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PubMed1991

Rat obesity gene fatty (fa) maps to chromosome 5: evidence for homology with the mouse gene diabetes (db).

The autosomal recessive mutations fa (rat) and db (mouse) cause obesity syndromes that develop early and ultimately become severe. Although both fa/fa rats and db/db mice have been studied extensively as models of human obesity and diabetes, the molecular bases of these phenotypes remain unknown. We have mapped fa in 50 fa/fa (obese) offspring of a (13M x Brown Norway) F1 fa/+ intercross relative to two molecular markers, Ifa and Glut-1, which flank db on mouse chromosome 4 and which are located on rat chromosome 5. Ifa and Glut-1 are linked to fa, with a gene order, Ifa-fa-Glut-1, that is identical to that for the region around db in the mouse genome. These results place fa on rat chromosome 5 and suggest that db and fa are mutations in homologous genes.

Animals↗
PubMed2002

In vitro effects of incinerator fly ash on pulmonary macrophages and epithelial cells.

Fly ash from a municipal waste incinerator was used as a model for atmospheric particles in order to identify parameters relevant for the induction of adverse health effects. The aim of this study was to compare the biological effects of the total incinerator fly ash (IFA), the soluble and the insoluble fraction with the effects of quartz by in vitro toxicity studies. The previously sized fly ash (< 20 microns) was characterized by elemental composition and particle size distribution. The particles were administered to rat alveolar macrophages (NR8383) and human bronchial epithelial cells (BEAS-2B) at different amounts via the medium. The total IFA and its insoluble fraction were shown to induce cytotoxicity and cytokine release in a dose-dependent manner. The soluble fraction was nearly unable to induce cytotoxicity and TNF-alpha release but showed potent induction of IL-8 release in BEAS-2B cells at increasing concentrations. Quartz caused similar effects compared to IFA in NR8383 but was less effective in BEAS-2B.

Air Pollutants↗
PubMed1979

Abnormal haemoglobins in the Sudan savanna of Nigeria. II. Immunological response to malaria in normals and subjects with sickle cell trait.

Children born in areas hyperendemic for Plasmodium falciparum are protected by maternal antibodies for up to about five months of life, after which they are subject to intense infection until they acquire sufficient immunity--by about five years of age. Children with sickle cell trait (Hb.AS) are at an advantage during these critical years, probably because of preferential phagocytosis of parasitized red cells. This could lead to either (i) early processing of antigen by macrophages and an accelerated immune response, or (ii) less antigenic stimulus and hence lower antibody production. Immunoglobulin (Ig)G and IgM determinations, agar gel diffusion (Ouchterlony) against soluble P. falciparum antigen, the indirect fluorescent antibody (IFA) test using P. falciparum and P. malariae antigens, and the indirect haemagglutination (IHA) test with P. falciparum antigen were performed on sera from a population with different Hb electrophoretic types in the hyperendemic malarial area of Garki, Kano State, Nigeria. Plasma immunoglobulins and antimalarial antibodies rose with age. After the first year of life, lower mean concentrations of immunoglobulins (especially IgM), and lower mean titres of antibodies specific against P. falciparum (Ouchterlony, IHA and less significantly IFA) were present in Hb.AS compared to Hb.AA; these differences increased with age. Antimalarial intervention was followed by a decline of all values and final levels showed little difference between haemoglobin types. It was unlikely that either a relative inability to produce antibody or a more rapid catabolism of immunoglobulins was responsible for the lower levels in sickle cell trait. The observations are more easily explained by the hypothesis that Hb.AS persons have less antigenic stimulus due to the early removal of parasitized sickled cells by macrophages, which then degrade the antigens. The antibody difference between Hb.AA and Hb.AS increased throughout life, suggesting that this process remained a feature of sickle cell trait even after parasite frequencies and densities were similar in the two Hb groups. These observations have implications in the aetiology of tropical splenomegaly syndrome, which is rarely seen in sickle cell trait subjects. Mean IgG and IgM were slightly higher in Hb.AS than Hb.AA infants, the difference for IgG achieving significance. This suggested that during infancy early phagocytosis of parasitized cells had led to enhanced processing of antigen and hence an earlier immune response in Hb.AS, but this was unlikely to be a major factor in survival. IFA titres against P. malariae were slightly but not significantly lower in Hb.AS, possibly as a result of cross-reaction with P. falciparum antibody or of a slight degree of protection against P. malariae.

Adolescent↗
PubMed1987

Malaria chemoprophylaxis to pregnant women provided by community health workers in Saradidi, Kenya. III. Serologic studies.

Parasitaemia and antimalarial antibodies were examined from May 1983 to March 1984 in monthly samples taken from 930 pregnant women attending antenatal clinics in Saradidi, Kenya, and 317 of their infants; 104 women were taking chloroquine phosphate 300 mg base weekly for chemoprophylaxis. Seropositivity rates in pregnant women were uniformly high, and mean enzyme-linked immunosorbent assay (ELISA) absorbance values were not related to presence of parasitaemia or history of chemoprophylaxis. Parasitaemia was present in 26.5% of 1677 slides from pregnant women and there was little variation by month of sample. Mean ELISA absorbance values varied by month of sample. Seropositivity rates in infants were high as measured in both the indirect fluorescent antibody (IFA) test (81.6% of 938) and ELISA at 1:100 (83.8% of 1025) and 1:1000 (34.8% of 1025) serum dilutions. Seropositivity rates decreased slightly after birth but by four months of age rates were again high. Parasitaemia was present in 26.5% of 1677 slides from pregnant women. Paired comparisons were made on maternal samples collected less than two months before parturition and samples from the infants collected within two months after birth. The paired antibody response by IFA or ELISA was not dependent on the presence of detectable parasitaemia in the mother. Infants from mothers with a history of antimalarial chemoprophylaxis had significantly (P = 0.04) lower IFA titres than other infants. Measuring the absorbance of a 1:100 serum dilution by ELISA appeared to be an excellent method with which to measure longitudinal serologic changes in a population.

Animals↗
PubMed1999

Serologic response to Bartonella henselae in patients with cat scratch disease and in sick and healthy children.

Indirect fluorescent antibody assay (IFA) is the most reliable test for detecting antibody to Bartonella henselae in the diagnosis of cat scratch disease (CSD). Recently, an ELISA test has been proposed, but conflicting results are reported. We compared IgG-IFA and IgG-IgM ELISA methods in CSD patients and in healthy children. We also tested ELISA specificity in a large group of healthy controls and in children with lymphoma-associated lymphadenopathy and with pyogenic lymphadenitis. The ELISA procedure was positive in 69/78 patients with CSD (sensitivity 89.6%), in 5/100 healthy children (specificity 95%), in 2/51 patients with non-Hodgkin's lymphoma or pyogenic lymphadenitis (specificity 96%) and in 27/296 blood donors (specificity 91.6%). In 34 patients with CSD, ELISA IgM and IgG responses decreased significantly between time of diagnosis of the disease and recovery. We found significantly higher IgG-ELISA titres in cat-owners, whether blood donors or healthy children, than in non-cat-owners. The IgG-IFA test gave positive results in 69/78 patients with CSD (sensitivity 89.6%) and in 5/62 healthy controls (specificity 92.5%). The ELISA method is a cheap, sensitive method for determining antibody response to Bartonella henselae infection and is also important for evaluating the clinical course of the disease and the efficacy of antibiotic therapy. The high specificity of ELISA in patients with non-Hodgkin's lymphoma will help the clinician to exclude a potentially life-threatening disease associated with lymphadenopathy.

Adolescent↗
PubMed2002

Inhibitory effects of incomplete Freund's adjuvant on experimental autoimmune encephalomyelitis.

Freund's incomplete adjuvant (IFA), an aqueous/oil emulsion that is widely used in combination with antigenic proteins and peptides to induce tolerance, is considered to be immunologically inert. However, sporadic reports indicate that IFA may itself have inhibitory properties on induction of adjuvant induced arthritis and spontaneous diabetes. In the current study, the effects of IFA/saline were evaluated on the induction of experimental autoimmune encephalomyelitis (EAE) in three different strains of mice. IFA/saline given i.p. in two doses of > 100 microl 10 days apart were found to inhibit EAE induction to varying degrees in all three strains of mice in a dose dependent fashion. The IFA/saline injections inhibited both mitogen and antigen-induced T cell proliferation, induced elevated secretion of IFN-gamma and IL-10 by neuroantigen specific T cells, and reduced expression of cytokines, chemokines, and chemokine receptors of CNS-infiltrating mononuclear cells. These data demonstrate for the first time a direct inhibitory effect of IFA/saline on EAE, and re-emphasize the need to properly control experiments using IFA to induce antigen-specific tolerance.

Animals↗
PubMed1999

Inhibitory effect of ferulic acid and isoferulic acid on the production of macrophage inflammatory protein-2 in response to respiratory syncytial virus infection in RAW264.7 cells.

We investigated the effect of ferulic acid (FA) and isoferulic acid (IFA), which are the main active components of the rhizoma of Cimicifuga heracleifolia (CH), an anti-inflammatory drug used frequently in Japanese traditional medicine, on the production of macrophage inflammatory protein-2 (MIR-2) in a murine macrophage cell line, RAW264.7, in response to respiratory syncytial virus (RSV) infection. Following the exposure of cells to RSV for 20h, the MIP-2 level in condition medium was increased to about 20 ng/ml, although this level in mock-infected cells was negligible. In the presence of either FA or IFA, RSV-infected cells reduced MIP-2 production in a dose-dependent manner. These data suggest that FA and IFA might be responsible, at least in part, for the anti-inflammatory drug effect of CH extract through the inhibition of MIP-2 production.

Animals↗
PubMed1985

Intermediate filaments in non-neuronal cells of invertebrates: isolation and biochemical characterization of intermediate filaments from the esophageal epithelium of the mollusc Helix pomatia.

To screen invertebrate tissues for the possible expression of intermediate filaments (IFs), immunofluorescence microscopy with the monoclonal antibody anti-IFA known to detect all mammalian IF proteins was used (Pruss, R. M., R. Mirsky, M. C. Raff, R. Thorpe, A. J. Dowding, and B. H. Anderton. 1981. Cell, 27:419-428). In a limited survey, the lower chordate Branchiostoma as well as the invertebrates Arenicola, Lumbricus, Ascaris, and Helix pomatia revealed a positive reaction primarily on epithelia and on nerves, whereas certain other invertebrates appeared negative. To assess the nature of the positive reaction, Helix pomatia was used since a variety of epithelia was strongly stained by anti-IFA. Fixation-extraction procedures were developed that preserve in electron micrographs of esophagus impressive arrays of IFs as tonofilament bundles. Fractionation procedures performed on single cell preparations document large meshworks of long and curvilinear IF by negative stain. These structures can be purified. One- and two-dimensional gels show three components, all of which are recognized by anti-IFA in immunoblotting: 66 kD/pl 6.35, 53 kD/pl 6.05, and 52 kD/pl 5.95. The molar ratio between the larger and more basic polypeptide and the sum of the two more acidic forms is close to 1. After solubilization in 8.5 M urea, in vitro filament reconstitution is induced when urea is removed by dialysis against 2-50 mM Tris buffer at pH 7.8. The reconstituted filaments contain all three polypeptides. The results establish firmly the existence of invertebrate IFs outside neurones and demonstrate that the esophagus of Helix pomatia displays IFs which in line with the epithelial morphology of the tissue could be related to keratin IF of vertebrates.

Amino Acids↗
PubMed1975

Induction of T-lymphocyte responses to a small molecular weight antigen. II. specific tolerance induced in azebenzenearsonate (ABA)-specific T cells in Guniea pigs by administration of low doses of an ABA conjugate of chloroacetyl tyrosine in incomplete Freund's adjuvant.

The experiments presented in this paper demonstrate that the induction of tolerance on the one hand and the induction of delayed sensitivity on the other hand can be accomplished by administration of similar doses of azobenzene-arsonate conjugated to N-chloracetyl tyrosine (ABA-T) to guinea pigs with the determining factor being the absence or presence, respectively, of activating bacterial products in the adjuvant mixture used. Thus, complete, persistent ABA-T-specific T-cell tolerance can be induced in adult guinea pigs with 20 mug of ABA-T given intradermally in incomplete Freund's adjuvant (IFA) whereas this same dose of ABA-T induces ABA-specific immunity when administered in complete Freund's adjuvant. This tolerance was not reversible by administration of ABA-T and IFA in the presence of bacterial lipopolysaccharide, was generated before the formation of primed T cells, and persisted for at least 3 mo after initiation. Moreover, cell transfer studies performed herein demonstrate that the unresponsiveness resulting from administration of ABA-T in IFA reflects the activity of suppressor cells to induce and maintain a state of unresponsiveness could only be demonstrated in unprimed animals may indicate a severe limitation on the potential clinical usefulness of such an approach to regulation of the immune system.

Acetates↗
PubMed1988

Allelic forms of gp195, a major blood-stage antigen of Plasmodium falciparum, are expressed in liver stages.

Mature exoerythrocytic (EE) forms of two cloned lines (3D7 and HB3) of Plasmodium falciparum were obtained in the livers of splenectomized chimpanzees. Sectioned preparations were examined by immunofluorescence (IFA) using mAbs that distinguished allelic variants of the blood-form antigen gp195 and mAbs that recognized multiple conserved epitopes of gp195. EE forms and blood schizonts exhibited identical IFA reactions for each respective clone, showing that the antigen was expressed identically in liver and blood-stage parasites. A third chimpanzee was infected with sporozoites derived from a mixture of 3D7 and HB3 gametocytes that had undergone cross-fertilization in the mosquitoes. IFAs on the EE forms in this animal showed that segregation of each gp195 allele had occurred earlier in the life cycle, providing evidence that the parasite is haploid for the whole of its mammalian development.

Alleles↗
PubMed1998

Reactivity of human sera to different strains of granulocytic ehrlichiae in immunodiagnostic assays.

Sera from 35 patients diagnosed with human granulocytic ehrlichiosis in Connecticut were tested by indirect IFA staining methods with 5 strains of Ehrlichia equi or the human granulocytic ehrlichiosis agent to assess the suitability of different strains in laboratory analyses. Antigens included horse-derived infected neutrophils (MRK and BDS strains) and human isolates cultured in human promyelocytic leukemia cells (NCH-1, RCH, and Webster). Of 35 sera, 23 (65.7%) reacted to all 5 strains. Seropositivity was highest (97.1%) in assays that contained the MRK strain from California and lowest (71. 4%) in tests with the NCH-1 strain from Nantucket, Massachusetts. In parallel testing of 32 sera with the NCH-1 strain by indirect IFA and Western blot analyses, results were concordant for 30 samples (93.8%). All strains of ehrlichiae can be used in IFA analyses for antibody detection, but assay sensitivity varied with the strain used.

Animals↗
PubMed1998

Seroprevalence of human granulocytic ehrlichiosis among permanent residents of northwestern Wisconsin.

Four-hundred seventy-five permanent residents of Wisconsin were tested for antibodies to the agent of human granulocytic ehrlichiosis (HGE) by indirect immunofluorescent antibody (IFA) testing with Ehrlichia equi as antigen marker. Each resident completed a standard survey questionnaire about outdoor activities, animal and tick exposure, and any febrile illness during the preceding 12 months. Seventy-one serum samples (14.9%) contained E. equi antibodies. The mean IFA titer for seropositive residents was 250 (range, 80-10,240). Seropositive residents were older than seronegative ones (62 vs. 56 years; P = .019). None of the seropositive residents had a history suggestive of ehrlichiosis. There was no association between the IFA test outcome and specific demographic variables or history of tick bites. HGE appears to be a common subclinical or mild infection among residents in northwestern Wisconsin.

Adolescent↗
PubMed2001

Infection and transovarial transmission of rickettsiae in Dermacentor variabilis ticks acquired by artificial feeding.

In this study we examined the efficiency of an in vitro feeding technique using glass microcapillaries as a method of establishing rickettsiae-infected lines of ticks. To quantify the volume ingested by ticks during microcapillary feeding, the incorporation of radiolabeled amino acids in tick gut and hemolymph was calculated. Fifteen of 18 ticks consumed between 0.06 and 6.77 microl. However, ingestion of fluid was not correlated to weight gain during capillary feeding. Uninfected and partially fed laboratory-reared female Dermacentor variabilis ticks were exposed to either Rickettsia montana- or Rickettsia rhipicephali-infected Vero cells via microcapillary tubes, returned to rabbit hosts, and allowed to feed to repletion. All tissues collected from ticks allowed to feed overnight on rickettsiae-infected fluids were found to be infected when examined by IFA. When rickettsiae-infected and uninfected capillary-fed ticks were allowed to feed to repletion and lay eggs, no significant differences in mean engorgement weight or fecundity was observed. When we assessed the efficiency of transovarial transmission of rickettsiae by ticks that imbibed rickettsiae-infected cells by polymerase chain reaction (PCR) and IFA, infection was detected by PCR in the eggs from 85% of the ticks exposed to R. montana and 69% of the ticks exposed to R. rhipicephali. Rickettsial genes were not amplified in samples of the uninfected controls. Examination by IFA of egg samples from females exposed to rickettsiae-infected cells identified rickettsiae in 100% of the samples tested, while the uninfected controls were negative.

Amino Acids↗
PubMed1991

Vaccine protection against HIV-2 infection in cynomolgus monkeys.

The aim of this study was to determine if protection against an infectious human immunodeficiency virus type 2 (HIV-2) challenge could be obtained in cynomolgus macaques by active immunization using whole killed virus vaccine. Four monkeys were immunized with killed HIV-2SBL-6669, two of them with five intramuscular (im) injections of viral preparation containing 100 or 300 micrograms protein emulsified in incomplete Freund's adjuvant (IFA) and the two remaining received four im injections of 25-50 micrograms viral protein in iscoms. Each of the four vaccinated cynomolgus monkeys, along with four unvaccinated controls, were challenged intravenously two weeks after the last booster with approximately 100 animal infectious doses (ID50) of live HIV-2SBL-6669. All four immunized monkeys developed antibodies to HIV-2 envelope and core proteins before challenge exposure to HIV-2, but only the two animals vaccinated with virus in IFA developed detectable neutralizing antibodies. The two monkeys immunized with killed virus in IFA have shown no evidence of infection nine months after challenge with live virus. When blood and lymph node cells from these animals were transfused into naive cynomolgus monkeys, the recipients remained free of infection. In contrast, virus was recovered repeatedly in all nonimmunized animals and in the two animals immunized with iscom-associated viral antigens, which had a low content of envelope gp125 antigen. The demonstration of vaccine-induced protection against HIV-2 in a nonhuman primate raises hope for effective immunization against HIV infections in humans as well.

Animals↗
PubMed1996

Immune response to 17 beta-estradiol in polyelectrolyte complex: antigen specificity and affinity of hybridoma clones.

Complex formation between synthetic polyelectrolytes (PE) [poly-4-vinyl-N-ethyl (cetyl), pyridine bromides-PVP(R2, R16)], bovine serum albumin (BSA), or 17 beta-estradiol-BSA conjugate (BSA.E) was studied in neutral water. Weakly water-soluble (colloidal) complex was formed upon addition of BSA.E to PVP (R2, R16) solution at pH 7. A nonrandom distribution of the protein molecules between the coils of polycations and self-assembly in the nonstochiometric polycomplex particles took place. The immunogenic properties of PVP (R2, R16)-BSA.E polycomplex were investigated and the specificities of produced antibodies analyzed. 17 beta-Estradiol introduced in polyelectrolyte complexes (PE-BSA) was found to invoke considerable increases in the steroid-specific immunoresponse. However, a comparative study of immunogenic activity of polycomplexes versus BSA.E+incomplete Freund's adjuvant (IFA) mixtures revealed some differences in regards to the specificity of antibody production. In contrast to IFA+BSA.E systems, polycomplexes were able to generate estradiol-as well as BSA-specific antibodies. Such a carrier-directed response may be determined by increase in immunogenicity of weak antigenic determinants and/or by the exposure of internally located determinants upon complex formation with polyelectrolytes. Fusions following the two different immunization procedures resulted in the growth of comparable numbers of estradiol-specific monoclonal antibodies with apparently similar antigen affinities. Thus, immunizations using antigens in PEC appear to provide an efficient alternative to IFA.

Animals↗
PubMed2005

Efficacy of twice-weekly multiple micronutrient supplementation for improving the hemoglobin and micronutrient status of anemic adolescent schoolgirls in Bangladesh.

BACKGROUND: Although iron deficiency is a major cause of anemia, other micronutrient deficiencies may also play a role. OBJECTIVE: We examined whether multiple micronutrient supplementation is more efficacious than is supplementation with iron and folic acid alone for improving the hemoglobin and iron status of anemic adolescent girls in Bangladesh. DESIGN: Anemic (hemoglobin < 12.0 g/dL) girls (n = 197) aged 14-18 y from rural schools in Dhaka District were entered into a randomized double-blind trial and received twice-weekly supplements of iron and folic acid (IFA group) or multiple micronutrients (15 micronutrients, including iron and folic acid; MMN group) for 12 wk. RESULTS: At recruitment, the characteristics of the girls in the 2 groups were not significantly different, except for family size and body mass index. At the end of the study, although both groups benefited significantly from supplementation, mean changes in hemoglobin and serum ferritin concentrations were not significantly different between groups. Compared with the IFA group, girls in the MMN group had significantly greater increases in mean serum vitamin A, plasma vitamin C, red blood cell folic acid, and riboflavin concentrations (assessed as erythrocyte glutathione reductase activation coefficient). After 12 wk of supplementation, only the prevalence of vitamins A and C and riboflavin deficiencies decreased more significantly in the MMN group than in the IFA group. CONCLUSIONS: Twice-weekly MMN supplementation for 12 wk significantly improved the status of the micronutrients assessed but was not more efficacious than was supplementation with iron and folic acid alone in improving the hematologic status of anemic adolescent girls. More frequent doses may be needed to achieve full benefit.

Adolescent↗
PubMed1997

Immunologic response to Bartonella henselae as determined by enzyme immunoassay and Western blot analysis.

Bartonella henselae is now regarded as the etiologic agent of cat-scratch disease and a cause of bacillary angiomatosis. We examined the human immune response to Bartonella henselae infection using a newly developed enzyme immunoassay (EIA) and a Western blot procedure using outer-membrane proteins. The EIA showed 98.6% and 91.4% agreement with an indirect fluorescence method (IFA) for detection of IgM and IgG antibodies, respectively. By using Western blot analysis, reactivity to an 8-kd band showed significant correlation with positive results by the IgM IFA and EIA. In contrast, reactivity to 209-, 208.5-, 208-, 116-, and 80-kd bands was identified only in positive IgG IFA serum samples. The EIA and Western blot should be useful tests in determining the antibody response to B. henselae infection and may also be important in determining the critical epitopes in the host-parasite interaction of this organism.

Adolescent↗
PubMed1985

Serologic diagnosis of toxoplasmosis with emphasis on the detection of Toxoplasma-specific immunoglobulin M antibodies.

Commercially available kits were used for the detection of Toxoplasma-specific IgM antibodies. False positive IgM results were observed in whole sera containing Toxoplasma-specific antibodies together with rheumatoid factor when tested by immunofluorescence (IFA). False negative IgM results occurred in whole sera containing competing levels of Toxoplasma-specific IgG antibodies, as indicated by the IFA:IgM-M ratio. False positive and false negative IgM results often occurred when whole sera were tested. These false reactions were eliminated by fractioning IgM from IgG using the Isolab's IgM Isolation System. All five sera in this study with an IFA titer to Toxoplasma of greater than or equal to 1:16,384 also contained Toxoplasma-specific IgM antibodies. This suggests that sera with high titers to Toxoplasma should be tested for Toxoplasma-specific IgM antibodies.

Antibody Specificity↗