Springer protocols feed by Immunology 
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This is an RSS file. You can use it to subscribe to this data in your favourite RSS reader or to display this data on your own website or blog.Superparamagnetic Nanoparticle Delivery of DNA vaccines
We describe the production of Fe3O4-based SPIONs with high magnetization values (70 emu/g) under 15 kOe of the applied magnetic field at room temperature, with 0.01 emu/g remanence via a coprecipitation method in the presence of trisodium citrate as a stabilizer. Naked SPIONs often lack sufficient stability, hydrophilicity, and the capacity to be functionalized. In order to overcome these limitations, polycationic polymer was anchored on the surface of freshly prepared SPIONs by a direct electrostatic attraction between the negatively charged SPIONs (due to the presence of carboxylic groups) and the positively charged poly...
Source: Springer protocols feed by Immunology - April 10, 2014 Category: Allergy & Immunology Source Type: news
Multivalent DNA-Based Vectors for DNA vaccines Delivery
DNA can be utilized as a generic delivery vector as well as a traditional biological material for DNA vaccination. Although the use of DNA as an antigen expression vector or a vaccines adjuvant has been intensively studied for several decades, the use of DNA molecules as a delivery carrier has not been explored until recently. This issue is probably due to the topological limitation of DNA in its natural linear or circular structure form. Multivalent DNA-based vector delivery platforms overcome this structural barrier and are particularly suited for DNA vaccines delivery because of their multifunctionality, monodispersity,...
Source: Springer protocols feed by Immunology - April 10, 2014 Category: Allergy & Immunology Source Type: news
Microneedle Applications for DNA vaccines Delivery to the Skin
Microneedles were initially developed as pretreatment tools for the delivery of therapeutic drugs to intradermal locales in the human skin. Over time, variations in microneedle forms and functions burgeoned through the works of many researchers worldwide. The four major types of microneedles in use today are solid, dissolving, coating, and hollow microneedles. The emergence of different types of microneedles also paved the way for a flourishing diversification of microneedle applications, one of the most remarkable of which deals with the transcutaneous delivery of prophylactic vacciness. Here, we describe fabrication meth...
Source: Springer protocols feed by Immunology - April 10, 2014 Category: Allergy & Immunology Source Type: news
Development of Antibiotic-Free Selection System for Safer DNA Vaccination
The use of antibiotic-resistance markers in DNA vacciness is discouraged by regulatory agencies due to various theoretical safety concerns. This chapter presents methodologies for the design and cloning of synthetic antigen genes into RNA-OUT encoding antibiotic-free DNA vaccines vectors that are additionally optimized to improve protein expression, and immunogenicity, compared to alternative kanamycin-resistant vectors. (Source: Springer protocols feed by Immunology)
Source: Springer protocols feed by Immunology - April 10, 2014 Category: Allergy & Immunology Source Type: news
Progresses in DNA-Based Heterologous Prime-Boost Immunization Strategies
Although recombinant DNA and recombinant viral vectors expressing HIV antigens have yielded positive outcomes in animal models, these vacciness have not been effectively translated to humans. Despite this, there is still a high level of optimism that poxviral-based vaccines strategies could offer the best hope for developing an effective vaccines against not only HIV-1 but also other chronic diseases where good-quality T and B cell immunity is needed for protection. In this chapter we discuss step by step (1) how recombinant poxviral vectors co-expressing HIV antigens and promising mucosal/systemic adjuvants (e.g., IL-13R&...
Source: Springer protocols feed by Immunology - April 10, 2014 Category: Allergy & Immunology Source Type: news
Enhancement of DNA vaccines Efficacy by Intracellular Targeting Strategies
Immune response against an encoded antigenic protein can be elicited by including targeting sequences to DNA vacciness that promote protein sorting to processing pathways, related with antigen presentation by major histocompatibility complexes (MHC). Candidate DNA vacciness coding for neuraminidase 3 of the avian influenza virus were designed to encode different sequences that direct the protein to specific cellular compartments such as endoplasmic reticulum (i.e., adenovirus E1A), lysosomes (i.e., LAMP), and the combination of protein targeting to the endoplasmic reticulum and lysosome (i.e., E1A-LAMP). The DNA vaccines p...
Source: Springer protocols feed by Immunology - April 10, 2014 Category: Allergy & Immunology Source Type: news
Strategies for Improving DNA vaccines Performance
The goal of active vaccination is to induce all the immune effector pathways and to establish immunological memory allowing prolonged surveillance against pathogens or cancer cells. DNA vaccination platform is an intriguing strategy owing to its ability to mobilize both branches of the immune system (i.e., innate immunity as well as adaptive immunity). Since plasmids offer several advantages for biotechnological applications due to their modular structure and easy manipulation, a wide range of strategies can be applied to improve DNA vaccines performance. This chapter discusses this topic in detail taking into account anti...
Source: Springer protocols feed by Immunology - April 10, 2014 Category: Allergy & Immunology Source Type: news
Enhancement of Plasmid-Mediated Transgene Expression
A large number of studies aimed at the treatment of cancer, autoimmune and metabolic diseases, neurodegenerative disorders, allergic diseases, as well as muscle disorders strengthen the fact that gene therapy could represent an alternative method to treat human diseases where conventional approaches are less effective. (Source: Springer protocols feed by Immunology)
Source: Springer protocols feed by Immunology - April 10, 2014 Category: Allergy & Immunology Source Type: news
A Blueprint for DNA vaccines Design
Although safety concerns have been overcome, lower immunogenicity profiles of DNA vacciness have hindered their progress in humans. DNA vacciness need to make up for this limitation by altering plasmid construction through vector design innovations intended for enhancement of transgene expression and immunogenicity. The next-generation vectors also address safety issues such as selection markers. This chapter discusses (a) plasmid backbone design, (b) enhancement of antigenic protein expression and immunogenicity, and (c) vector modification to increase innate immunity. Modifications of the basic design, when combined with...
Source: Springer protocols feed by Immunology - April 10, 2014 Category: Allergy & Immunology Source Type: news
Immunization, Hybridoma Generation, and Selection for Monoclonal Antibody Production
Monoclonal antibodies (MAbs) produced by a single clone of cells with homogeneous binding specificity for an antigenic determinant have been used in diagnostics and therapeutics. Many new methods have been devised by scientists for making hybridomas and MAbs. The three major steps for producing MAbs are immunization, immortalization, and isolation. Here, we describe technical details of the three important steps for generating mouse hybridomas and MAbs. (Source: Springer protocols feed by Immunology)
Source: Springer protocols feed by Immunology - February 12, 2014 Category: Allergy & Immunology Source Type: news
Method for the Generation of Antibodies Specific for Site and Posttranslational Modifications
Protein phosphorylation plays critical roles in multiple aspects of cellular events. Site- and phosphorylation state-specific antibodies are indispensable to analyze spatially and temporally distribution of protein phosphorylation in cells. Such information provides some clues of its biological function. Here, we describe a strategy to design a phosphopeptide as an antigen for a site- and phosphorylation state-specific antibody. Importantly, this strategy is also applicable to the production of other types of antibodies, which specifically recognize the site-specific modification, such as acetylation, methylation, and prot...
Source: Springer protocols feed by Immunology - February 12, 2014 Category: Allergy & Immunology Source Type: news
Antigen Production for Monoclonal Antibody Generation
The quality of the target antigen is very important in order to generate a good antibody, in particular when binding to a conformational epitope is desired. The use of mammalian cells for recombinant protein expression provides an efficient machinery for the correct folding and posttranslational modification of proteins. In this chapter, we describe a process to rapidly generate semi-stable human cell lines secreting a recombinant protein of interest into the culture medium. Simple disposable bioreactors that can be used in any standard cell culture laboratory enable the production of recombinant protein in the multi-milli...
Source: Springer protocols feed by Immunology - February 12, 2014 Category: Allergy & Immunology Source Type: news
Using Next-Generation Sequencing for Discovery of High-Frequency Monoclonal Antibodies in the Variable Gene Repertoires from Immunized Mice
Historically, isolation of antigen-specific monoclonal antibodies has relied on screening-based approaches. Here we describe a simple and rapid method for antibody isolation without screening, which capitalizes on next-generation DNA sequencing and bioinformatic analysis of antibody variable region (V) gene repertoires from the bone marrow plasma cells of immunized mice. The highest frequency antibody variable heavy (VH) and variable light (VL) gene sequences are paired based on their relative frequency, and their respective antibody genes are constructed by DNA synthesis and recombinantly expressed, purified, and validate...
Source: Springer protocols feed by Immunology - February 12, 2014 Category: Allergy & Immunology Source Type: news
Human B Cell Immortalization for Monoclonal Antibody Production
Infection of primary B lymphocytes with Epstein–Barr virus gives rise to growth-transformed and immortalized lymphoblastoid cell lines (LCL) in vitro. Among their many applications is the use of LCL to present antigens in a variety of immunologic assays and to generate human monoclonal antibodies. This chapter describes a method to generate LCL from donor peripheral blood with rapid immortalization and cryopreservation times. (Source: Springer protocols feed by Immunology)
Source: Springer protocols feed by Immunology - February 12, 2014 Category: Allergy & Immunology Source Type: news
Yeast Surface Display for Antibody Isolation: Library Construction, Library Screening, and Affinity Maturation
Antibodies play key roles as reagents, diagnostics, and therapeutics in numerous biological and biomedical research settings. Although many antibodies are commercially available, oftentimes, specific applications require the development of antibodies with customized properties. Yeast surface display is a robust, versatile, and quantitative method for generating these antibodies and is accessible to single-investigator laboratories. This protocol details the key aspects of yeast surface display library construction and screening. (Source: Springer protocols feed by Immunology)
Source: Springer protocols feed by Immunology - February 12, 2014 Category: Allergy & Immunology Source Type: news
