a successful virus infection and paralysis induction in Fv-1n mice.

Mink type C virus: biochemical characterization of the structural polypeptides.

Mink type C virions contained six major protein species of approximate M.W. of 90,000, 70,000, 30,000, 15,000, 12,000 and 10,000. The two largest polypeptides were glycosylated and the 12,000 M.W. polypeptide was the major phosphoprotein of the virion. Two-dimensional tryptic peptide map of the 30,000 M.W. major structural protein of MiLV showed a pattern distinct from those of analogous proteins from mouse and endogenous cat type C viruses. Significant peptide homology of this protein was, however, found with the corresponding protein of infectious feline type C virus (FeLV).

Characterization of a highly oncogenic murine leukemia virus from wild mice.

We have isolated a highly lymphomagenic wild mouse virus by passage of a weakly oncogenic amphotropic murine leukaemia virus (MuLV-A) in NIH Swiss mice. This virus is similar in host range, interference and neutralization properties to that of the inoculated amphotropic virus but is distinct from it biochemically and causes lymphomas in 90-100% of mice within 1-2 months. Our results indicate that this highly oncogenic virus is a recombinant of wild mouse MuLV-A and sequences related to env gene region of the endogenous xenotropic virus of NIH Swiss mice.

RNA tumor virus phosphoproteins: primary structural analysis and identification of phosphopeptides.

Two-dimensional tryptic peptide mapping was used to compare the peptide sequences of the phosphoprotein (pp12) of cloned ecotropic and amphotropic wild mouse leukemia viruses, strains 1504 and 292. The maps of two ecotropic isolates were very similar to one another, as were the maps of two amphotropic isolates. There was also extensive similarity between the maps of this protein from ecotropic and amphotropic viruses, although characteristic peptide differences were readily recognized. These differences were consistent with the general type specificity of oncovirus phosphoproteins. The pp12 of the field isoalte of 292 virus contained five phosphopeptides, and the non-phosphorylated and variously phosphorylated species of this pp12 showed identical peptide maps, indicating differential phosphorylation of a single polypeptide.

Stereospecific and nonspecific interactions of the morphine congener levorphanol in subcellular fractions of mouse brain.

A METHOD IS DESCRIBED FOR ANALYZING THE ASSOCIATION OF THE OPIATE NARCOTIC LEVORPHANOL WITH BRAIN TISSUE INTO THREE COMPONENTS: nonsaturable, saturable nonspecific, and saturable stereospecific. The method may be of general applicability for the study of the interaction of drugs with body tissues. In mouse brain the stereospecific binding of levorphanol represents only 2% of the total association of drug with tissue, and it was found only in certain membrane fractions. The material responsible for the stereospecific binding might be the opiate receptor.

Allele-specific enzymatic amplification of beta-globin genomic DNA for diagnosis of sickle cell anemia.

A rapid nonradioactive approach to the diagnosis of sickle cell anemia is described based on an allele-specific polymerase chain reaction (ASPCR). This method allows direct detection of the normal or the sickle cell beta-globin allele in genomic DNA without additional steps of probe hybridization, ligation, or restriction enzyme cleavage. Two allele-specific oligonucleotide primers, one specific for the sickle cell allele and one specific for the normal allele, together with another primer complementary to both alleles were used in the polymerase chain reaction with genomic DNA templates. The allele-specific primers differed from each other in their terminal 3' nucleotide. Under the proper annealing temperature and polymerase chain reaction conditions, these primers only directed amplification on their complementary allele. In a single blind study of DNA samples from 12 individuals, this method correctly and unambiguously allowed for the determination of the genotypes with no false negatives or positives. If ASPCR is able to discriminate all allelic variation (both transition and transversion mutations), this method has the potential to be a powerful approach for genetic disease diagnosis, carrier screening, HLA typing, human gene mapping, forensics, and paternity testing.

Comparative studies on the structural phosphoproteins of mammalian type C viruses.

The major phosphoprotein common to woolly monkey sarcoma virus, gibbon ape lymphosarcoma virus, and type C viruses of the lower mammalian species (mouse, rat, cat), with the exception of the endogenous cat virus (RD-114), is the polypeptide of about 12,000 molecular weight. The protein-phosphate bond in this polypeptide of several viruses is of the phosphoserine variety excepting gibbon ape virus, which contains both phosphoserine and phosphothreonine. The primary phosphoprotein of RD-114 virus and the endogenous baboon type C virus, on the other hand, is the polypeptide of about 15,000 molecular weight which contains phosphothreonine as its phosphoamino acid. A second major phosphoprotein of molecular weight of 10,000 is detected only in viruses genetically related to rat species including those derived from the RPL cell line, from Sprague-Dawley rat embryo cells, and the Kirsten mouse sarcoma virus which was recovered from a mouse erythroblastosis virus after in vivo propagation through rat. These phosphorylated polypeptides of molecular weight 15,000, 12,000, or 10,000 are present in the virion structure in several different but nonrandom phosphorylated states.

Biochemical properties of wild mouse oncornaviruses with lymphomagenic and neurotropic activities.

The close immunologic relation between the group-specific and polymerase proteins of the wild mouse derived and the established strains of mouse type C oncornaviruses, and the lack of any unusual structural polypeptide or RNA components in the wild mouse virions, eliminate the possibility of detectable contamination by a class of virus different from the mouse oncornavirus class in the wild mouse virus preparations. Liquid hybridization studies with a wild mouse viral 70S 3H-RNA and cellular DNA under conditions of DNA excess, suggest that a significant fraction but not all of the virus-specific nucleotide sequences is present in all normal and tumored wild mouse tissues tested. These virus related sequences may possibly be attributed to a hypothetical endogenous inherited type C virus genome(s) carried by all wild mice or to an infection by one or more but not all of the different exogenous strains of wild mouse type C viruses which could possibly be present in the virus preparation used. The findings are consistent with wild mouse derived type C viruses being either entirely exogenous or a mixture of endogenous and exogenous viruses. This interpretation is also consistent with the earlier observation that certain wild mouse type C viruses are exogenously transmitted, transplacentally, and via milk. The possible relation of the virus heterogeneity or the distinct characteristics of the virus surface molecules to the diverse pathogenicity of the wild mouse oncornaviruses is discussed.