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Flow Cytometric Analysis

Flow cytometry, as the name suggests, is the analysis of cells (which carry one or more fluorescent labels) moving in a fluid flow (1,2). This technique has become widely used because of the enormous increase in the number and range of specificities of antibodies to cell determinants. Monoclonal and ...

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Immunolabeling for Electron Microscopy

The protocols in this chapter concern postembedding immunolabeling for transmission electron microscopy; other schedules, such as pre-embedding methods, frozen tissue processes, and procedures for scanning electron microscopy, can be found elsewhere (1). In principle, imm ...

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PCR of the V Region

To engineer or manipulate antibody or Fv-based molecules, isolation of the V region of the antibodies is necessary. A number of strategies can be adopted for amplification; one such approach is to use subgroup-specific oligonucleotides for the amplification of the V-region genes. Small di ...

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Phage Display Technology: Protocols

Over the past few years, considerable effort has gone into genetic engineering and manipulation of antibody molecules. One consequence of this research has been the use of filamentous phage as a vehicle on which to display antibody fragments. It is possible to express a variety of molecules on the ...

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Meningococcal Vaccines and Vaccine Developments

Despite rapid advances in the diagnosis of bacterial infections and the availability of effective antibiotics, meningococcal disease continues to represent a substantial public health problem for most countries (1–4). Disease usually develops rapidly, is notoriously diffi ...

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Immunoblot Analysis of Sera from Patients and Vaccinees

This chapter will describe the use of the immunoblotting method for analysing antibody specificities towards meningococcal antigens in sera from vaccinees, receiving group B meningococcal outer-membrane vesicle (OMV) vaccines, or from patients, falling ill with meningococ ...

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Serogroup B and C Serum Bactericidal Assays

Meningococci are usually classified based on serological reactivity of their polysaccharide capsules with serogroups A, B, and C, currently the most common causes of disease. Serogroup A meningococci can cause massive outbreaks particularly in areas such as the African meningitis b ...

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A Modified ELISA for Measurement of High-Avidity IgG Antibodies to Meningococcal Serogroup C Polysaccharide that Correlate with Bactericidal Titers

This chapter describes a modified enzyme-linked immunosorbent assay (ELISA) employing assay conditions that ensure specificity of antibody binding and favor detection primarily of high-avidity serum IgG antibodies to meningococcal serogroup C polysaccharide (1). Antib ...

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Whole-Blood Model

Neisseria meningitidis is an obligate human pathogen. When it interacts with the host, it can establish a commensal relationship or can, on a minority of occasions, invade and cause systemic disease. Protection against systemic disease, particularly for serogroup A and C infections, has b ...

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Antibody-Induced Opsonophagocytosis of Serogroup B Meningococci Measured by Flow Cytometry

Antibodies can protect against meningococcal infection by at least two mechanisms: complement-dependent serum bactericidal activity (SBA), and/ or opsonophagocytosis (OP), leading to destruction of the bacteria (1–4). Regarding group C meningococci, there seems to be a correla ...

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T-Cell Responses Against Meningococcal Antigens

T-cells recognize protein antigens as short peptide fragments (8–20 amino acids) bound to major histocompatibility complex (MHC) molecules on the surface of antigen-presenting cells (APCs). A prerequisite for antigen-specific T-cell activation is antigen uptake, enzymatic de ...

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CD4 T-Cell Epitope Mapping

The majority of T cells recognize peptide epitopes bound to major histocompatibility complex (MHC)-encoded glycoproteins on the surface of professional antigen-presenting cells (APC), principally dendritic cells, macrophages, and B cells (1–3). Most T cells are specific for pept ...

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The Introduction of Group C Conjugate Meningococcal Vaccine into the UK

Throughout the 1990s, the incidence of meningococcal disease was higher in England and Wales than in most other European countries (1). In addition to the high incidence, the pattern of disease changed, with shifts to more cases caused by serogroup C strains, and more cases occurring in older teena ...

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Meningococcal Vaccine Trials

The evaluation of meningococcal vaccines in humans is a challenging task. Issues of safety, and benefit to the individual patient and to the community may raise difficult ethical problems. The inherent variability of human responses, the rarity of clinical disease, age-dependence in the i ...

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Purification of Capsular Polysaccharide

Meningococcus is an aerobic, fastidious, Gram-negative diplococcus that is found only in humans. Its cell wall has a cytoplasmic membrane, a peptidoglycan layer, and an outer membrane. Meningococci isolated from the bloodstream or the spinal fluid are almost always encapsulated. The ca ...

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Immune Response and Host-Pathogen Interactions

For the most part, the relationship between the pathogen, Neisseria meningitidis, and humans is uneventful. Colonization of the human nasopharynx at various times during life is an almost universal experience but clinically overt disease is unusual except during epidemics. This ove ...

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Protein-Polysaccharide Conjugation

The use of bacterial capsular polysaccharides as immunoprophylactic agents in human diseases caused by encapsulated bacteria is now firmly established (1). However, despite their many advantages, they do have serious limitations. First, they induce an inadequate immune response ...

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N-Propionylation

Serogroup B Neisseria meningitidis remains a major world health problem and currently there is no fully efficacious vaccine available. The poor immunogenicity of the group B meningococcal polysaccharide both in adults and infants prevents the formulation of a comprehensive poly ...

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Outer Membrane Protein Purification

The major outer membrane proteins (OMPs) from Neisseria meningitidis, which are expressed at high levels, are subdivided in five classes based on molecular weight (1,2) (see Table 1). Table 1 Major Meningococcal Outer-Membrane Proteins

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Outer Membrane Protein Vesicle Vaccines for Meningococcal Disease

Alternative strategies exist for prevention of group B Neisseria meningitidis (meningococcal) disease through vaccination (see Chapters 5, 8, 13, 14 in this volume). However, the most promising approach to date has been the use of outer-membrane vesicle (OMV) vaccines for induction of ba ...

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