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Biomedical subjects

K Hedman

Publications and source records attributed to K Hedman.

At least 19 recordsLinked to original sources
PubMed1992

Prokaryotic expression of a VP1 polypeptide antigen for diagnosis by a human parvovirus B19 antibody enzyme immunoassay.

To produce parvovirus B19 antigen for diagnostic purposes, partially overlapping segments covering the genes encoding the viral structural proteins VP1 and VP2 were cloned into expression vectors. The constructs were induced in Escherichia coli, resulting in the expression of beta-galactosidase fusion proteins. In immunoblotting experiments with sera from patients with erythema infectiosum, immunoglobulin G (IgG) and IgM antibodies bound to a single polypeptide of 235 amino acids at the N terminus of VP1. The DNA fragment encoding this polypeptide was amplified by the polymerase chain reaction and cloned into an expression vector. The viral capsid antigen expressed in E. coli was purified by preparative agarose gel electrophoresis and used in IgG and IgM solid-phase enzyme immunoassays. Comparison with reference gamma- and mu-capture radioimmunoassays using whole virus antigen showed that these antibody tests are suitable for the serodiagnosis of human infections caused by parvovirus B19.

Antibodies, Viral↗
PubMed1992

Immunoglobulin G antibody avidity in patients with respiratory syncytial virus infection.

The titer and avidity of respiratory syncytial virus-specific antibodies were measured in 196 serum specimens from 93 children with an acute, laboratory-confirmed respiratory syncytial virus infection. An enzyme immunoassay method based on the ability of urea to dissociate the bound antibodies with low avidity from the antigen was used. Three patterns of immune responses were observed. Children less than 6 months of age usually had low titers of antibodies with high avidity in their acute-phase serum samples. These antibodies were concluded to be of maternal origin, since their reaction pattern was similar to that of healthy adults. During the next few weeks, a slight increase in titers with a concurrent decrease in antibody avidity was observed. All children 6 to 24 months of age had low-avidity antibodies in their acute-phase serum samples, which matured to high avidity during the follow-up. On the contrary, about half of the children greater than 24 months of age had high-avidity antibodies already in the acute-phase serum samples. We conclude that the former children were experiencing primary infections with respiratory syncytial virus and the latter were experiencing reinfections. All adults with remote immunity had antibodies with high avidity.

Adult↗
PubMed1992

Incidence of primary toxoplasma infections during pregnancy in southern Finland: a prospective cohort study.

Congenital toxoplasmosis may develop after maternal primary infection during pregnancy. Since toxoplasmosis in adults is usually asymptomatic, the only way to find the pregnancies at risk of acute toxoplasmosis (i.e. seronegative mothers) is to perform serological screening of all pregnant women. We studied toxoplasma-specific antibodies in 16,733 pregnant women in the first, second, and third trimesters over an 18-month period in the Helsinki area, with a population of 820,700. The participation rate in our study was 90.2% of all pregnancies in the area. The overall prevalence of toxoplasma seropositivity was 20.3%. The incidence of primary infection was 2.4/1000 pregnancies at risk. Our data on incidence suggest an annual occurrence of 131 cases of primary toxoplasma infections during pregnancy in Finland. Primary toxoplasma infection appears to be a significant risk to the fetus even in countries where the prevalence of toxoplasma seropositivity is low. The feasibility of mass screening for toxoplasma infections during pregnancy should be considered in order to reduce the morbidity and mortality due to congenital toxoplasmosis.

Adolescent↗
PubMed1992

The program for antenatal screening of toxoplasmosis in Finland: a prospective cohort study.

16,733 pregnant women were studied for toxoplasma specific antibodies during an 18-month period. The number of specimens was 44,181. The IgG seropositivity of mothers was 20.3%. The incidence of primary toxoplasma infections during pregnancy was 4.2 per 1000 seronegative pregnancies by using the traditional criteria, IgG seroconversion and/or a positive IgM. By using the IgG activity test as an additional criterion, the incidence was 2.4 per 1000 seronegative pregnancies. Due to transmission rate of 40%, the number of congenital toxoplasma infections in Finland is more than 50 per year. The evaluation is going on and the cost-benefit analysis will reveal the effectiveness of antenatal screening.

Antibodies, Protozoan↗
PubMed1991

Rapid diagnosis of hantavirus disease with an IgG-avidity assay.

Nephropathia epidemica (NE), due to Puumala virus, is endemic in eastern Europe and Scandinavia. Serodiagnosis of NE relies on conventional indirect immunofluorescence to detect IgG against Puumala virus. However, in conventional serology, most patients with acute NE have raised but stable non-diagnostic antibody titres. For better serodiagnosis of NE, we have developed a test that measures the avidity (functional affinity) of IgG antibodies against Puumala virus. This new assay was highly specific and sensitive; a diagnosis of NE could be confirmed or excluded rapidly from an early single serum sample. With this test we have now verified the diagnosis of NE in more than 1300 Finnish patients during 22 months in 1989-91. Our findings point to an incidence of confirmed hantavirus disease much higher than previously shown.

Antibodies, Viral↗
PubMed1989

Measurement of avidity of specific IgG for verification of recent primary rubella.

Sixty-four subjects' serum samples, positive or equivocal by rubella IgM assays and containing rubella IgG, were examined for the avidity of rubella IgG. Four of the sera originated from rubella reinfections; others had false-positive IgM results due to interference by parvovirus infection or by other mechanisms; and the remaining were sera from the acute phase or convalescence of primary rubella. A novel IgG avidity test, avidity-ELISA, and a semiquantitative haemolysis typing assay were used. According to the avidity-ELISA, 29 subjects had recent primary rubella (low IgG avidity), and another 29 had previous rubella immunity (high IgG avidity), whereas 6 serum samples gave borderline avidity values. Comparison of these results with pre-existing clinical records and laboratory data showed that all samples with low IgG avidity were obtained during or shortly after acute primary rubella. All sera with high IgG avidity originated from the previously immune subjects; the rubella reinfections were confined within this group. Five of the six sera with borderline avidity values were obtained within 2 months from primary rubella. In conclusion, the measurement of IgG avidity is a powerful tool for the distinction of acute or recent primary rubella from pre-existing rubella immunity, including rubella reinfections.

Adult↗
PubMed1989

Maturation of immunoglobulin G avidity after rubella vaccination studied by an enzyme linked immunosorbent assay (avidity-ELISA) and by haemolysis typing.

Two tests were introduced recently for assessment of the avidity of rubella immunoglobulin antibodies. In the quantitative test--avidity-enzyme linked immunosorbent assay (ELISA)--IgG antibodies obtained from individuals shortly after primary infection with rubella virus are distinguished from those with past immunity by their antigen-elution characteristics. This method uses agents that disrupt hydrophobic bonds in proteins [Kamoun PP (1988): Denaturation of globular proteins by urea: Breakdown of hydrophobic bonds? Trends in Biological Sciences 13:424-425.]. In the semiquantitative, presumptive test--haemolysis typing--the low-avidity rubella-IgG antibodies are distinguished from the high-avidity antibodies by the quality of their haemolytic zones in a radial haemolysis test. In the present study, both tests were applied to sera taken before and after vaccination with two different strains (Cendehill or RA 27/3) of live attenuated rubella virus. It was found that after vaccination of previously nonimmune subjects, IgG synthesized during the first 2 months had a very low avidity; IgG avidity increased dramatically during the subsequent 4 months and less markedly between 6 and 12 months after vaccination. On the contrary, the initially high IgG avidity of previous immune vaccinees remained at an elevated level postvaccination. These results provide a basis for identification of recent primary rubella virus infections, or vaccination reactions, by the avidity of specific IgG and also for their separation from rubella reinfections.

Adult↗
PubMed1989

Recent primary toxoplasma infection indicated by a low avidity of specific IgG.

An unusual assay was developed for the serologic diagnosis of recent primary infection by Toxoplasma gondii. This test measures the antigen-binding avidity of toxoplasma-specific IgG antibodies. Serum samples from 5 patients with recent primary toxoplasma infection were compared with those from 21 subjects with preexisting toxoplasma immunity. Patients with primary infection exhibited a low avidity of toxoplasma-specific IgG, which persisted for several months after the onset of symptoms of toxoplasmosis. In contrast, all subjects with past immunity had a high avidity of toxoplasma-IgG. This IgG avidity assay should assist in the diagnosis of acquired toxoplasmosis and may be used to identify pregnancies that are at risk for congenital toxoplasmosis.

Adult↗
PubMed1988

Recent rubella virus infection indicated by a low avidity of specific IgG.

Rubella-specific IgG in acute-phase sera produces a characteristically altered zone termed "soft hemolysis" in the radial hemolysis test. Here, the soft hemolysis was shown to be a product of the purified IgG1 subclass isolated from acute-phase sera. In contrast, ordinary hemolysis was produced by IgG1 isolated from sera of previous rubella immunity, indicating that the subclass composition of IgG was not involved in the mechanism of soft hemolysis. A novel type of solid-phase immunoassay was developed for the avidity of virus-specific IgG. Acute-phase IgG (with soft hemolysis) was dissociated from rubella antigen in an enzyme immunoassay (EIA) test by hydrogen-bond disrupting agents under conditions where IgG of previous immunity (showing ordinary hemolysis) remained mostly bound. These data suggest that the mechanism of soft hemolysis is the avidity of rubella-specific IgG. The new quantitative avidity EIA was tested with sera taken from 169 subjects. Recent infection could be shown from sera taken weeks or months after primary rubella.

Antibodies, Viral↗
PubMed1988

Inactivation of human immunodeficiency virus in serum specimens as a safety measure for diagnostic immunoassays.

Since the human immunodeficiency virus (HIV) may be transmitted accidentally to laboratory personnel analyzing patient sera, the efficiency of a non-ionic detergent, Triton X-100, in inactivation of HIV in human serum as a safety measure was studied. Semliki Forest virus, an enveloped toga virus, was used as a model virus to create optimal treatment conditions. In the presence of 50% serum, complete inactivation (i.e. no residual virus detected, greater than 7 log reduction of virus titre) was achieved by incubation with 0.2% Triton X-100 for 1 h at 37 degrees C. Under these conditions HIV was also completely inactivated (i.e. no residual infectious virus detected, greater than or equal to 5 log reduction of virus titre). Both treated and untreated serum specimens were also tested with several enzyme immunoassays used in virological laboratories to determine whether the inactivation treatment interfered with the assays. The treated specimens, further diluted as recommended for each assay, were subjected to 15 enzyme immunoassays for microbial antibodies and antigens (HIV IgG, hepatitis A IgG and IgM, hepatitis B s, c, and e antigens and antibodies, cytomegalovirus IgG, mumps virus IgG, poliovirus IgG, rubellavirus IgM, toxoplasma IgG, and chlamydia IgG). Clearly decreased sensitivity was found only with two hepatitis B tests (e antigen and antibody to the surface antigen). It is concluded that safe inactivation of HIV in serum is achieved by 0.2% Triton X-100, but the treatment may decrease the sensitivity of some tests in which low specimen dilution is used.

Acquired Immunodeficiency Syndrome↗
PubMed1988

Ultrastructural localization of plasma membrane-associated urokinase-type plasminogen activator at focal contacts.

We have recently shown that urokinase-type plasminogen activator (u-PA) and plasminogen activator inhibitor type 1 are both found extracellularly beneath cultured human skin fibroblasts and HT-1080 sarcoma cells, but in distinct localizations. Here, the ultrastructural distribution of u-PA was studied using immunoferritin electron microscopy. In HT-1080 cells, u-PA on the extracellular aspect of the plasma membrane was detected at sites of direct contact of the cell with the growth substratum beneath all parts of the ventral cell surface. The ferritin-labeled adhesion plaques, which were enriched in submembraneous microfilaments, were frequently seen at the leading lamellae of the cells as well as in lamellipodia and microspikes. Besides the cell-substratum adhesion plaques, ferritin label was detected at cell-cell contact sites. Double-label immunofluorescence showed a striking colocalization of u-PA and vinculin in both HT-1080 cells and WI-38 lung fibroblasts, which is consistent with u-PA being a focal contact component. The u-PA-containing focal contacts of WI-38 cells had no direct codistribution with fibronectin fibrils. In WI-38 cells made stationary by cultivation in a medium containing 0.5% FCS, vinculin plaques became highly elongated and more centrally located, whereas u-PA immunolabel disappeared from such focal adhesions. These findings show that plasma membrane-associated u-PA is an intrinsic component of focal contacts, where, we propose, it enables directional proteolysis for cell migration and invasion.

Cell Adhesion↗
PubMed1988

Pre-lysosomal divergence of alpha 2-macroglobulin and transferrin: a kinetic study using a monoclonal antibody against a lysosomal membrane glycoprotein (LAMP-1).

We have used electron microscopic immunocytochemistry to compare the distribution of LAMP-1, a marker for lysosomal membranes, with the intracellular localization of alpha 2-macroglobulin (alpha 2-M) and transferrin at various time points after their endocytosis into cultured NIH 3T3 cells. The purposes of this study were (a) to determine how soon endocytic ligands reach lysosomal organelles, (b) to examine whether the intermediate endocytic vesicles gained lysosomal markers gradually or in a precipitous, discrete event, and (c) to examine the relationship, if any, between the pathway of recycling ligands and lysosomes. At early time points (0-5 min) after initiation of endocytosis, most structures containing alpha 2-M labeled with colloidal gold (receptosomes) were not labeled by anti-LAMP-1 detected using ferritin bridge or peroxidase immunocytochemistry. At late time points (greater than or equal to 15 min), the structures containing alpha 2-M (lysosomes) were strongly labeled by anti-LAMP-1. In contrast, transferrin that was directly labeled with ferritin was mostly located in LAMP-1-negative structures at all time points studied. The proportion of alpha 2-M-gold containing vesicles strongly labeled for LAMP-1 roughly paralleled the proportion of alpha 2-M-gold-containing structures positive for cytochemically detectable acid phosphatase. Our data indicate that ligands such as transferrin that are internalized through coated pits and receptosomes, but not delivered to lysosomes, do not traverse a lysosomal organelle compartment as marked by LAMP-1 content. Ligands such as alpha 2-M that are destined for lysosomal delivery reach a LAMP-1-positive organelle compartment only after they traverse LAMP-1-negative, non-lysosomal vesicles previously described as receptosomes.

Acid Phosphatase↗
PubMed1987

Comparison of the intracellular pathways of transferrin recycling and vesicular stomatitis virus membrane glycoprotein exocytosis by ultrastructural double-label cytochemistry.

Transferrin is taken up by receptor-mediated endocytosis into intracellular vesicles and tubules, and then recycles rapidly to the plasma membrane (diacytosis). We applied double-label cytochemistry to study whether the recycling structures containing transferrin fuse with the intracellular membranous structures that deliver newly synthesized membrane glycoproteins from the ER to the plasma membrane (exocytosis) or whether they remain independent. KB and Vero cells were infected with the temperature-sensitive transport mutant 0-45 of vesicular stomatitis virus (VSV). Temperature-regulated exocytosis of membrane glycoprotein "G" occurred simultaneously with diacytosis of transferrin. The exocytic "G" protein, as detected by immunoperoxidase electron microscopy, passed through the cisternal Golgi stacks and vacuolar, tubular, vesicular, and pit-like structures of the Golgi system. A transferrin-ferritin conjugate used in ultrastructural double-label experiments was detected in diacytic vesicles and tubules that accumulated in the proximal (trans-reticular) Golgi area of the cell. The ferritin-labeled vesicles/tubules were often close to and intermixed with the VSV-"G" containing membranous structures, but in most cases at early times (15-20 min) the transferrin and VSV-"G" containing vesicular structures remained distinct. At later times (30-45 min), the two labels were occasionally found in the same structures. These results indicate that rapid recycling of endocytosed materials and exocytosis of membrane glycoproteins to the cell surface usually occur in distinct vesicles, possibly along the same general morphologic exit pathway.

Exocytosis↗