bluuebunny/crossref_metadata_2025_split huggingface dataset:
Non-radioactive antibody probes
Authors: Wisdom G Brian 
Published: November 2023

Abstract
The high specificity of monoclonal antibodies makes them valuable reagents, however, to exploit them fully requires a means of detecting them at low con centration. This is usually done by labelling the antibody in one of various ways. Radioisotopes are widely used (Chapter 9) as are non-isotopic labels such as enzymes, fluorogenic and luminogenic molecules, as well as various low molecular weight groups that can themselves be easily detected by a secondary reagent. The non-isotopic labels provide low detection limits in a wide range of techniques without the complications of handling radioisotopes, although it should be noted that all the non-isotopic labelling procedures have the hazards associated with protein-modifying reagents.
Monoclonal antibody therapy in organ transplantation
Authors: Wise Matt , Zelenika Diana 
Published: November 2023

Abstract
In the two decades since they were first described by Kohler and Milstein mono clonal antibodies have proved to be invaluable tools for research, not only in immunology, but also in many aspects of biomedical research. Monoclonal anti bodies revolutionized lymphocyte biology by permitting the precise characterization of cell surface molecules which until that time were ill-defined by polyclonal sera. To date numerous mAbs have been used in animal models as either immunosuppressants or agents to induce tolerance. It may therefore come as some surprise to the reader that since their inception only two mAbs have been licensed for this purpose in humans.
Analysis of protein kinase interactions using biomolecular interaction analysis
Authors: Herberg Friedrich W , Zimmermann Bastian 
Published: November 2023

Abstract
Novel technologies employing surface plasmon resonance (Biacore AB) or resonance mirrors (lAsys, Lab Systems) can be used for the characterization of protein-protein, protein-DNA and ligand-receptor interactions (for review see refs 1-3). These techniques directly measure the binding of a molecule in the soluble phase (the ‘analyte’, also called the ‘ligate’) to a ‘ligand’ molecule immobilized on a sensor surface. Thereby the association (k a) and dissociation rate constants (k d) are determined separately. With a known concentration of the analyte, apparent equilibrium binding constants (K D or K A) can be calculated. The chemistries for coupling molecules to the surface via amine, sulfhydryl, carboxyl, and other groups are well defined and reproducible. Non-covalent, site-directed immobilization can be achieved by avidin-biotin interaction, by the use of antibodies, or by binding fusion proteins to specific surfaces. Surfaces containing lipid monolayers are also available. This technology can be used for monitoring interactions, in real time without the use of labels, between two or more molecules, e.g. between proteins, peptides, nucleic acids, or pharmaceuticals.
Basic immunofluorescence
Authors: Allan Victoria J 
Published: November 2023

Abstract
Immunofluorescence on fixed samples forms a vital part of most investigations into protein function, since it provides an idea of the whereabouts of a protein within the cell, which in turn can give important clues as to the protein’s function. In addition, immunofluorescence can tell us whether this location varies through the cell cycle or development, and whether this is consistent in cells from different tissues or species. All of this information is very valuable, particularly now that genome sequencing data are available. However, in order to obtain meaningful results, and to interpret them correctly, it is important that the methods and limitations of immunofluorescence are understood.
Regulation of heterologous proteins by fusion to a hormone binding domain
Authors: Picard D 
Published: November 2023

Abstract
It is often helpful or even a necessity in biological studies to be able to regulate.: the expression or the activity of a protein of interest. Intracellular receptors by virtue of being ligand-regulated transcription factors have naturally been examined as regulatory tools. The application that is the focus of this chapter exploits the normal regulatory activity of the hormone binding domain (HBD) of vertebrate steroid receptors for the regulation of heterologous proteins in cis.Upon fusion to a heterologous protein. (he ITRD can function as an autonomous regulatory cassette and subject various protein functions to hormonal control (1. 2). Thus. one or several functional activities of the heterologous moiety may be turned offin the absence of steroid ligand. While the chimeric protein can he expressed constitutively, it only become-, active upon addition of hormone. Within at most a few minutes the hormonally-induced conformational change in the HBD results in the relieve of its protein inactivation function and activation of the chimeric protein.
Construction and screening of a subtractive cDNA library
Authors: Akopian Armen N , Wood John N 
Published: November 2023

Abstract
Subtractive library production and screening are powerful methods to isolate transcripts that are exclusively or selectively expressed either in certain tissues, or during development or in disease states. This chapter describes procedures for the isolation of specific transcripts using construction and screening of sub tractive libraries, and illustrates examples of typical problems arising during the experiments. The original method was developed by Davis and colleagues in classical work on the isolation of T-lymphocyte-specific transcripts, including a third type of murine T-cell receptor gene (1, 2). Over the years this approach has been modified. However, the principle behind the method is essentially the same, and only a few modifications have improved on the remarkable sensitivity and efficiency of the original method. Construction and screening of a sub tractive cDNA library can be divided into several steps Figure 1).
SADE: a microassay for serial analysis of gene expression
Authors: Cheval Lydie , Virion Bérangère , Elalouf Jean-Marc 
Published: November 2023

Abstract
SAGE has been shown to provide rapid and detailed information on transcript abundance and diversity (7-10). It involves several steps for mRNA purification, cDNA tags generation and isolation, and polymerase chain reaction (PCR) ampli fication. We reasoned that increasing the yield of the various extraction pro cedures, together with slight modifications in the number of PCR cycles could enlarge SAGE potentiality. Here we present a mkroadaptation of SAGE, referred to as SADE (11) as, in contrast to the original method, it allows to provide quantitative gene expression data on a small number (30000-50000) of cells.
Protein folding and human disease
Authors: Raso Stephen W , King Jonathan 
Published: November 2023

Abstract
The earliest studies on the refolding of purified proteins in the test tube identified the failure of such processes, in the form of inactive and insoluble chains (1, 2). For decades these aggregated states were thought to be unrelated to physiological processes taking place within cells. However, the recognition that the failure of the expression of cloned genes in bacteria was often due to aggregation of intracellular folding intermediates focused attention on these processes (3-5). The discovery that heat-shock proteins in almost all organisms include chaperonins that aid protein folding, established that failure of protein folding is a very general phenomenon, at least at elevated temperatures and under other stressful circumstances (6). In parallel, the discovery that amyloid deposits recovered from human tissues represent fibrillar forms of particular protein fragments revealed that protein aggregation was associated with a number of human pathologies (7, 8).
Analysis of protein phosphorylation
Authors: Walaas S Ivar , ∅stvold Anne Carine 
Published: November 2023

Abstract
Extensive studies have demonstrated that extracellular signals, e.g. hormones, neurotransmitters, growth factors, as well as intracellular signals and effectors, e.g. cyclic nucleotides, Ca2+ ions, and phospholipid derivatives, mediate many of their effects by regulating the activities of protein phosphorylation systems (1-3). The role of reversible protein phosphorylation in regulating and integrating the activity of eukaryotic cells has thus become widely appreciated during the last decades (4). Cellular protein phosphorylation systems consist of a minimum of three components: (a) The phosphorylated proteins themselves. (b) A large number of protein kinases which catalyse the incorporation of phosphate from the terminal phosphate of ATP into specific serine, threonine, tyrosine, and possibly histidine (5) residues in the substrate phosphoproteins.
Carbohydrate recognition in leukocyte-endothelial cell interactions
Authors: Homeister Jonathon W , Lowe John B 
Published: November 2023

Abstract
Mammalian cell surfaces are decorated with complex carbohydrate molecules, also known as oligosaccharides or glycoconjugates. These molecules display a remarkable degree of structural diversity, which is in turn a function of the cell type or tissue, or the particular stage of development of the cell or tissue. Cell surface oligosaccharide structure varies with the differentiation state or lineage of a cell. These observations suggest that these molecules play important roles in mediating information transfer into and out of the cell. During mammalian development, for example, dynamic changes are observed in cell surface glycoconjugate expression. This work suggests that complex mechanisms have evolved to regulate these processes, and that they have important functional correlates. The existence of an ever-expanding group of mammalian carbohydrate binding proteins, or lectins (1), suggests that cell surface carbohydrate molecules may participate as ligands in adhesive processes mediate by mammalian lectins. Indeed, oligosaccharides have been shown to operate in processes whereby lysosomal enzymes are targeted to the lysosome (via the mannose-6-phosphate receptor; ref. 2), in the clearance of desialylated serum glycoproteins (via the hepatic asialoglycoprotein receptor; ref. 3), and in the clearance of circulating pituitary glycoprotein hormones like leutropin (4). Experimental studies during the past several years have implicated members of a family of mammalian lectins, known as the selectins, and a specific group of cell surface oligosaccharide determinants, in adhesive processes that mediate leukocyte adhesion to vascular endothelium. This chapter will summarize experimental approaches that have led to the discovery of these processes, and will discuss the enzymes, selectins and other molecules that participate in carbohydrate dependent leukocyte adhesion.
Polysialic acid in neural cell developtnent: roles, regulation and tnechanistn
Authors: Bruses Juan L , Rutishauser Urs 
Published: November 2023

Abstract
Polysialic acid (PSA) was discovered as a major component of vertebrate brains, largely on the basis of its unusual composition and size (1). It is a linear homopolymer of a-2,8-linked sialic acid (from about eight to over 100 residues) attached to an otherwise typical N-linked carbohydrate core (for review, see ref. 2, see also Fig. 10 in chapter 1). However, unlike most carbohydrates found on the cell surface, PSA is attached exclusively to the polypeptide of the neural cell adhesion molecule (NCAM) (3-5). PSA’s function was first recognized through its ability to decrease NCAM mediated membrane-membrane adhesion in vitro (6, 7). It was later observed that PSA could also affect other cell interactions that do not appear to depend directly on NCAM’s intrinsic binding function (5, 8). For example, PSA can attenuate the function of the Ll class of axon-associated adhesion molecules, apparently without any direct association of L1 with NCAM or PSA.
Single-sperm typing: a rapid alternative to family-based linkage analysis
Authors: Cullen Michael , Carrington Mary 
Published: November 2023

Abstract
Single-sperm typing provides an efficient means of measuring recombination frequencies relative to the traditional methods of genetic cross and pedigree analysis. Using this method, it is possible to rapidly determine the haplotype of each meiotic product directly using PCR to identify alleles at multiple poly morphic markers along the chromosome. Traditional pedigree analysis, on the other hand, requires genotypes collected from children of parents informative (heterozygous) for markers of interest. For example, mapping resolutions of 1–2 cM were reported for the Genethon human linkage map based on 5264 microsatellites genotyped for eight CEPH (Centre d’Etudes du Polymorphisme Humaine) families, to talling 134 individuals (186 meioses) (1). However, markers used to construct a family-based linkage map must have high joint polymorphic information content (PIC), i.e. a high proportion and number of informative meioses per marker pair (2). In regions of low marker density this is problematic since the total number of markers with high PIC values is decreased. This issue becomes less relevant with the use of single-sperm typing because donors who are heterozygous at the appropriate markers can be selected regardless of the PIC value of those markers. Furthermore, unlimited numbers of meiotic events provide a means of analysing tightly linked markers at resolutions of 0.1–0.2 cM, presently not possible by pedigree analysis (3–5).
HLA typing methods
Authors: Krausa Pete , Browning Michael 
Published: November 2023

Abstract
The human major histocompatibility complex or HLA system is a set of highly polymorphic genes clustered on chromosome 6. There are three main groups of HLA genes, namely: HLA class I, class II, and class III. HLA class I and class II genes code for molecules that present antigen in the form of peptides at the cell surface. HLA class I molecules are expressed on most nucleated cells and platelets, and present antigen to CD8+ T cells. HLA class II genes are constitutively expressed on professional antigen-presenting cells (B lymphocytes, macrophages, monocytes, and dendritic cells). In addition, HLA class II expression can be induced on a range of other cell types, including activated T cells, activated endothelium, and many epithelial cells. HLA class II molecules present antigen to CD4+ T cells. HLA class III genes code for certain complement components, cytokines (TNF-a and -13), and a variety of non-immunological products, but their identification will not be covered in this chapter.
Preparing suspensions of single cells
Authors: Ormerod Collated by Michael G 
Published: November 2023

Abstract
A suspension of single cells is the essential element of flow cytometry. The sample should contain as little debris and as few dead cells and clumps as pos sible. A set of protocols for the preparation of cell suspensions has been collected in this chapter. It is by no means comprehensive but should form a good starting point for any new investigation. Extensive sets of methods have also been described by Cheetham et al. (1) and Pallavicini (2).
The first six protocols have been taken from N. P. Carter’s Chapter 3 in the first edition of this book.
Flow sorting
Authors: Carter Nigel P , Ormerodt Michael G 
Published: November 2023

Abstract
Flow sorting enables subpopulations of cells or particles to be separated from the sample suspension with a high degree of purity. The purified fractions (usually95% pure) are thus made available for morphological examination or use in functional or other assays, so greatly extending the research and diagnostic potential of the flow cytometer. Any combination of parameters can be applied as criteria to decide whether a cell is or is not sorted, thereby harnessing the full analytical potential of the instrument in the separation process. One of the main original applications of flow sorting was the purification of morphologically similar, but functionally distinct, lymphocyte subpopulations using fluorescent staining with polyclonal antibodies. However, the ability to utilize any analytical parameter or combinations of parameters has led to flow sorting being used in diverse applications, ranging from cloning rare cell transfectants directly into multi-well plates to the bulk sorting of human chromosomes for the production of DNA libraries.
The application of fluorescence in situ hybridization to chromosome analysis
Authors: Kearney L , Buckle V J 
Published: November 2023

Abstract
For over two decades banding has remained the ‘gold standard’ of cytogenetic analysis, providing the first genome-wide screen for abnormalities. However, conventional cytogenetic banding techniques are limited to the detection of rearrangements involving more than 2 Mb of DNA. In addition, the identification of de nova unbalanced chromosome rearrangements provides a particular challenge for chromosome banding to decipher. In recent years a number of techniques based around FISH have evolved, all of which complement the conventional banding approaches and which have steadily increased the accuracy of cytogenetic diagnosis.
The lymphomas and chronic lymphoid leukaemias
Authors: Harrison C J 
Published: November 2023

Abstract
Malignant lymphomas and chronic lymphoid leukacmias rcpresc not two highly heterogeneous groups of malignant disease. Both arise as a result of neoplastic proliferation of lymphoid cells. A number of chromosomal abnormalities have been associated with histopathology. disease staging, cmd prognosis. Although these are fewer than any of the other haematological disorders, cytogenetic information is increasing in importance as an aid to diagnosis and patient management. This chapter presents those techniques that have been the most successful in producing cytogenetic results and a SL1mma1y of the impo11ant cytogenetic findings.
The role of cytogenetics in the diagnosis and classification of haematological neoplasms
Authors: Bain B J 
Published: November 2023

Abstract
Cytogenetic analysis is undertaken in patients with haemalological disorders for two main reasons: firstly, to aid in the management of the patient and secondly, for the advancement of scientific knowledge. Often, both purposes are served simultaneously. The patient may benefit from cor1firmation of a provisional diagnosis and from the availability of prognostic information that can influence m;inagcmcnt. Cytogc-111:tic analysis can be very useful during follow up. firstly for monitoring the response to therapy and secondly for detecting rebpse and distinguishing relapse from a new therapy-related leukaemia. Cytogenetic analysis may also be required before patients are entered into therapeutic trials and may influence stratification within these trials.
Introduction to optical microscopy for plant cell biology
Authors: Shaw P J 
Published: November 2023

Abstract
The development of the optical microscope in the seventeenth Century opened up new areas of study in many fields of science, In particular, the observations of plant and animal tissues and micro-organisms gave rise to cell biology, although our modern idea of what a ‘cell’ actually is arose somewhat later. Some of the data recorded by the early microscopists, using only primitive microscopes and drawing freehand what they saw, are quite remarkable. Hooke, Malpighi, and Leuwenhoek are the best known seventeenth century microscopists, but Nehemiah Grew was the first true specialist in plant microscopy. He produced detailed descriptions of plant microanatomy which have proved remarkably accurate. An example is shown, along with Grew’s original legend (1), in Figure 1. Although there is only space to show one such picture, the original volumes contain many pages of such detailed, painstaking hand-engravings. With today’s access to photography, video cameras, and computer image processing, this should serve as a reminder of how much can be achieved with careful observation and a very simple microscope. Many biologists overlook the useful information that can be rapidly obtained by even a very simple laboratory microscope.
Transient expression, a tool to address questions in plant cell biology
Authors: Hadlington Jane L , Denecke Jurgen 
Published: November 2023

Abstract
Transient expression in plant systems is a rapidly expanding technique, which has opened up new strategies to approach biological problems without the necessary delay and environmental risks that are associated with the production of stable transformants. DNA is introduced into cells via naked DNA transfer or biological vectors such as bacteria or viruses, and gene expression is assayed for a limited time after the initial transfer. Some methods allow the introduction of RNA or proteins as well. Results can be obtained much faster without extensive resources and time, and in case of cell suspensions or protoplasts, experiments are more reproducible due to the high number of cells and the homogeneous nature of the suspensions. In some cases, i.e. the analysis of cytotoxins or dominant mutant approaches disrupting essential pathways or mechanisms, transient expression is the only option available, because transgenic plants would not be viable. Finally, with the current debate on the safety and necessity of genetically modified organisms, it is important to introduce alternative ways of addressing biological questions in plants. In many cases, transient expression has been shown to provide ultimately the same information as approaches using transgenic plants, thus justifying a more consequent and widespread use of the technique.
Electrophysiological methods: monitoring exo- and endocytosis in real time
Authors: Thiel* Gerhard , Sutter, Jens-Uwe , Homann* Ulrike 
Published: November 2023

Abstract
About two decades ago Neher and Marty (1) showed that the patch-clamp technique in combination with some impedance analysis could be used for monitoring the membrane capacitance as a quasi real time assay for exocytosis and endocytosis. This method allows recording of exo- and endocytosis in single animal cells in vivo with such a high resolution, that single exo- and endocytotic events from vesicles with a diameter under 100 nm can be recorded with a temporal resolution in the order of some 10 msec. This method has been adopted with success in plant physiology laboratories to examine these processes in plant protoplasts (reviewed in ref. 2). Here we aim to present the range of problems which can be addressed with this methodology and provide guidance to the methodological approach.
Organelle isolation
Authors: Robinson David G , Hinz Giselbert 
Published: November 2023

Abstract
Subcellular fractionation has, as its general principle, the application of three consecutive procedures. The first, homogenization, results in the abolition of cellular integrity. Depending on the forces involved, individual compartments or organelles may also no longer remain intact. The second step entails imposing a new system of order among the randomly dispersed organelles in the homogenate. This is usually achieved through separations based on size (gel filtration), mass (differential or rate zonal centrifugations), density (isopycnic centrifugation), or surface charge (electrophoresis, phase-partitioning). Finally, the subcellular units or fractions obtained have to be checked for their homogeneity. For this, electron microscopical (negatively stained or thin sectioned samples), chemical (enzyme tests), and immunological (Western blotting) methods can be employed.