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  • Role of the first and third extracellular domains of CXCR-4 in human immunodeficiency virus coreceptor activity. 9151868

    The CXCR-4 chemokine receptor and CD4 behave as coreceptors for cell line-adapted human immunodeficiency virus types 1 and 2 (HIV-1 and HIV-2) and for dual-tropic HIV strains, which also use the CCR-5 coreceptor. The cell line-adapted HIV-1 strains LAI and NDK and the dual-tropic HIV-2 strain ROD were able to infect CD4+ cells expressing human CXCR-4, while only LAI was able to infect cells expressing the rat homolog of CXCR-4. This strain selectivity was addressed by using human-rat CXCR-4 chimeras. All chimeras tested mediated LAI infection, but only those containing the third extracellular domain (e3) of human CXCR-4 mediated NDK and ROD infection. The e3 domain might be required for the functional interaction of NDK and ROD, but not LAI, with CXCR-4. Alternatively, LAI might also interact with e3 but in a different way. Monoclonal antibody 12G5, raised against human CXCR-4, did not stain cells expressing rat CXCR-4. Chimeric human-rat CXCR-4 allowed us to map the 12G5 epitope in the e3 domain. The ability of 12G5 to neutralize infection by certain HIV-1 and HIV-2 strains is also consistent with the role of e3 in the coreceptor activity of CXCR-4. The deletion of most of the amino-terminal extracellular domain (e1) abolished the coreceptor activity of human CXCR-4 for ROD and NDK but not for LAI. These results indicate that HIV strains have different requirements for their interaction with CXCR-4. They also suggest differences in the interaction of dual-tropic HIV with CCR-5 and CXCR-4.
    Dokumententyp:
    Referenz
    Produkbestellnummer:
    Mehrere
    Produktbezeichnung:
    Mehrere

  • Anti-acetylcholine antibodies and first immunocytochemical application in insect brain. 3897911

    A specific immunological approach was developed to enable acetylcholine (ACh) to be visualized in biological tissues. A variety of ACh-like immunogens were synthesized, and injected into rabbits. Antibody specificity was tested using an enzyme-linked immunosorbent assay (ELISA) method. The most immunoreactive ACh derivative was found to be choline-glutaryl-lysine. A mixture of allyl alcohol and formaldehyde was found to be the best fixative of ACh in tissues. The specificity of this antibody recognition was tested in vitro and in immunochemistry. There was excellent agreement between the in vitro results and the ACh staining. Moreover, visualization using these anti-ACh antibodies appeared identical to the results using anti-choline acetyltransferase antibodies.
    Dokumententyp:
    Referenz
    Produkbestellnummer:
    AB5522

  • Intraprotomer masking of third variable loop (V3) epitopes by the first and second variable loops (V1V2) within the native HIV-1 envelope glycoprotein trimer. 22128330

    Within the trimeric HIV-1 envelope (Env) spike, the first and second variable loops (V1V2 region) and the third variable loop (V3) of the gp120 subunit play dual roles in antibody recognition, because they contain neutralization epitopes and also participate in epitope masking. The spatial relationships between V1V2 and V3 and the associated mechanisms of epitope masking remain unclear. Here we investigated interactions between these domains using two monoclonal antibodies recognizing distinct conserved linear epitopes that are subject to masking in the functional trimer, which limits their neutralizing activities. Using Env pseudotype virus infection assays, we found that deleting the V1V2 region greatly enhanced neutralization by both antibodies, leading us to consider two alternative models: V1V2 on one gp120 protomer masks V3 on the same protomer (intraprotomer or cis masking) versus on an adjacent protomer (interprotomer or trans masking). Our experimental approach exploited a previously described complementation system wherein two variant Envs harboring different inactivating mutations (one in gp120, the other in gp41) are coexpressed in the same cell; functional Env results only from cooperative interactions within mixed trimers, thereby enabling selective examination of mixed trimer activity. We introduced additional mutations that either promoted (V1V2 deletion, i.e., unmasking) or prevented (GPGR to GPGQ mutation, i.e., epitope destruction) interaction with the antibodies. The observed neutralization sensitivities of mixed trimers produced from various combinations of constructs support the intraprotomer (cis) model of V1V2 masking of V3 epitopes.
    Dokumententyp:
    Referenz
    Produkbestellnummer:
    MAB447

  • Anti-myelin antibodies predict the clinical outcome after a first episode suggestive of MS. 17468447

    The aim of this study was to test the contribution of anti-myelin antibodies in predicting conversion from clinically isolated syndrome (CIS) to multiple sclerosis (MS) when considering either Poser's or McDonald's diagnostic criteria. Fifty-one patients with CIS and abnormal brain MRI were imaged monthly for six months and then at 12, 18, 24, 36 months. At baseline serum samples testing antibodies against myelin oligodendrocyte glycoprotein (anti-MOG) and myelin basic protein (anti-MBP) were collected. During the 36-month follow-up, 26 (51%) patients developed a relapse thus becoming clinically definite MS (CDMS) according to Poser's criteria; 46 (90.2%) patients converted to MS according to McDonald's criteria. Out of 51 patients, 28 (54.9%) had either double or single positivity for anti-myelin antibodies. Antibody status significantly predicted MS according to Poser's criteria (P=0.004), but did not according to the McDonald's criteria. When compared to antibody negative patients, the risk of developing a relapse was 8.9 (95% CI: 2.7-29.8; P0.001) for anti-MBP positive (anti-MBP+) patients and 1.5 (95% CI: 0.4-5.4; P=0.564) for those anti-MOG positive (anti-MOG+); double positive patients (ie, anti-MBP+/anti-MOG+) had a risk of relapse's occurrence equal to 3.4 (95% CI: 1.1-10.2; P=0.031). Also, the antibody status predicted the median time span from CIS to CDMS, that was of 36 months in the anti-MOG-/anti-MBP- group, 33 months in the anti-MOG+/anti-MBP- group, 24 months in the anti-MOG+/anti-MBP+ group and 12 months in the anti-MOG-/anti-MBP+ patients (P=0.003 by ANOVA). Our data support the prognostic value of anti-myelin antibodies in CIS patients at risk of CDMS, with positive patients showing shorter time interval to relapse occurrence than negative patients.
    Dokumententyp:
    Referenz
    Produkbestellnummer:
    MAB381

  • Antimyelin antibodies as a predictor of clinically definite multiple sclerosis after a first demyelinating event. 12853586

    BACKGROUND: Most patients with multiple sclerosis initially present with a clinically isolated syndrome. Despite the fact that clinically definite multiple sclerosis will develop in up to 80 percent of these patients, the course of the disease is unpredictable at its onset and requires long-term observation or repeated magnetic resonance imaging (MRI). We investigated whether the presence of serum antibodies against myelin oligodendrocyte glycoprotein (MOG) and myelin basic protein (MBP) in patients with a clinically isolated syndrome predicts the interval to conversion to clinically definite multiple sclerosis. METHODS: A total of 103 patients with a clinically isolated syndrome, positive findings on cerebral MRI, and oligoclonal bands in the cerebrospinal fluid were studied. At base line, serum samples were collected to test for anti-MOG and anti-MBP antibodies with Western blot analysis, and the lesions detected by cerebral MRI were quantified. Neurologic examinations for relapse or disease progression (defined as conversion to clinically definite multiple sclerosis) were performed at base line and subsequently every three months. RESULTS: Patients with anti-MOG and anti-MBP antibodies had relapses more often and earlier than patients without these antibodies. Only 9 of 39 antibody-seronegative patients (23 percent) had a relapse, and the mean (+/-SD) time to relapse was 45.1+/-13.7 months. In contrast, 21 of 22 patients (95 percent) with antibodies against both MOG and MBP had a relapse within a mean of 7.5+/-4.4 months, and 35 of 42 patients (83 percent) with only anti-MOG antibodies had a relapse within 14.6+/-9.6 months (P0.001 for both comparisons with antibody-seronegative patients). The adjusted hazard ratio for the development of clinically definite multiple sclerosis was 76.5 (95 percent confidence interval, 20.6 to 284.6) among the patients who were seropositive for both antibodies and 31.6 (95 percent confidence interval, 9.5 to 104.5) among the patients who were seropositive only for anti-MOG antibodies, as compared with the seronegative patients. CONCLUSIONS: Analysis of antibodies against MOG and MBP in patients with a clinically isolated syndrome is a rapid, inexpensive, and precise method for the prediction of early conversion to clinically definite multiple sclerosis. This finding may be important for the counseling and care of patients with a first demyelinating event suggestive of multiple sclerosis.
    Dokumententyp:
    Referenz
    Produkbestellnummer:
    MAB381

  • Immunohistochemical studies on phosphorylation of tyrosine hydroxylase in central catecholamine neurons using site- and phosphorylation state-specific antibodies. 9483531

    Antibodies raised to phosphorylated forms of tyrosine hydroxylase, the first and rate-limiting enzyme in the catecholamine biosynthesis, were applied in immunohistochemical studies on rat brain slices incubated in vitro with a phosphodiesterase inhibitor (3-isobutyl-1-methylxanthine, IBMX) and on forskolin on formalin-perfused rat brains. Four antisera/antibodies were used: polyclonal rabbit antisera to (i) tyrosine hydroxylase phosphorylated at serine 40 (THS40p antiserum), (ii) tyrosine hydroxylase phosphorylated at serine 19 (THS19p antiserum), (iii) the native enzyme (pan-tyrosine hydroxylase antiserum), and mouse monoclonal antibody to (iv) native tyrosine hydroxylase. In the in vitro studies THS40p-like immunoreactivity was not observed unless slices were treated with IBMX-forskolin after which a dense fibre network was found in the striatum, and immunoreactive cell bodies were found in the ventral mesencephalon, especially in the ventral tegmental area. Although these cells were pan-tyrosine hydroxylase-positive, several of them were not stained with the tyrosine hydroxylase-monoclonal antibody. Moreover, there was a marked reduction of tyrosine hydroxylase-monoclonal antibody-immunoreactive fibres in drug-treated slices, suggesting that this tyrosine hydroxylase-monoclonal antibody does not recognize the serine 40-phosphorylated form of tyrosine hydroxylase. Treated slices did not show any THS40p-immunoreactive cell bodies in the dopaminergic A11 cell group and only a few, weakly fluorescent neurons were observed in locus coeruleus. However, a sparse fibre plexus was observed in locus coeruleus, possibly reflecting epinephrine fibres. In the perfused brains THS40p-like immunoreactivity could be visualized in some dopamine neurons in the ventral mesencephalon, especially the A10 area, and in noradrenergic locus coeruleus neurons, whereas THS19p-like immunoreactivity was found in all catecholamine groups studied, similar to the results obtained with the pan-tyrosine hydroxylase antiserum and the tyrosine hydroxylase-monoclonal antibody. In forebrain areas known to be innervated by mesencephalic dopamine neurons, no THS40p-positive fibres were observed, whereas THS19p-immunoreactive fibres were found in subregions of the striatum, olfactory tubercle and nucleus accumbens, essentially overlapping with dopamine fibres previously shown to contain cholecystokinin-like immunoreactivity. The present results suggests that antibodies directed against phosphorylated forms of tyrosine hydroxylase can be used to evaluate the state of tyrosine hydroxylase phosphorylation in individual neuronal cell bodies and processes both in vitro and in vivo.
    Dokumententyp:
    Referenz
    Produkbestellnummer:
    AB5986P
    Produktbezeichnung:
    Anti-Tyrosine Hydroxylase Antibody

  • DNase X is a glycosylphosphatidylinositol-anchored membrane enzyme that provides a barrier to endocytosis-mediated transfer of a foreign gene. 17416904

    DNase X is the first mammalian DNase to be isolated that is homologous to DNase I. In this study, we have examined its function using a novel monoclonal antibody and showed it to be expressed on the cell surface as a glycosylphosphatidylinositolanchored membrane protein. High level expression was observed in human muscular tissues and in myotubes obtained in vitro from RD rhabdomyosarcoma cells. We observed that RD myotubes incorporated a foreign gene, lacZ, by endocytosis but that expression of the encoded coding product, beta-galactosidase, was strongly inhibited. Overexpression of DNase X inhibited endocytosis-mediated gene transfer, whereas knockdown of DNase X with small interfering RNA had the opposite effect. These results reveal that DNase X provides a cell surface barrier to endocytosis-mediated gene transfer.
    Dokumententyp:
    Referenz
    Produkbestellnummer:
    MAB374
    Produktbezeichnung:
    Anti-Glyceraldehyde-3-Phosphate Dehydrogenase Antibody, clone 6C5

  • Beacon immunoreactivity in the rat hypothalamus. 16511859

    Beacon (BC) is a peptide of 73 amino acids, whose gene expression was first reported in the hypothalamus of Psammomys obesus (or Israeli sand rat). To appreciate better the functional role of BC in normal rats and sand rats, the distribution of BC immunoreactivity (irBC) and its subcellular localization were studied in the brain of Sprague-Dawley rats. In the hypothalamus, intense staining was present in neurons of the supraoptic (SO), paraventricular (PVH), and accessory neurosecretory nuclei and in cell processes of median eminence. Double labeling of the hypothalamic sections with mouse monoclonal oxytocin (OT) antibody and rabbit polyclonal BC antiserum revealed that nearly all OT-immunoreactive cells from SO, PVH, and accessory neurosecretory nuclei were irBC. Double labeling of the sections with guinea pig vasopressin (VP) antiserum and BC antiserum showed that a population of VP-immunoreactive neurons was irBC. By immunoelectron microscopy, immunoreactive product was associated with mitochondrial membranes or appeared as electron-dense bodies in many PVH and SO neurons. Most of the neurosecretory granules were unstained for BC. Taken together, our results indicate the presence of beacon in the OT-containing neurons and a population of VP-containing neurons, mostly associated with mitochondrial membrane. Insofar as the amino acids sequence of beacon is identical to that of ubiquitin-like 5, it is possible that the distribution of BC immunoreactivity noted in our study is that of ubiquitin-like 5 peptide in the rat hypothalamus.
    Dokumententyp:
    Referenz
    Produkbestellnummer:
    MAB5296
    Produktbezeichnung:
    Anti-Oxytocin Antibody, clone 4G11

  • Localization of the binding site for the monocyte immunoglobulin (Ig) A-Fc receptor (CD89) to the domain boundary between Calpha2 and Calpha3 in human IgA1. 8666916

    Immunoglobulin (Ig) A serves as the first line of humoral defense at all mucosal surfaces and is present in large quantities of blood. In playing its role in humoral immunity, IgA interacts with a variety of effector molecules present both in serum and on the surfaces of immune and inflammatory cells. To study these interactions, we previously established expression of human IgA1 in insect cells using recombinant baculoviruses and showed that the expressed antibody is a structurally and functionally intact polypeptide useful for examining the molecular properties of IgA. Indeed, since the C alpha 2 N-linked glycosylation site lies near the Fab-distal pole of C alpha 2, the inability of a mutant IgA1 lacking C alpha 2 N-glycosylation to bind its cognate receptor suggested that the monocyte Fc alpha receptor (mFcalphaR) recognizes IgA at a hinge-distal site encompassing the boundary between the C alpha 2 and C alpha 3 domains. In this report, we utilize both domain-swapped IgA/IgG and point-mutated IgA chimeras to verify the above hypothesis. Using an antigen-specific rosetting assay and a mFc alpha R-expressing cell line, we show that (a) C alpha 2 and C alpha 3 together are necessary and sufficient for binding; (b) neither the IgA hinge nor the tailpiece is necessary for binding; (c) mutations away from the interdomain boundary do not affect binding; and (d) mutations located near the three-dimensional boundary between C alpha 2 and C alpha 3 completely disrupt binding. Taken together, these results localize the mFc alpha R recognition site on IgA to the boundary region between the second and third constant domains--a site analogous to that recognized by Staphylococcus aureus protein A on IgG. The use of this hinge-distal site is, to date, unique among Fc receptors of the Ig superfamily.
    Dokumententyp:
    Referenz
    Produkbestellnummer:
    04-1579

  • Human herpesvirus-6-associated exanthema in a patient with acute lymphocytic leukaemia. 8616091

    Summary: We report the first case of human herpesvirus-6 (HHV-6) associated exanthema in a patient with acute lymphocytic leukaemia (ALL). We analysed DNA extracted from an exanthematous lesion using the polymerase chain reaction (PCR). DNA was positive for HHV-6 but negative for herpes simplex virus, varicella zoster virus, and cytomegalovirus. Immunohistochemical staining of the skin with monoclonal antibody against (HHV-6 confirmed the infection. The possibility of HHV-6 infection should be considered when an atypical skin rash is seen in patients with ALL.
    Dokumententyp:
    Referenz
    Produkbestellnummer:
    MAB8535
    Produktbezeichnung:
    Anti-Herpes Virus 6 Antibody, 101kDa protein, clone C3108-103