bluuebunny/crossref_metadata_2025_split huggingface dataset:
Systetnatic approaches for the identification of imprinted genes
Authors: Feinberg Andrew P 
Published: November 2023

Abstract
Genomic imprinting has been shown to influence development, growth, and behaviour in mammals as a result of parent of origin-dependent expression of imprinted genes (1-3). Some human genetic diseases and cancers are also attributable to genomic imprinting (4, 5). These effects are apparently due to loss or gain of function of imprinted genes when one or more of these genes affect specific tissues and developmental stages. How many imprinted genes in total are there throughout the lifespan in mammals and what are their precise functions? How are imprinted genes regulated and what are the molecular mechanisms controlling their expression? Answers to many of these questions are still rudimentary. For further progress in the field, it is desirable to identify imprinted genes so that a comprehensive understanding of the mechanism and role of genetic imprinting in development and disease can be attempted.
lmprinting in the Prader-Willi/Angelman syndrome region on human chromosome 15
Authors: Horsthemke Bernhard 
Published: November 2023

Abstract
The Prader-Willi syndrome (PWS) and the Angelman syndrome (AS) are distinct neurogenetic disorders which are caused by a deficiency of paternal (PWS) or maternal (AS) contributions for chromosome 15. The affected genes are located in an imprinted chromosomal domain, which is controlled by an imprinting centre. Mutations of this centre block the paternal - maternal or the maternal - paternal imprint switch during gametogenesis. DNA methylation appears to be instrumental in regulating individual gene activity and can be employed for diagnostic purposes.
The p53 pathway, past and future
Authors: Lane David P 
Published: November 2023

Abstract
Mutation of the p53 tumour suppressor gene and the accumulation of the p53 protein are amongst the most common specific changes found in human cancer. An understanding of the precise biochemical and biological affects of these changes will provide new insights into the fundamental nature of the neoplastic process. New results show that the p53 protein is a critical part of a damage-induced cell cycle checkpoint control that maintains genetic stability. The p53 protein provides an attractive target for the development of new diagnostic and therapeutic agents that will hopefully reduce the impact of cancer on society.
Happy mapping
Authors: Dear Paul H 
Published: November 2023

Abstract
Most methods for genome mapping rely on some form of cloning to isolate a subfraction of the genome for analysis, whether into yeast or bacterial hosts (as in physical mapping, Chapters 10 and 11), into hybrid cells (as in radiation hybrid mapping, Chapter 4), or as offspring amongst which polymorphic markers segregate (as in linkage mapping, Chapters 1-3).
Fluorescence in situ hybridization
Authors: Leversha Margaret a. 
Published: November 2023

Abstract
In situ hybridization enables the visualization of a probe of interest within a cytological context. For the purposes of gene mapping, the probe is usually a particular segment of cloned DNA which is labelled to allow detection. The target is normally a preparation of chromosomal DNA on a microscope slide. Both target DNA and probe are denatured, and then annealed together to allow the labelled probe to hybridize to the complementary sequences in the target. Unbound probe is then washed away and the specific hybridization signals revealed. Non-isotopic probe labelling methods, with the advantages of improved speed and safety, have largely replaced the earlier isotopic procedures (1-3). An increasing range of fluorescent labelling or cytochemical detection systems enables simultaneous hybridization of multiple probes to a single cytological preparation (4).
Fixation of tissues for the polymerase chain reaction
Authors: Hopwood David 
Published: November 2023

Abstract
Cell and tissue fixation was originally devised to produce good morphology. The aims of fixation have been defined variously but a number of features are in common (1,2):to prevent autolysis and bacterial attack to preserve cell shape and volume during subsequent procedures to retain as life-like a condition as possible to facilitate subsequent processing and staining These aims are met in part and workable and reproducible morphological results achieved. Some substances, such as lipids, glycogen, and iron, are lost to different degrees during routine processing. Nonetheless, the microanatomy of the tissues is well enough preserved to allow a histopathological diagnosis to be established in most cases. The needs of hospital histopathology are the processing of large numbers of specimens received under a variety of conditions and from a multitude of anatomical sites. The system must be robust, simple, and reproducible. It is important to remember that it has been devised and has evolved to give good morphology rather than the optimum preservation of DNA and RNA.
Spectroscopic characterization of immobilized proteins
Authors: Bright Frank V 
Published: November 2023

Abstract
As a nation the United States spends billions of dollars annually on diagnostic assays. Unfortunately, most of these assays are performed in well-outfitted laboratories, and require skilled personnel, large amounts of costly reagents, and often demand long analysis times to quantify clinically important analytes. In order to overcome the disadvantages inherent with this approach, one must develop new sensing schemes that are reliable, inexpensive, fast, simple to construct and operate, redundant and self-checking, accurate, and precise, with adequate detection limits. Finally, the demand of a given sensing scheme to be used in point of care or field situations requires small, simple, and robust sensing and detection platforms.
Immobilization using electrogenerated polymers
Authors: Schuhmann Wolfgang 
Published: November 2023

Abstract
The reproducible immobilization of biological recognition elements, preserving their catalytic activity and biological selectivity on the surface of a suitable transducer, is a fundamental presupposition for the successful development of biosensors. As future applications will aim at miniaturized biosensors and multisensor arrays which integrate several different biological recognition elements, immobilization procedures have to be developed which allow one: (a) To pre-define the immobilization site (i.e. the miniaturized transducer surface) avoiding manual deposition procedures.
Analysis of protein: protein interactions by gel electrophoresis
Authors: Coghlan Vincent M 
Published: November 2023

Abstract
With the application of modern techniques for ultrastructural imaging and macromolecular characterization has come an appreciation for the high degree of structural organization found within cells. An emerging theme in cellular organization is that for many metabolic pathways, cells utilize large multienzyme complexes that are maintained through specific protein: protein interactions (1). Defining these interactions has become, and will remain, an essential part of the functional characterization of proteins.
Engineering of novel neurotrophins
Authors: Ilag Leopold L , Ibanez Carlos F 
Published: November 2023

Abstract
Neurotrophins constitute a class of growth factors that is essential for neuronal survival and differentiation in both the central and peripheral nervous systems. This class of proteins includes nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and neurotrophin-4 (NT-4). Neurotrophins influence the differentiation, survival, and regeneration of vertebrate neurones and although there are specific activities associated with each neurotrophin, overlapping functions also exist. Illustrating this point is the fact that all members of the family support, in varying degrees, the survival of embryonic neural crest-derived sensory neurones, NGF.specifically supports the survival of embryonic sympathetic neurones but not placode-derived sensory neurones, which are supported by BDNF and NT-3 (1).
Immunolocalization and RT-PCR for the detection and quantification of growth factor and receptor gene expressi•on
Authors: Myers Simon R , Navsaria Harshad A 
Published: November 2023

Abstract
In many research fields that impact directly on clinical medicine (e.g. wound healing) a new emphasis on the examination of human tissue has emerged facilitated by the availability of more sophisticated laboratory techniques. This immediately defines the limiting factor in choosing an assay as the volume of human tissue available. To investigate human growth factor and re ceptor gene expression in any depth, assessment of both protein and messenger RNA (mRNA) for ligand and receptor must be made, in terms both of quantity and of distribution, on small tissue samples. 6 mm punch biopsy wounds of skin for example can be sutured directly, or converted into a small ellipse. In the former case the sutures tend to pull out to leave a pale, circular scar, and in the latter a linear scar of&lt; 1 cm length remains. A 6 mm punch biopsy of skin provides around 60 mg wet weight of tissue. For protocols that require repeated biopsy of an individual, 3 mm punch biopsies are preferable. These can be left to heal by secondary intention, leaving a minimal ‘pit’ scar. Note: researchers must meet all local and/or national ethical requirements before taking biopsies for experimental purposes from human subjects.
Nucleotide- and dye-ligand chromatography
Authors: Mattiasson Bo , Galaev Igor Yu. 
Published: November 2023

Abstract
A.s the level of competition between the manufacturers of rival biotechnological products increases, downstream processing costs, which can account for up to 80% of the overall production costs, will become more and more critical in determining product competitiveness (1). Since the late 1960s bio specific separation methods have come into focus and have been continuously improved. A fundamental advantage of affinity techniques is their predictive and rational character, since the ligand selected is designed to interact specifically with the protein to be purified. Among main important developments in affinity techniques is the use of more robust adsorbents and coupling chemistry that are not prone to leaking immunogenic degradation products. One such group of robust adsorbents is presented by matrices with nucleotide-ligands or dye-ligands, the latter exhibit a remarkable propensity to bind, sometimes specifically, to a plethora of proteins and enzymes.
Immunoaffinity chromatography
Authors: Cutler Paul 
Published: November 2023

Abstract
The principle of immunoaffinity or immunoadsorption chromatography is based on the highly specific interaction of an antigen with its antibody (1). Immunoaffinity chromatography is a specialized form of affinity chromatography and as such utilizes a ligand (either antigen or antibody) immobilized onto a solid support matrix in a manner which retains its binding capacity. Although the technique includes the separation of antibodies using immobilized antigens (2-4), it is more commonly performed for the identification, quantification, or purification of antigens (Figure 1). The crude extract is pumped through the column and the unbound material washed clear prior to elution of the retained antigen by alterations to the mobile phase conditions which weaken the antibody-antigen interaction.
Purification of DNA-binding proteins
Authors: Hanoa Hiroshi , Yamaguchi, Yuki , Wada Tadashi 
Published: November 2023

Abstract
Sequence-specific DNA-binding proteins are involved in many biological processes, such as DNA repair, recombination, replication, and transcription. To characterize these processes, it is quite important to purify the products that participate in them to homogeneity. The purified proteins can then be used to analyse their biochemical properties, to raise antibodies against them, and to isolate the genes encoding them.
Preparation of liposomes
Authors: Lasch J , Weissig V , Brandl M 
Published: November 2023

Abstract
Store organic solutions of phospholipids in a glass container layered with argon or nitrogen below -20 °c, preferably at -78 °C. The closure for the vial should be lined with Teflon. Alternatively, the vial can be sealed with Parafilm®. Never store organic solutions in polymer or plastic containers as this will leach impurities out of the container material (2). When transferring a portion of the material, remove the container from the freezer and allow it to reach room temperature before opening the bottle.
Long-Circulating Sterically Protected Liposomes
Authors: Klibanov Alexander L , Torchilin Vladimir P , Zalipsky Samuel 
Published: November 2023

Abstract
Soon after liposomes were proposed as vehicles for drug delivery and targeting, their major disadvantage was discovered: Typical liposome compositions rapidly exited the bloodstream and accumulated in the Kupffer cells in the liver, as well as in spleen macrophages (RES) (1). Various approaches to overcome this problem have been tested. Initially, the simple approach of blocking RES cells by administering an excess of ‘empty’ liposomes, a colloid dispersion or dextran sulfate was tested (2, 3). However, blocking RES is not the most appropriate approach for human therapy. On the other hand, to target liposomal pharmaceuticals to organs other than the liver and spleen, liposomes have been modified with specific targeting moieties, e.g. antibodies. However, immunoliposomes also do not exhibit prolonged circulation times and fail to accumulate sufficiently in the targets with limited blood supply and/or low antigen concentration (i.e. tissues and organs that require increased circulation times for liposome-to-target interaction). It is evident that a longer circulation time increases the probability that the liposomes will interact with the target tissue, yielding better targeting. Therefore, numerous attempts have been made to prolong the circulation time by varying liposome properties.
Gene expression databases
Authors: Davidson Duncan , Baldock Richard , Bard Jonathan , Kaufman Matthew , Richardson Joel E , Eppig, Janan T , Ringwald Martin 
Published: November 2023

Abstract
The rapid accumulation of large amounts of information from in situ hybridization experiments, together with the spatial and temporal complexity of this kind of data, presents serious difficulties for conventional publishing methods and for the individual researcher who wants to collate and analyse other people's results in relation to his own studies. The solution to these problems is to construct a database of gene expression and several are now being built; most apply to embryos, but in the future it is likely that they will be extended to include the adult.
Basic optical microscopy
Authors: Lacey A J 
Published: November 2023

Abstract
The perception of, or the sight of, an object whether in a microscope or in an everyday situation is the result of a complex process. It begins with light inter acting with the object. The modified light is collected by the lens of the eye and brought to a focus on the retina of the eye. The nerve responses in the retina are interpreted by the eye/brain system programmed to a greater or lesser extent by previous experience. The area of the specimen seen by the eye is the perception of the extension in x and y directions, while the experience of the of the z direction is the perception of depth.
Gene targeting vectors -considerations and use of loxP
Authors: Torres Raul M , Kühn Ralf 
Published: November 2023

Abstract
In general, the experimental outline of Cre/loxP mediated gene targeting is accomplished in two steps. The first is the incorporation of loxP sites into the genome of ES cells via a corresponding targeting vector and the identification of these homologous recombinant clones. The final genetic modification is obtained by a Cre-mediated recombination event, either stably in cultured ES cells (and the germline of mice derived from these ES cells) or in a conditional manner, in mice by the expression of Cre as a transgene. Because the potential of each targeting construct is greatly expanded with the use of this system, loxP sites should be generally considered for all targeting vectors even if the initial goal is simply to inactivate gene function.
ES cell transfection overview
Authors: Torres Raul M , Kühn Ralf 
Published: November 2023

Abstract
Thinking about addressing a biological question by gene targeting is an exciting aspect of the experiment and one that requires relatively little knowledge about ES cells and the practicalities of gene targeting. Similarly, constructing a targeting vector can also be accomplished with little knowledge of ES cells. The time when ES cells are often first considered, at least in a practical sense, is when the targeting construct is completed and you are ready to introduce the DNA into the ES cells in attempts at homologous recombination. Without any ES cell experience it is at this point that a slight trepidation is often experienced. This feeling is enhanced for the newcomer by the apparent confusion which surrounds the juggling of EF cells for each ES cell manipulation. This section attempts to provide an overview of these manipulations, including the transfection of ES cells, followed by detailed protocols which we have used in generating homologous recombinant ES cell clones and the transfer of the altered allele to the germline. Of course, once your mutation has been transmitted to the germline, the breeding, maintenance, and analysis of the mouse strain is necessary. The topic of mouse genetics is not addressed here but we refer you to an excellent book on this topic (134).
Identification of homologous recombinants
Authors: Torres Raul M , Kühn Ralf 
Published: November 2023

Abstract
One of the biggest strategical questions you must ask about your targeting experiment (preferably before vector construction) is whether or not to screen by PCR. A major advantage to screening by PCR is that the number of colonies which can feasibly be analyzed is much larger than that for screening by Southern analysis. This will of course be important if the locus you wish to target has a low efficiency of undergoing homologous recombination (for unknown reason(s)) and may make the difference between finding a homologous recombinant or not. Another advantage to PCR screening is being able to very quickly discard clones which have not homologously recombined, greatly reducing the amount of tissue culture required when screening by Southern analysis.
Generation of mice -preparation
Authors: Torres Raul M , Kühn Ralf 
Published: November 2023

Abstract
There are several methods in the literature which result in ES cell-directed development and germ.line colonization of a developing mouse blastocyst (16, 140, 147, 168-170). We describe here two methods: the physical injection of ES cells into the blastocoel of blastocysts and the aggregation of ES cells with eight-cell stage/morulas. Although perhaps not as useful for most purposes, ES cells have also been shown to be able to direct the development of a fetus when aggregated together with tetraploid embryos, possibly resulting in viable offspring (147). In our view, the decision as to which of the two general methods to use depends on personal preference but may also be determined by the equipment available, the number of chimeras that need to be produced, the ES cell line used, and the individual’s skill in micromanipulation. In contrast to blastocyst injection, aggregation methods require no expensive equipment (microscope, micromanipulators) and allow the processing of larger numbers of embryos per day. For the production of low numbers of chimeras for transmission of a targeted mutation through the germline, however, both methods are equally useful.
Characterization of macrophage antigens and receptors by immunochemistry and fluorescent analysis: expression, endocytosis, and phagocytosis
Authors: Peiser Leanne , Gough Peter J , Darley Elizabeth , Gordon Siamon 
Published: November 2023

Abstract
This chapter describes techniques to identify macrophages and measure their endocytic and phagocytic capabilities. These methods can assist in the analysis of the physiological and pathological situations in which macrophages play a central role including homeostasis, wound repair, and immunity. The expression of cell surface proteins and other molecules varies greatly among macrophage populations as a result of heterogeneity in their differentiation, activation state, or response to external influences in their microenvironment or cell culture conditions (1-3). Inununochemical and fluorescent techniques can beused to investigate the basal expression of antigens, the changes induced in response to a particular stimulus, and to detect antigens in tissues.
Standards and immunogens
Authors: Chaudhry M Q 
Published: November 2023

Abstract
Standards are preparations of analyte that are used for calibration, that is, in an assay ‘unknown’ samples are compared with them to estimate concentrations of analyte in the samples. While for hapten immunoassays standards are chem­ically quite different from the corresponding immunogens, for immunoassays that measure naturally immunogenic antigens the same purified preparation may be used as both standard and immunogen. However, the criteria for a good standard may be quite different from those for a good immunogen. Standards are considered in Section 6.
Preparation of recombinant antibodies from immune rodent spleens and the design of their humanization by CDR grafting
Authors: Leger* Olivier J P , Saldanhat Jose w. 
Published: November 2023

Abstract
The biotechnological generation of high affinity monoclonal antibodies has traditionally involved the production of hybridomas from spleen cells of immunized animals. Recent advances in recombinant technology, using phage dis play technology, have opened a new route for the generation of antibodies. Instead of immortalizing B cells for production of monoclonal antibodies, the antibody heavy (H) and light (L) chain V genes are immortalized by gene technology. The IgG mRNA of B cells of immunized animals can be used as a source of variable region (V) gene libraries. Murine VH and VL genes are amplified using PCR and used to construct libraries of millions of recombinant single chain Fvs (scFvs). ScFv antibodies consist of the variable regions of the light and heavy chains of antibodies linked via a short peptide spacer (1). Repertoires of scFv are cloned into the gene encoding the minor coat protein (gene III) of filamentous bacteriophage, thus creating a large library of phage, each displaying an individual heavy and light chain combination. The fusion protein created, consisting of the antibody fragment at the N-terminus of the coat protein, is incorporated into the phage particle (a phage antibody). Each recombinant phage genome contains the DNA encoding the specific antibody displayed on its surface, allowing the phage particle carrying antibody gene to be selected directly using the binding properties of the expressed protein.
Laboratory based methods for small scale production of monoclonal antibodies
Authors: Griffiths Bryan 
Published: November 2023

Abstract
Although hybridoma cells are spherical and well adapted to suspension culture they do grow well in stationary culture resting on the substrate, sometimes with very light attachment. This means that a wide selection of culture vessels and systems are available for their culture ranging from a simple tissue culture plate or flask to highly sophisticated bioreactors with full instrumentation to control the physiological environment (1-3). Commercially hybridoma cell lines are grown at scales up to 2000 litres and beyond in culture units scaled-up from laboratory size vessels. Laboratory, pilot, and production scale is ill-defined so in this chapter laboratory scale will be taken as 10 litre volume cultures, and below.