Immunohistochemical demonstration of FC receptors in rat tissues using immune complexes as ligand

Summary To demonstrate the presence and localization of Fc receptors, rat liver cryostat sections were incubated with heterologous and autologous immune complexes (ICx) and immunoglobulin (Ig) aggregates. Binding was demonstrated using the immunoperoxidase technique. Autologous and heterologous ICx as well as aggregates from human and rat Ig appeared to bind to the sinusoidal wall. ICx bind in preference to aggregates. Monomeric Ig and aggregated Ig from swine and rabbit did not bind. The results demonstrated that ICx and rat and human Ig aggregates were bound via an Fc receptor. This Fc receptor was still intact in livers from carbontetra chloride and galactosamine treated rats. The receptor could also be demonstrated on spleen macrophages and on kidney interstitial cells. This method turned out to be an useful functional histochemical method to localize Fc receptors and to demonstrate their affinity and species specificity in tissues.     

The use of backscattered electrons in analytical electron microscopy for the measurement of the mass of individual rat blood platelets

Summary The backscattered electron signal, generated in individual cells, has been used to measure the dry mass of these cells. Absolute mass values were obtained by comparing the backscattered electron signals of cells to the signals of polystyrene-latex spheres of known mass. The technique was carried out in an automated analytical scanning transmission electron microscope and applied to rat blood platelets.The resulting mass distributions agreed well with the distribution measured with a method that uses the transmitted electron signal by means of densitometric analysis of electrographs. Also the range of masses was in agreement with values deduced from data in the literature.The fully automated technique has the advantage that it is direct, fast, and that thicker specimens can be measured than is possible using the transmitted electron signal. The method is intended for use in combination with quantitative electron probe X-ray microanalysis and is then able to produce elemental mass fractions of biological specimens at the subcellular level.In honour of Prof. van Duijn     

Light dependence of sexual agglutinability in Chlamydomonas eugametos

The mating activity of mating-type plus gametes of Chlamydomonas eugametos depends on light. Cells lost their ability to agglutinate with mating-type minus gametes after a dark period of 30 min. They regained their agglutinability after 10 min exposure to light. Other mating reactions, such as tipping and flagellar tip activation, were not dependent upon light. Since cycloheximide and tunicamycin did not affect the light-induced activation of flagellar agglutinability, no protein synthesis or glycosylation is involved in this process. Equal amounts of biologically active agglutination factor could be extracted from cells placed either in light or in darkness. A minor portion of the active material was found to be located on the flagellar surface of illuminated cells. No active material was found on the flagellar surface of dark-exposed cells, whereas their cell bodies contained the same amount of active material as the cell bodies of illuminated cells. Since a light-induced flow of agglutination factors from the cell body to the flagella could not be detected and dark-exposed cells could be slightly activated by amputation or fixation by glutaraldehyde, we propose that light affects flagellar agglutinability by an in-situ modification of the agglutination factor on the flagella. When mt ⁺ and mt ^- strains were crossed and the progeny examined for light-sensitivity, it was apparent that this phenomenon is not mating type-linked.Abbreviations and symbols FTA flagellar tip activation - mt ^+/- mating type plus or minus - WGA wheat-germ agglutinin     

Correlation between infection by Rhizobium leguminosarum and lectin on the surface of Pisum sativum L. roots

The lectin on the surface of 4- and 5-dold pea roots was located by the use of indirect immunofluorescence. Specific antibodies raised in rabbits against pea seed isolectin 2, which crossreact with root lectins, were used as primary immunoglobulins and were visualized with fluorescein- or tetramethylrhodamine-isothiocyanate-labeled goat antirabbit immunoglobulin G. Lectin was observed on the tips of newly formed, growing root hairs and on epidermal cells located just below the young hairs. On both types of cells, lectin was concentrated in dense small patches rather than uniformly distributed. Lectin-positive young hairs were grouped opposite the (proto)xylematic poles. Older but still-elongating root hairs presented only traces of lectin or none at all. A similar pattern of distribution was found in different pea cultivars, as well as in a supernodulating and a non-nodulating pea mutant. Growth in a nitrate concentration which inhibits nodulation did not affect lectin distribution on the surface of pea roots of this age. We tested whether or not the root zones where lectin was observed were susceptible to infection by Rhizobium leguminosarum. When low inoculum doses (consisting of less than 10⁶ bacteria·ml^-1) were placed next to lectin-positive epidermal cells and on newly formed root hairs, nodules on the primary roots were formed in 73% and 90% of the plants, respectively. Only a few plants showed primary root nodulation when the inoculum was placed on the root zone where lectin was scarce or absent. These results show that lectin is present at those sites on the pea root that are susceptible to infection by the bacterial symbiont.Abbreviations FITC fluorescein isothiocyanate - TRIC tetramethylrhodamine isothiocyanate     

Laying, development and hatching of eggs of the fish ectoparasite Argulus japonicus (Crustacea: Branchiura)

Egg laying of the fish-louse Argulus japonicus was observed and examined experimentally. The effect of temperature on development time and hatching yielded an inverse exponential function. Hatching started after 61–10 days in a temperature span of 15–35 °C. Eggs are laid in strings on hard substrata and covered by a gelatinous material. Females lay between 1–9 strings, 5–226 eggs per string, arranged in 1–6 rows. Four embryonic developmental stages were recognized and the mean hatching efficiency was 50% in the optimal temperature range of 20–30 °C. Hatching efficiency was not related to either the number of eggs in a string or the total number of eggs laid by any particular female. Argulus japonicus displays continuous egg-laying activity with a possibility of an overwintering mechanism which suggests a seasonality of a sort.     

Characterization of the folic acid C9-N10-cleaving enzyme of Dictyostelium minutum V3

Folic acid is a chemoattractant for the slime mold Dictyostelium minutum V3. The activity of extracellular folic acid is regulated by a folic acid C9-N10 splitting enzyme (FAS). The products were identified as pterin-6-aldehyde and p-amino-benzoylglutamic acid. The enzyme was stabilized by EDTA. For the extracellular enzyme, the Km was 10(-7) M, and the optimal pH was 4.0. During starvation, FAS activity was mainly secreted into the medium; after 3 h, a plateau was reached. The membrane-bound activity was constant, but only 12% of the extracellular activity at 3 h. Intracellular activity also increased up to 3 h to a level of 23% of the extracellular FAS. The substrate recognition of FAS was found to be based on 4-O or N3 or both, N5 or N8 or both, N10, and the p-aminobenzoic acid moiety, whereas 2-NH2, N1, and the glutamic acid moiety were not recognized. Other slime mold species were found to secrete FAS with 20-fold or more reduced activity than D. minutum V3.     

Effect of pKM101 on cell killing and specificity of mutation induction by cis-diaminedichloroplatinum(II) in Escherichia coli K-12.

Cell killing and mutation induction in the lacI gene of Escherichia coli by cis-Pt(NH3)2Cl2 were studied in cells with different repair capacities, with and without pKM101. The presence of the plasmid pKM101 made repair-proficient cells more susceptible to killing by cis-Pt(NH3)2Cl2 and strongly enhanced mutation induction by that compound. Both effects were shown to be dependent upon excision repair. Characterization of the induced mutations in the lacI gene after cis-Pt(NH3)2Cl2 treatment of E. coli cells, by the LacI system, revealed that the mutagenic specificity of the Pt compound was strongly influenced by the presence of the pKM101 plasmid. With pKM101, 23% of the induced amber and ochre mutations resulted from substitutions at AT base pairs, whereas these mutations were hardly induced in cells without pKM101. These results suggest that pKM101-induced repair differs from normal SOS repair.     

Inhibition of DNA replication by transformation in a Haemophilus influenzae mutant carrying an altered Rec-1 protein.

In the HM5 mutant of Haemophilus influenzae, which carries a mutation in the rec-1 gene region and in which the replication of donor-recipient DNA complexes formed in transformation is inhibited, the transformation frequency could be greatly enhanced by inhibition of protein synthesis during transformation, indicating that transformation in the HM5 mutant induces the synthesis of a protein that inhibits the replication of the donor-recipient DNA complexes. This induction occurred in an early step of the recombination. Synthesis of the wild-type Rec-1 protein after transformation of the HM5 mutant with wild-type DNA could diminish the inhibiting effect on DNA replication. The HM5 mutant synthesized an altered Rec-1 protein (molecular weight, 38,000) whose pI differed from that of the wild type. As a result of the mutation in the rec-1 gene, two other proteins (molecular weights, 37,500 and 43,000) are lacking in the HM5 mutant.     

In vivo 31P and 13C nuclear magnetic resonance studies of acetate metabolism in Chromatium vinosum

31P and 13C nuclear magnetic resonance (NMR) experiments were performed on suspensions of the phototrophic bacterium Chromatium vinosum incubated anaerobically in the dark. 31P NMR spectra revealed that during prolonged dark incubation high ATP levels are maintained. This phenomenon was independent of the presence of the energy reserves polyglucose and polyphosphate. 13C NMR experiments revealed that the amino acids glutamate, aspartate, and alanine are the major products of acetate incorporation in the dark. Apart from these amino acids, poly-beta-hydroxybutyrate was also formed. Acetate metabolism was markedly stimulated by the presence of polyglucose. The specific 13C activity of glutamate C-2 was approximately 50% that of glutamate C-4. The idea is discussed that this difference is the consequence of the maintenance of redox balance during entry of acetate into cell metabolism.