Neutralizing antibodies and the course of HIV-induced disease.

The capacity to neutralize the human immunodeficiency virus (HIV) in vitro was examined in 52 sera obtained from 23 seropositive individuals in addition to 7 negative control sera. Neutralization was measured as the activity of a serum to protect MT-4 cells against the cytopathic effect of HTLV-IIIB. Virus neutralization depended on HIV antibodies. Some sera had HIV neutralizing antibody titers of several thousands. All serum samples had been titrated in two ELISAs based either on disrupted HTLV-IIIB or on a bacterially synthesized polypeptide (ENV-80) of gp41 as a test antigen. The correlation of neutralizing activity of the sera with ELISA titers was low. A correlation of serum neutralizing titers with the stage of the disease could not be observed. However, in a longitudinal study with 6 patients over up to 22 months an increase in neutralizing antibodies seemed to protect against progression of the disease. The implications of these findings for antibody treatment and vaccine development are discussed.

Multiple sclerosis and human T-cell lymphotropic retroviruses: negative serological results in 135 German patients.

A total of 135 sera and 18 cerebrospinal fluid (CSF) samples from patients with multiple sclerosis (MS) were screened for antibodies to human T-cell lymphotropic virus type I (HTLV-I) and human immunodeficiency virus (HIV) by ELISA tests. None of the sera or CSF reacted with HIV antigens. Only 3 out of 135 MS sera but no MS CSF showed increased reactions in the ELISA test for HTLV-I. However, these positive reactions were classified as non-specific by immunoprecipitation. Thus no serological evidence for infection with HIV, HTLV-I, or a related retrovirus was found in the MS patients.

Capability of ICP-MS (pneumatic nebulization and ETV) for Pt-analysis in different matrices at ecologically relevant concentrations.

ICP-MS both with conventional nebulization and with ETV (Electro Thermal Vaporization) have been applied for the determination of Pt in different matrices, e.g. occupational samples like urine and dust samples. It can be used also for other matrices like soil, plants, tissues etc. dependent on the concentration ranges and on a suitable decomposition method. The evaluation, based on the different Pt-isotopes and the quality criteria (detection limit, precision, accuracy) is discussed. Very low determination limits in the range of 1 ng/l can be achieved by ICP-MS-ETV using the standard addition method. This method allows to determine Pt in urine without any sample pretreatment.

Total digestion of silicate containing matrices (plants, soil, sludges) using a pressure ashing device with PFA-vessels.

A digestion procedure for silicate containing matrices like soil, sludge, etc. by using a commercially available pressure ashing device (Seif-Aufschlubetatechnik, Germany) with PFA (perfluoroalkoxy) sample vessels is described at the results for different matrix- and trace elements in different suitable certified standard reference materials. The advantages of this technique compared with the use of Teflon(R)-(PTFE, polytetrafluoroethylene) vessels and the open treatment in Pt-crucibles will be discussed.

Biokinetic studies in humans with stable isotopes as tracers. Part 1: A methodology for incorporation of trace metals into vegetables.

The metabolism and biokinetics of trace metals in humans can be successfully studied employing stable isotopes of the investigated elements as tracers. For the estimation of the bioavailability and the intestinal absorption from solid food, materials are required which have been intrinsically labelled with the chosen stable tracer, since the use of an extrinsic label may lead to erroneous results. Here a technique for producing intrinsically labelled vegetables is presented and optimized with regard to molybdenum, gadolinium and ruthenium, elements of interest in the field of radiation protection and/or nutrition. These feasibility studies were aimed to determine the most favourable conditions for the production of vegetables containing the selected tracers in amounts high enough to enable successful biokinetic studies in humans. In this optimization study the natural elements were used instead of the more expensive stable isotopes. Mo is readily absorbed both into cress (Lepidium sativum) and into french beans (Phaseolus vulg. var. nanus). Gd uptake into cress is moderate, while Ru may be easily and successfully incorporated only into sprouts of mung beans (Vigna radiata).

Orthopithonella collaris sp. nov., a new calcareous dinoflagellate cyst from the K/T boundary (Fish Clay, Stevns Klint/Denmark).

A new calcareous dinoflagellate cyst species, Orthopithonella collaris sp. nov., is described from the Cretaceous/Tertiary (K/T) boundary clay (Fish Clay) of Stevns Klint, Denmark, on the basis of SEM studies and light-microscopic analyses of thin sections of single specimens. The species has been found exclusively in the Fish Clay and as such may be a potential marker for the K/T boundary. Its pulse-like occurrence is thought to be due to the abrupt, relatively short-term ecological catastrophe associated with the K/T boundary event.

Application of ICP-MS for the assessment of thorium excretion in urine.

A method for rapid and sensitive determination of thorium in urine by inductively coupled plasma mass spectometry (ICP-MS) is described. The method is sufficiently sensitive to detect 1 ng/L 232Th in urine without any sample preparation. The mean urinary 232Th excretion in 23 unexposed subjects was 6.2 +/- 3.3 ng/d.

Thorium and uranium contents in human urine: influence of age and residential area.

Inductively coupled plasma mass spectrometry (ICP-MS) has been used for the determination of (232)Th and (238)U in urine of unexposed Jordanian subjects living in six cities. The range of (232)Th excretion in all subjects was found to be 1.4-640 microBq d(-1) with an average of 34.8 microBq d(-1) (geometric mean 15.8 microBq d(-1)). Results showed no statistically significant correlation with age and residential area. The average value obtained is in agreement with levels considered normal in some recent publications. The average value of (238)U in all samples was found to be 3955 microBq d(-1) (geometric mean 1107 microBq d(-1)), which is higher than reported figures from Germany and India, but in agreement with those figures given in ICRP publication, number 23. The mean values of the different groups were found to be proportional to age up to 60 years. A noticeable drop is observed for subjects greater than 60 years old.

A blotting procedure which preserves specific protein-DNA interactions.

A quick, quantitative, and nonselective electrophoretic transfer of proteins from acetic acid-urea gels onto nitrocellulose, which preserves their ability to interact specifically with DNA, is achieved when exposure to dodecyl sulfate ions is avoided and a special type of nitrocellulose is used which contains cellulose phosphate ester. Filter-adsorbed histone H1 and other nuclear proteins from an insect, Chironomus thummi, were tested for binding of an AT-rich DNA sequence from the heterochromatin of the same organism under competitive conditions. On the blots, histone H1 exhibited the dependency of DNA binding on NaCl concentration and the preference for AT-rich DNA or poly[d(A-T)] found in quantitative filter-binding studies. By stepwise alteration of the NaCl molarity and competing Escherichia coli DNA concentration, respectively, in the binding buffer, two minor protein fractions could be identified in the heterogeneous extracts, one of which bound preferentially to AT-rich DNA, and the other bound to this sequence at up to 500 mM NaCl. Exposure to dodecyl sulfate led to a disappearance of the ability of these proteins to interact specifically with DNA. While nondenaturing transfer by diffusion (L. Levinger and A. Varshavsky (1982) Proc. Natl. Acad. Sci. USA 79, 7152) is a procedure that requires about 2 days, the present technique of gentle protein transfer for DNA binding studies requires only 2 to 3 h.

The determination of metals (antimony, bismuth, lead, cadmium, mercury, palladium, platinum, tellurium, thallium, tin and tungsten) in urine samples by inductively coupled plasma-mass spectrometry.

OBJECTIVE: An analytical method has been established to determine the concentration of antimony (Sb), bismuth (Bi), lead (Pb), cadmium (Cd), mercury (Hg), Palladium (Pd), platinum (Pt), tellurium (Te), tin (Sn), thallium (Tl) and tungsten (W) in urine. The aim was to develop a method which is equally suitable for the determination of environmentally as well as occupationally caused metal excretion. METHODS: Inductively coupled plasma-mass spectroscopy (ICP-MS) was used for the determination of metals. Calibration was done using aqueous solutions and standard addition respectively. RESULTS: Urine samples of 14 persons occupationally non-exposed to metals were analysed. With the exception of Pt and Bi all the metals were found in these urine samples. The detection limits for these metals lie between 5 and 50 ng/l. CONCLUSIONS: For some metals, which are important from an occupational as well as an environmental viewpoint, ICP-MS is more sensitive than atomic absorption spectrometry (AAS). ICP-MS, moreover, is welcome as a reference method for AAS with the additional advantage of multi-element measurement.

Efficient replication of HTLV-III and STLV-IIImac in human Jurkat cells.

High producer cell lines were established by infecting Jurkat cells with either HTLV-IIIB from H9 cells or with STLV-IIImac from HUT-78 cells. The Jurkat cells produced both viruses at least 10 times more efficiently then the original cell lines; the pelleted virus from Jurkat cells was also less contaminated with non-virion proteins. Accordingly, a higher specificity was attained in an ELISA for antibodies to HTLV-III if the antigenic activity was derived from the virus from Jurkat cells, as opposed to that from H9 cells.

Antibodies to human immunodeficiency virus (HTLV-III/LAV) in people from Lagos and Cross River States of Nigeria.

Using enzyme-linked immunosorbent assay (ELISA) and confirmatory immunoprecipitation tests, sera from 640 Nigerians from Lagos and Cross River States were examined for antibodies against HIV. These comprised 570 blood donors and their family members, 56 patients with various haematological conditions and 14 patients with acute Plasmodium falciparum infection. None of the sera was positive for HTLV-III/LAV antibodies by immunoprecipitation, although 12 (1.9%) sera were positive by ELISA.

Seroepidemiology of human immunodeficiency virus in Africa.

Serum samples from 6015 African subjects without symptoms of the acquired immune deficiency syndrome (AIDS) or contact with the disease were examined for antibodies to the human immunodeficiency virus by a combination of an enzyme linked immunosorbent assay and radioimmunoprecipitation (2567 samples) or by immunofluorescence (3448 samples). Serum samples had been collected between 1976 and 1984 in Senegal (n = 789), Liberia (935), Ivory Coast (1195), Burkina Faso (299), Nigeria (536), Gabon (1649), Zaire (15), Uganda (164), and Kenya (433). Only four samples contained antibodies. Three of these were from attenders at the Lambarene clinic in Gabon and one from a villager in Senegal. By contrast, two out of six AIDS suspects from Guinea-Bissau, all 13 patients with AIDS from Kinshasa (Zaire), and two out of three of their contacts were seropositive, all these specimens having been collected in 1985. These data show that fewer than one in a 1000 subjects were seropositive for AIDS at the time of sampling before 1985 and do not support the hypothesis of the disease originating in Africa.

Correlation of immune defects in hemophilia with HTLV-III antibody titers.

Of 170 hemophilia patients, 22% had high-titer, 29% had low-titer, and 49% had no antibodies against HTLV-III. The strength of HTLV-III antibodies was correlated significantly with a decreased OKT4/T8 ratio (p less than 0.0005), decreased in vitro response to pokeweed mitogen (p less than 0.025), and elevated serum neopterin (p less than 0.05) and serum IgG (p less than 0.0005). The fraction of patients with abnormal immunological findings was consistently greater among patients with high-titer than among patients with low-titer HTLV-III antibodies. Testing these immunological parameters may be useful for monitoring the breakdown of immune functions leading to AIDS.

A new ELISA test for HIV antibodies using a bacterially produced viral env gene product.

Screening tests for antibodies to the human immunodeficiency virus (HIV), based on the indirect ELISA principle using viral preparations as antigen, yield a substantial number of false-positive and false-negative results. These failures are due to the lack of certain viral polypeptides or contaminating cellular polypeptides in viral preparations. Therefore, the accuracy of the screening tests should be improved by using highly purified, synthetic viral antigens. With establishment of such an ELISA antigen in mind, we examined a bacterially synthesized polypeptide [ENV(80)] that corresponds to 80 conserved amino acids of the HIV gp41 transmembrane glycoprotein. ENV(80) was expressed as a DHFR fusion protein in Escherichia coli. Results obtained by HIV ELISA and immunoprecipitation with 497 serum samples from various groups at risk of AIDS were compared with those obtained with the ENV(80) ELISA. The ENV(80) ELISA was found to be superior to the H9/HTLV-III ELISA with respect to sensitivity and specificity and is almost equivalent in accuracy to immunoprecipitation.

Subregions of a conserved part of the HIV gp41 transmembrane protein are differentially recognized by antibodies of infected individuals.

A 240-bp DNA fragment encoding a peptide, designated ENV(80), homologous to a conserved part of the gp41 transmembrane glycoprotein of human immunodeficiency virus (HIV) was chemically synthesized and inserted into different plasmid expression vectors. Escherichia coli transformants containing these plasmid constructs produced upon induction high amounts of either an ENV(80) peptide of relative molecular mass (Mr) of 10,000 or the same ENV(80) peptide N-terminally fused to E. coli chloramphenicol acetyltransferase (CAT) or to mouse dihydrofolate reductase (DHFR) having Mr of 36,000 and 31,000 respectively. All polypeptides containing the ENV(80) sequences were strongly reactive with antibodies present in sera from AIDS virus-infected individuals, but not with control sera. The strategy of gene assembly allowed the expression of ENV(80) subfragments fused to DHFR. The serodiagnosis of 15 positive sera by Western blot analysis using these bacterially synthesized ENV(80) subfragments revealed the presence of several immunoreactive epitopes on the 80-amino acid polypeptide which were recognized differently by the various patients.