bluuebunny/crossref_metadata_2025_split huggingface dataset:
Stability, Storage, and Sterilization of Liposomes
Authors: Zuidam Nicolaas Jan , Winden Ewoud van , Vrueh Remco de , Crommelin Daan J A 
Published: November 2023

Abstract
During the preparation process or upon storage liposomes may undergo chemical degradation or physical changes. In this chapter, the types of degradation, the consequences, and measures of prevention or minimization are discussed. Methods to establish the degree of degradation are presented in Chapter 2. Most of the relevant data on liposome stability published in the public domain was collected with bilayers based on phosphatidylcholine, phosphatidylglycerol, and phosphatidylethanolamine often in combination with cholesterol. Therefore, our conclusions are particularly relevant for liposomes composed of those lipids. A number of review articles/chapters dealing with liposome stability issues have been published by our group (1-5).
Vesicular Phospholipid Gels
Authors: Brandl M , Massing U 
Published: November 2023

Abstract
This chapter discusses some of the common limitations with small liposomes as drug carriers and how vesicular phospholipid gels can be used to circumvent them. The purpose of an intravenously injected liposomal drug carrier usually is to circulate in the bloodstream and carry the drug to the desired target organ or tissue. Under ideal circumstances the pharmacokinetics and biodistribution of the carrier-associated drug is merely dependent on the characteristics of the liposomes, not on the drug itself. Size and surface characteristics of the liposomes primarily determine the degree to which they are eliminated from the bloodstream by macrophages and end up in the RES organs spleen and liver (1, 2). Big liposomes (&gt;200 nm) are rapidly eliminated, thus they are inappropriate for i.v. drug delivery. Smaller liposomes are more likely to escape the RES and may thus circulate long enough to reach targets within or nearby the vasculary bed (2). Smaller liposomes furthermore are more likely to extravasate.
Liposome-based DNA Vaccines: Procedures for Entrapment
Authors: Gregoriadis Gregory , McCormack Brenda , Perriea Yvonne , Bacon Andrew , Caparros-Wanderley, Wilson , Zadi Brahim 
Published: November 2023

Abstract
Numerous workers (1-3) have shown that intramuscular injection of naked plasmid DNA elicits humoural and cell-mediated immune responses against the encoded antigen. It appears {2, 3) that promotion of immunity is the result of DNA uptake by muscle cells which leads to the expression and extracellular release of the antigen to be subsequently taken up by antigen-presenting cells (APC). It is also believed (3) that some of the injected DNA is taken up directly by APC. There are two main disadvantages of immunization with naked DNA. First, DNA is taken up by only a minor fraction of muscle cells which, at any rate, are not professional APC. Secondly, naked DNA is exposed to deoxyribonuclease in the interstitial fluid thus necessitating the use of relatively large quantities of DNA.
Detection of mutations by hybridization with sequencespecific oligonucleotide probes
Authors: Saiki Randall K , Erlich Henry A 
Published: November 2023

Abstract
Genetic analysis by hybridization with sequence-specific oligonucleotide probes (SSOP) is one of the oldest methods commonly used to examine nucleic acids for the presence of known sequence variants (1, 2). The procedure takes advantage of the fact that under the appropriate reaction conditions, a short oligonucleotide probe will hybridize to its target only when it is perfectly matched; a single base pair mismatch is often sufficiently destabilizing to prevent a stable probertarget duplex from forming. By testing a specimen with a pair of probes—one complementary to the normal sequence and one to the mutant—the genotype at that position can be determined. Using a panel of probes, an entire series of mutations or polymorphisms can be surveyed.
Functional assay of the p53 tumour suppressor gene
Authors: Frebourg Thierry , Flaman Jean-Michel , Estreicher, Anne , Iggo Richard 
Published: November 2023

Abstract
Screening methods generally used to detect mutations are commonly based on structural analysis of genes. These methods are often labour-intensive and fail to distinguish inactivating mutations from polymorphisms or functionally silent mutations. The alternative is to use biological assays which test the function of the gene product. This chapter describes a functional assay which detects mutations in human p53 cDNA expressed in the yeast Saccharomyces cerevisiae. Somatic p53 mutations are the most common genetic defect known to occur in human tumours (for a review see ref. 1) and more than 2000 different mutations have been described. Germline p53 mutations predispose to the development of multiple tumours and are responsible for the majority of cases of the familial Li-Fraumeni cancer syndrome.
Analysis of antigen processing and presentation
Authors: Kaye P M 
Published: November 2023

Abstract
This chapter seeks to identify the key issues relating to the study of macrophages as antigen-presenting cells (APC) and presents protocols for the major assays used to measure antigen processing and presentation. In addition, it discusses some of the theoretical and practical issues related to the processing and presentation of intracellular pathogens for which macrophages are the major host cell. The methods presented are, for the most part, those used in the author’s laboratory or by colleagues. Whilst these provide working examples, it is stressed that the nature of individual host-pathogen interactions may necessitate alterations on a case-by-case basis. Potential areas for such modifications are highlighted, as appropriate.
Pattern recognition: classification of chromosomes
Authors: Graham Jim 
Published: November 2023

Abstract
The topic of pattern recognition is often associated with image analysis. Indeed several textbooks combine the topics (1, 2) and the titles are often used interchangeably. While the two topics intersect, they are distinct. Pattern recognition concerns classification: assigning labels to things on the basis of their properties. This is clearly important in image analysis, and Chapter 3 would not have been complete without some discussion of classification. In this chapter I will deal with classification methods and associated issues, which could be applied more widely ( e.g. in signal analysis, speech analysis, medical diagnosis, etc.). However, in keeping with the context of the book it will be illustrated by examples from image analysis. There are many textbooks which deal with the topic of pattern recognition in great depth, from the classic work of Duda and Hart (1 ), through several worthy contributions (2-5) to the recent comprehensive expositions of Ripley (6) and Bishop (7), which relate the topic to neural networks. The purpose of this chapter is to present a ‘user’s guide’, for which I will draw on all of this substantial body of work. I will make little attempt at mathematical or statistical justification and direct myself to those topics which I believe most ‘users’ (as distinct from pattern recognition practitioners) would need to be aware of. Where appropriate I will guide readers with an appetite for a greater depth of exposition to the appropriate sources. I have tried to keep mathematical and statistical argument to a minimum, but have found it necessary in defining some basic terms to make use of some vector-matrix notation. Where this has occurred I have tried to explain what it means. I do not think that a reader who is familiar with the basic concepts of probability, mean, and covariance will find their understanding impaired.
Projective stereology in biological croscopy
Authors: Carothers Andrew D 
Published: November 2023

Abstract
Stereology can be defined as the three-dimensional interpretation of flat images, such as sections and projections, by criteria of geometric probability. Although the definition encompasses both sections and projections, the two approaches differ in both theoretical underpinning and practical application. Most standard theory is concerned almost exclusively with sectioned material. No doubt this is a result of the early historical development of the subject, which was largely driven by geological applications where the main requirement was to determine patterns of inclusions within solid rocks. Sectioning is necessary wherever there is a need to determine the internal composition of large, opaque objects, such as body organs. There is little point in attempting to reproduce here the many useful results from this body of theory. For these, the reader is referred to standard texts (see for example refs 1-3). In many other applications of biological microscopy, however, it is more relevant to consider plane projections since sectioning is invasive and may be technically difficult, and since objects of interest, such as subcellular elements, whole cells, and tissues, are often transparent or nearly so. Another, and perhaps the earliest, application of the stereology of projections was the inference by the ancient Greeks that the earth was spherical from the shape of its shadow on the moon during lunar eclipses.
Baculovirus-mediated overexpression of transport proteins
Authors: Litherland* Gary J , Baldwintꝉ Stephen A 
Published: November 2023

Abstract
A major limitation to our understanding of the molecular mechanisms of membrane transport processes is the lack of detailed structural information on transport proteins, such as might be obtained using X-ray crystallography and other biophysical techniques. One reason for the paucity of structural studies on transporters using such techniques has been the difficulty in producing sufficient quantities (10-100 mg) of pure transport protein. The natural abundance of most transporters is low, and purification difficult. While many prokaryotic transporters have been successfully expressed at high levels in Escherichia coli (see Chapter 6), the use of bacteria to overexpress complex, often glycosylated or otherwise modified membrane proteins from eukaryotic sources has rarely been successful.
Choosing and characterizing antibodies
Authors: Forde* A , Coley† J 
Published: November 2023

Abstract
Choosing an antibody or antibodies is usually the most important preliminary to developing any immunoassay. The assay developer may have just raised antibodies for the particular application, or candidate antibodies may have been obtained from one of the many commercial and noncommercial organizations that produce antibodies. In either case, the best must be selected, or they all must be rejected and the search continued. An immunoassay is defined primarily by its analyte molecule, which can range in size from an intact cell such as a microorganism, to a natural antigen (which could be almost any size above about 2 kDa), to a specific antibody and to a peptide or small organic molecule.
Labels and endpoints
Authors: Abdul-Ahad* Wajdi , Brett† Martin 
Published: November 2023

Abstract
The sophistication of current immunoassay methodology allows the assay developer a wide choice of labelling substances and assay endpoints, so that it is not usually difficult to find a combination that well suits the specification required for a new assay. This chapter will deal primarily with. the three major kinds of labels and endpoints that are used when immunoassays for research purposes are being developed, i.e. radioisotopic (1), enzymatic (2) and fluorescent (3). Information on the preparation of ligand labels containing biotin and fluorescein (4) is also given.
Solid-phase reagents
Authors: Matson Robert S 
Published: November 2023

Abstract
This chapter is concerned with the preparation and use of solid-phase reagents in immunoassay. The merits in using a solid-phase, as well as the various approaches to preparing solid-phase reagents will be addressed. Information on the kinds of supports available will be given. Finally, detailed protocols for various formats (microtitre plates, membranes, beads, tubes) will be provided, along with sources of commercial solid-phase reagents.
Analysis of signal transduction pathways using protein—serine/threonine phosphatase inhibitors
Authors: Grahame Hardie D 
Published: November 2023

Abstract
A widely used approach for studying protein phosphorylation is the pharmacological one, i.e. to study the effects of cell-permeable inhibitors or activators of the enzymes which catalyse changes in phosphorylation. In Chapter 2 the use of protein kinase activators and inhibitors is discussed. Over the past 10 years, a number of potent, cell-permeable protein phosphatase inhibitors have also become available, and their impact has been considerable. Their applications may be summarized as follows: (a) Improving the specificity of protein phosphatase assays in cell-free extracts. This approach is discussed in Chapter 7, and the present chapter will concentrate instead on their uses in intact cell studies.(b) Provision of initial evidence that a physiological system is regulated by protein phosphorylation. This is particularly valuable in the case of a system where the major components are not well characterized, in which case more direct methods for the study of protein phosphorylation cannot be utilized.(c) In cases where the role of protein phosphorylation is already established, the inhibitors may in some cases be used to identify the protein phosphatase(s) active against particular substrates in intact cells.
Assay and purification of protein (serine/threonine) phosphatases
Authors: Mackintosh Carol , Moorhead Greg 
Published: November 2023

Abstract
The eukaryotic protein phosphatases comprise several families of enzymes that catalyse the dephosphorylation of intracellular phosphoproteins, thereby controlling diverse cellular functions in all eukaryotic cells. The activities of these enzymes are highly regulated in response to extracellular stimuli, and in many cases regulation is mediated by interactions of the catalytic subunits with diverse targetting and regulatory subunits and domains. The protein phosphatases responsible for dephosphorylation of serine and threonine (and histidine) residues in the cytoplasmic and nuclear compartments of eukaryotic cells are derived from two distinct gene families, PPP and PPM, that have been classified both by phylogenetic analysis of their sequences, and by their three-dimensional structures (reviewed in refs 1 and 2).
Analysis of protein kinase specificity using oriented peptide libraries
Authors: Songyang Zhou , Cantley Lewis C 
Published: November 2023

Abstract
This chapter focuses on the use of oriented peptide libraries to study the specificities of protein kinases. This approach not only predicts an optimal sequence from a single experiment, with no prior knowledge of in vivo phosphorylation sites required, but it also provides information about the relative importance of each position for selectivity, and about which amino acids are tolerated. In the following sections we will explain peptide library design strategies, detail the experimental techniques for peptide synthesis and selection, and discuss how the data obtained are interpreted.
Phosphatidylinositol 3-kinase signalling: a tale of two kinase activities
Authors: Coffer Paul J 
Published: November 2023

Abstract
Over the last 25 years considerable emphasis has been placed on the phosphorylation of proteins on tyrosine and serine/threonine residues as a mechanism of regulating cellular function. The importance of protein phosphorylation is highlighted in the other chapters of this book. However, there is another class of kinases that has recently taken centre stage, those kinases that phosphorylate phosphoinositide (Pl) lipids. In the original concept of membrane structure, it was thought that the membrane proteins were solely responsible for regulating membrane functions. More recently however, membrane lipids have been found to play not only a structural role, by isolating the intracellular environment from the extracellular, but also a critical role in the generation of second messengers. For example, the hydrolysis of phosphatidyl inositol (4,5)-bisphosphate(Ptdlns(4,5)P2) to diacylglycerol(DAG) and inositol(l,4,5)P3 results in the release of Ca2+ from intracellular stores and the activation of protein kinase C (PKC). While the significance of Ptdlns(4,5)P2 hydrolysis has long been appreciated, the role of Pl-lipid phosphorylation is only now beginning to be fully understood. The reason for the relatively slow progress in this field of research is that, unlike protein kinases that were first cloned in the 1980s, the first PI-lipid kinase, phosphatidylinositol 3-kinase (Pl3K), was only cloned in 1992. This chapter will focus on recent discoveries concerning the structure and regulation of PI3K and its role in cellular functioning.
Genetic approaches to protein kinase function in lower eukaryotes
Authors: Plyte Simon E 
Published: November 2023

Abstract
The study of protein kinase function in higher eukaryotes such as Mammalia has been generally restricted to biochemical analysis of purified proteins, due to the limitations of genetic approaches in these species. Most of these enzymes were first identified biochemically and their genes isolated subsequent to peptide sequence analysis. However, the promiscuous nature of protein kinases in phosphorylating proteins in vitro has complicated study of their function in cells. In contrast, the tractability of lower eukaryotes to genetic analysis has resulted in a distinct approach in which genes involved in a given process are initially identified via their phenotypic effects. These genetic strategies, of course, do not specifically target protein kinase genes but this family of proteins has surfaced at a surprising frequency during analysis of regulatory genes, which is a testimony to their adaptability and ubiquity as biological tools. Once identified, the relative ease of genetic manipulation in lower eukaryotes enables the investigation of kinase function in vivo. Thus, mutational analysis can be used to determine whether a gene product is required in a particular pathway. Additionally, genetic epistatic experiments, between two genes in a particular pathway, may be applied to determine the order of action of those gene products. During the last decade, functional homologues of many mammalian protein kinases have been identified in lower eukaryotes, allowing a degree of extrapolation in the workings of the proteins in different species. In fact many of these kinases seem to function in an identical manner in lower eukaryotes and participate in evolutionary conserved kinase cascades, vindicating the use of lower eukaryotes as a model system in which to study protein kinase function.
Specificity in JAK-STAT signalling
Authors: Halupa Adrienne , Barber Dwayne L 
Published: November 2023

Abstract
Since the original identification of Janus kinases by Wilks et al. (l, 2) and the isolation of STATl and STAT2 by the Darnell laboratory, an entire new field of discovery has emerged and come to dominate research in cytokine signalling during the last decade. This wealth of information now provides us with some clues as to the function of these novel tyrosine kinases and transcription factors. To date, four Janus kinases (Fig. 1) and seven STAT (Fig. 2) transcription factors have been identified in mammalian species. With the exception of TYK2 and STAT2, gene-targeting experiments have been reported for all members of the JAK-STAT family. Drosophila expresses a Janus kinase homologue, called hopscotch, indicating that the JAK-STAT pathway is conserved throughout evolution. The STAT transcription factors are more widespread, having been isolated in Caenorhabditis elegans, Drosophila spp., and Dictyostelium spp.
Differential display analysis of alteration in gene expression
Authors: Cho Yong-Jing , Liang Peng 
Published: November 2023

Abstract
Temporal and spatial expression of the 100000 different genes in the genome of a mammal is a highly regulated process which determines the normal develop ment of an organism. The alterations in this process are therefore often the cause of developmental and pathological abnormalities such as cancer. Two-dimensional protein gel electrophoresis was developed for the purpose of comparative studies. However, the limitation of this method became evident mostly due to its sensitivity problem. as at most only up to 2000 of 10-15000 different types of protein expressed in any mammalian cell can be detected. Therefore, the majority of the genes, especially those expressed at lower levels, may not be represented. One other major pitfall of the method is the difficulty of being able to retrieve enough protein spots identified for future molecular characterizations, although there have been many recent technical advances in these areas (see Chapters 9 and 10 for further discussion of these issues).
Identification of differentially expressed genes by suppression subtractive hybridization
Authors: Desai Sejal , Hill Jason , Trelogan Stephanie , Diatchenko Luda , Siebert Paul D 
Published: November 2023

Abstract
SSH features several improvements over other cDNA subtraction methods that have been described in the literature (1-5). SSH eliminates any intermediate steps for physical separation of single-stranded (ss) and double-stranded (ds) cDNAs, requires only one round of subtractive hybridization, and can achieve greater than a 1000-fold enrichment for differentially expressed cDNAs (6-8). To overcome the problem of differences in mRNA abundance, SSH incorporates a hybridization step that normalizes (equalizes) sequence abundance by standard hybridization kinetics during the course of subtraction. This dramatically in creases the probability of obtaining low-abundance differentially expressed cDNAs and simplifies analysis of the subtracted library.
Gene expression analysis by cDNA microarrays
Authors: Mousses Spyro , Bittner Michael L , Chen Yidong , Dougherty Ed R , Baxevanis Andreas , Meltzer Paul S , Trent Jeffrey M 
Published: November 2023

Abstract
The Human Genome Project (1) has spurred the emergence of ‘genome-wide’ approaches to study gene function, regulation and interaction termed functional genomics. Comprehensive analysis of differential gene expression is becoming a cornerstone of functional genomics. To this end, several new technologies have been developed for high-throughput differential gene expression analysis (2-8). The deposition of cDNAs in an array format to filters or glass is a hybridization based analytical tool at the forefront of this field. The parallel gene expression of arrayed cDNAs on glass and subsequent analysis by means of two-colour fluor escence was pioneered in the laboratory of Dr P.O. Brown at Stanford. The first proof of principle paper showed gene expression analysis for an array with 45 Arabidopsis genes (2). This approach has since grown rapidly to high density microarrays with tens of thousands of genes. Today, many laboratories around the world have established cDNA microarrays for a range of applications and found it to be very useful for simultaneously profiling mRNA levels for thousands of genes.
Analysis of gene expression by two-dimensional gel electrophoresis
Authors: Tolkovsky A M 
Published: November 2023

Abstract
Two-dimensional (2-D) polyacrylamide gel electrophoresis using isoelectric focus ing (IEF) in the first dimension separates proteins by charge and size. Although simple in concept, the technique has been under-used for three main reasons: (i) the diffirulty in obtaining highly reproducible patterns of isoelectric focusing in the first (pH) dimension using tube gels and soluble carrier ampholytes; (ii) the lack of good analytical tools for analysis and assignation of differential patterns of gene expression; and (iii) inability to obtain sequencing information from other than the most abundant proteins. In the last few years, however, the practical and analytical tools have changed to such an extent that analysis of the proteome by 2-D gels can (and should) now be considered an accessible technique.