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The detection of substructures within proteoglycan molecules. Electron-microscopic immuno-localization with the use of Protein A-gold. 总被引:1,自引:1,他引:0 下载免费PDF全文
Proteoglycan monomers from pig laryngeal cartilage were examined by electron microscopy with benzyldimethylalkylammonium chloride as the spreading agent. The proteoglycans appeared as extended molecules with a beaded structure, representing the chondroitin sulphate chains collapsed around the protein core. Often a fine filamentous tail was present at one end. Substructures within proteoglycan molecules were localized by incubation with specific antibodies followed by Protein A-gold (diameter 4 nm). After the use of an anti-(binding region) serum the Protein A-gold (typically one to three particles) bound at the extreme end of the filamentous region. A small proportion of the labelled molecules (10-15%) showed the presence of gold particles at both ends. A monoclonal antibody specific for a keratan sulphate epitope (MZ15) localized a keratan sulphate-rich region at one end of the proteoglycan, but gold particles were not observed along the extended part of the protein core. This distribution was not changed by prior chondroitin AC lyase digestion of the proteoglycan. Localization with a different monoclonal antibody to keratan sulphate (5-D-4) caused a change in the spreading behaviour of a proportion (approx. 20%) of the proteoglycan monomers that lost their beaded structure and appeared with the chondroitin sulphate chains projecting from the protein core. In these molecules the Protein A-gold localized antibody (5-D-4) along the length of the protein core whereas in those molecules with a beaded appearance it labelled only at one end. Labelling with either of the monoclonal antibodies was specific, as it was inhibited by exogenously added keratan sulphate. The differential localization achieved may reflect structural differences within the proteoglycan population involving keratan sulphate and the protein core to which it is attached. The results showed that by this technique substructures within proteoglycan molecules can be identified by Protein A-gold labelling after the use of specific monoclonal or polyclonal antibodies. 相似文献
4.
Antigen-presenting cell-T cell interaction in the chicken is MHC class II antigen restricted 总被引:4,自引:0,他引:4
O Vainio T Veromaa E Eerola P Toivanen M J Ratcliffe 《Journal of immunology (Baltimore, Md. : 1950)》1988,140(9):2864-2868
The involvement of the MHC in the recognition of Ag by avian T lymphocytes was analyzed. PBL from chickens primed with keyhole limpet hemocyanin in vivo were induced to synthesize DNA in an in vitro response to specific Ag. Responding cells were T cells as judged by immunofluorescence staining. In vivo Ag-primed PBL were stimulated in vitro with specific Ag and further propagated in the presence of IL-2. Subsequent Ag-specific T cell proliferation required the presence of Ag-pulsed peripheral blood adherent cells (APC). T cell responses were restricted by the MHC of the APC; Ag presented by allogeneic APC did not support T cell proliferation. By using MHC-recombinant chicken lines, the gene products controlled by MHC class II loci were shown to restrict the T cell-APC interaction. This conclusion was substantiated by the inhibition of the Ag-specific T cell response by a mAb against chicken MHC class II gene products but not by a mAb against chicken MHC class I gene products. 相似文献
5.
Jean F. Emly Wendy A. Ratcliffe Elaine Green Sarah J. Bowden David A. Heath Ann Blight Susan Hughes John G. Ratcliffe 《生物化学与生物物理学报:疾病的分子基础》1992,1180(1):58-64
The molecular forms of parathyroid hormone-related protein (PTHRP) in conditioned media from the BEN human lung cancer cell line, rat parathyroid cells (PT-r) and human keratinocytes were studied by gel-filtraton chromatography with assay of PTHRP by immunoassays and bioassay. Immunoreactivity (1–86 and 1–34) and bioactivity (1–34) in conditioned media eluted as a coincident major peak (approx. molecular mass 19–22 kDa) and there was evidence of amino-terminal species in the molecular mass range 10–16 kDa in BEN and keratinocyte media. Western blotting of PTHRP affinity purified by monoclonal antibodies directed at regions 1–34 or 37–67, identified a major species in all cell cytosols and media with an apparent molecular mass of 24–25 kDa, consistently slighty larger than recombinant PTHRP(1–141) (mobility of 21 kDa) which may represent an intact or native form of PTHRP. Additional amino-terminal species were identified in medium from keratinocytes (16 and 7 kDa), BEN cells (18 and 14 kDa) and PT-R cells (17 kDa), suggesting that processing occurs at the C-terminus and within the mid-region to form a range of amino-terminal fragments. 相似文献
6.
The external morphology of the terminal region of the fifth tarsal segment of the blowfly, Calliphora vomitoria (L.) has been studied using light and scanning electron microscopy (S.E.M.). The pulvilli, with their numerous tenent hairs of spatulate form projecting from the ventral surface, are responsible for adhesion to smooth surfaces. The two large claws are believed to be important in clinging to irregularities in surfaces. Two footplates, possibly sensory organs, lie in close association with the base of the large ventral seta, the empodium. Blowflies release a non-volatile lipid secretion on to the spatulate ends of the tenent hairs and this secretion is essential to the adhesion process on smooth surfaces. The force of adhesion has been measured for tethered blowflies on glass using both vertical and lateral pulls; lateral pulls gave much greater forces. It is concluded that surface tension of the lipid secretion under tenent hairs is sufficient to enable successful adhesion to smooth surfaces by blowflies. 相似文献
7.
The structure and possible mode of functioning of the female reproductive system in Nasonia vitripennis (Hymenoptera: Pteromalidae) 总被引:1,自引:0,他引:1
Associated with the female insect reproductive system are a number of glands. Those present in Nasonia vitripennis are described, their microanatomy examined and where possible this is linked with their function during the process of drilling into the host puparium, feeding on the host fluids and oviposition. The structures dealt with are the ovaries, spermatheca, alkaline gland, colleterial gland and the acid gland with reservoir. 相似文献
8.
J. Sklenar G. G. Fox B. C. Loughman A. D. B. Pannifer R. G. Ratcliffe 《Plant and Soil》1994,167(1):57-62
Although the sensitivity of the plasma membrane H+-ATPase to vanadate is well known, the metabolic response of plant cells to vanadate is less well characterised in vivo and its use as an inhibitor in whole plant experiments has had mixed success. Experiments with maize (Zea mays, L.) roots and with purified plasma membrane fractions from the same tissues showed that exposure to vanadate caused: (i) a reduction in the capacity for phosphate uptake; (ii) a reduction in the extractable ATPase activity from the tissue; and (iii) a significant increase in the ATP level. The measurements on the extractable ATPase activity and the ATP level showed that the effect of vanadate developed slowly, apparently reflecting the slow accumulation of intracellular vanadate. The marked effect of vanadate on the ATP level-exposure to 500 M vanadate for 5 h doubled the ATP content of the roots tips-indicates that there is no stringent control over the ATP level in the roots and that the plasma membrane H+-ATPase activity is likely to have a significant role in determining the ATP level under normal conditions. 相似文献
9.
Nuclear magnetic resonance studies of the location and function of plant nutrients in vivo 总被引:2,自引:1,他引:1
The cytoplasmic and vacuolar pools of ammonium, inorganic phosphate and potassium can be studied non-invasively in plant tissues using high resolution nuclear magnetic resonance spectroscopy. The techniques that allow these pools to be discriminated in vivo are described and their application to plants is reviewed with reference to the phosphorus, nitrogen and potassium nutrition of root tissues. 相似文献
10.
Thomas F Floyd James M Clark Robert Gelfand John A Detre Sarah Ratcliffe Dimitri Guvakov Christian J Lambertsen Roderic G Eckenhoff 《Journal of applied physiology》2003,95(6):2453-2461
Breathing 100% O2 at 1 atmosphere absolute (ATA) is known to be associated with a decrease in cerebral blood flow (CBF). It is also accompanied by a fall in arterial Pco2 leading to uncertainty as to whether the cerebral vasoconstriction is totally or only in part caused by arterial hypocapnia. We tested the hypothesis that the increase in arterial Po2 while O2 was breathed at 1.0 ATA decreases CBF independently of a concurrent fall in arterial Pco2. CBF was measured in seven healthy men aged 21-62 yr by using noninvasive continuous arterial spin-labeled-perfusion MRI. The tracer in this technique, magnetically labeled protons in blood, has a half-life of seconds, allowing repetitive measurements over short time frames without contamination. CBF and arterial blood gases were measured while breathing air, 100% O2, and 4 and 6% CO2 in air and O2 backgrounds. Arterial Po2 increased from 91.7 +/- 6.8 Torr in air to 576.7 +/- 18.9 Torr in O2. Arterial Pco2 fell from 43.3 +/- 1.8 Torr in air to 40.2 +/- 3.3 Torr in O2. CBF-arterial Pco2 response curves for the air and hyperoxic runs were nearly parallel and separated by a distance representing a 28.7-32.6% decrement in CBF. Regression analysis confirmed the independent cerebral vasoconstrictive effect of increased arterial Po2. The present results also demonstrate that the magnitude of this effect at 1.0 ATA is greater than previously measured. 相似文献