Defined as red blood cell break down and the release of hemoglobin and intracellular contents into the plasma, haemolysis can seriously impact patient care as well as the laboratory's reputation through its affect on test results. Therefore, the European Preanalytical Scientific Committee, in collaboration with the International Federation of Clinical Chemistry Working Group on Patient Safety, have designed a questionnaire to collect data on prevalence and management of haemolytic specimens referred to the clinical laboratories for clinical chemistry testing. The new book will help identify the areas where haemolysis occurs most frequently, which can, in turn, guide further analysis about why it is occurring. Once these elements are known, practices and procedures can be implemented to dramatically reduce haemolysis and avoid erroneous laboratory results affecting patient care and increasing laboratory costs.

During the past two decades, many books, governmental reports and regu lations on safety measures against chemieals, fire, microbiological and radioactive hazards in laboratories have been published from various coun tries. These topics have also been briefly discussed in books on laboratory planning and management. The application ofvarious scientific instruments based on different ionizing and non-ionizing radiations have brought new safety problems to the laboratory workers of today, irrespective of their scientific disciplines, be they medicine, natural or life sciences. However, no comprehensive laboratory handbook dealing with aIl these hazards, some of which are recently introduced, had so far been available in a single volume. Therefore, it was thought worthwhile to publish this Handbook on safety and health measures for laboratories, with contributions from several experts on these subjects. As this second edition of the Handbook, like the first edition, is a multiauthor volume, some duplication in conte nt among chapters is unavoidable in order to maintain the context of a chapter as weIl as make each chapter complete. An attempt has also been made to maintain the central theme, which is how to work in a laboratory with maximum possible environmental safety."

Weareenteringintoaneraofre-awakeningoftheimportanceandinterestinthe role ofaltered intermediary metabolisminthedevelopmentandprogressionof malignancy. Therefore, thisbookMitochondriaandCanceristimelyandrelevant tocontemporaryandfutureresearch, training, andeducation. Thehallmarkstudies ofOttoWarburgetal. (1926)sparkedtheeraoftumorcellmetabolism. Fromthen untilaround1980, studiesofintermediarymetabolismofnormalandmalignant cellsweremajorareasofresearchandofgraduateandpostgraduatetrainingin biomedicalsciences. Thisperiodwasdominatedbymitochondrialstudiesandby thoseextraordinaryoutstandingscientistswhowereaffectionatelyreferredtoasthe "mitochondriacs." However, theadvent, development, andsubsequentdominance ofmoleculargenetics/molecularbiologyandmoleculartechnologyinbiomedical researchwereaccompaniedbythenearlycompletesubmersionofinterestinareas ofintermediarymetabolismandtumorcellmetabolism. Forthepast30orsoyears, graduateandpostgraduatetraining, experienceandfocusinbiochemistry, meta- lism, enzymology, cellphysiologyhavedeclined, beingdisplacedbythedo- nanceofmoleculartechnologies. Therecentrevelationsandadvancesinmolecular genetics/molecularbiologyandthedevelopmentofmolecularandnanotechnology providenewdimensionstoapplytotheunderstandingandelucidationoftheroleof intermediarymetabolismandmitochondrialfunctioninthedevelopmentandp- gressionofcancers. Toaddresstheissuesoftumormetabolism, thepresentand futuregenerationsofresearchersmustintegratetheknowledgeandmethods of biochemistrywiththeknowledgeandtechnologyofmolecularbiology. Thefocusonmitochondriaiscentraltodiscussionsandissuesofintermediary metabolism. Thegenetictransformationofnormalfunctionalcellstoneoplastic cellsimposestheneedforalterationsinintermediarymetabolismandmitochond- alfunctiontoprovidethebioenergetic/synthetic/growthrequirementsforthep- gression of the neoplastic cell to malignancy. In the absence of the metabolic transformation, theneoplasticmalignantcellwillnotachieveitsmalignantpot- tial. Thus, itiscriticaltoestablishtheroleofthemitochondriainthemalignant process. Moreover, speci?calterationsindifferentcancertypesmustbeidenti?ed. Such information will provide new approaches to the treatment and perhaps preventionofdifferingcancers. TheadventofMetabolomicsforthedetectionof v vi Preface cancer and progression of malignancy is dependent upon the elucidation and identi?cationofalteredmetabolismandmitochondrialfunction. Thus, thisexciting journeyincancerresearchhasbarelybegunandhasalongwaytogo. Thisbook, Mitochondria and Cancer, and its subject matter provide a roadmap and the preparationtoproceedonthisjourney. Wearegreatlyindebtedtothecontributingauthorsfortheirenthusiasticsupport andwritingachapterintheirareaofexpertise. WearegratefultoMs. DonnaOvak for secretarial help and our families for their patience and support in putting togetherthisbook. Thisbookisdedicatedtopast, present, andfuture"mitoch- driacs"aroundtheworld. Buffalo, NY KeshavK. Singh Baltimore, MD LeslieC. Costello Contents MitochondriaandCancer. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 KjerstinM. Owens, J. S. Modica-Napolitano, andKeshavK. Singh WarburgandhisLegacy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23 MichaelRistowandTimJ. Schulz TheLipogenicSwitchinCancer. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39 KoenBrusselmansandJohannesV. Swinnen CitrateMetabolisminProstateandOtherCancers . .

The purpose of the book is to provide an overview of clinical research (types), activities, and areas where informatics and IT could fit into various activities and business practices. This book will introduce and apply informatics concepts only as they have particular relevance to clinical research settings.

John Sinclair and a panel of expert investigators present a comprehensive collection of cellular and molecular techniques for the analysis of cytomegalovirus (CMV) biology and its pathogenetic mechanisms. The methods-all described in step-by-step detail with ready reproducibility in mind-range from basic virus culture to complex molecular analysis of CMV structure and function. Included are methods for CMV detection using both immunological and biological techniques, methods for analyzing fundamental aspects of the CMV infection cycle, and methods for analyzing T cell response to cytomegalovirus infection in the human host. Comprehensive and state-of-the-art, Cytomegalovirus Protocols provides investigators with a collection of the key methods that are illuminating not only the basic biology of this complex and intriguing human herpesvirus, but also its significant role in human infectious diseases and their emergent therapies.

When Volume 1 (Toxicolpgy) in this series of Standard Operating Procedures was pub lished in early 1979, the FDA's Good Laboratory Practice Regulations did not have the force of United States Law, but nevertheless had a substantial impact on the conduct of toxicology in laboratories throughout the world. These Regulations are now in force, and Volume 2 (Pathology) was published later the same year. Our critics have implied that we have attempted to reduce toxicology to the level of the cookery book, or alternatively that we seek to impose our standards on others, In some sinister way ensuring that the IRI code will become the international norm. We dismiss these criticisms as arrant nonsense. The many thousands of volumes already sold worldwide can provide at best a framework for adaptation to suit local laboratory condi tions, and thus speed to GLP compliance those organisations which might otherwise have remained foundering at the starting post. If Volumes 1 and 2 of this series have con tributed anything to the conduct of toxicology it must surely be in those non-English speaking nations which, because of the international nature of pharmaceutical and chemical trading, are required by commercial pressures to be in compliance with a foreign law formulated in unfamiliar terminology and introduced for reasons that are not immediately obvious. Much has happened in the short period of time since Volumes 1 and 2 were published."

This is the fourth volume of Standard Operating Procedures (SOPs) compiled from documents prepared in these laboratories in part fulfilment of the requirements of various Good Laboratory Practice (GLP) regulations and guidelines. SOPs have now become an everyday feature of work in most industrial and contract toxicology laboratories. They provide a written definition of the mechanics of unit operations which together comprise the framework for experiments in safety evaluation. Metabolic studies and analytical chemistry are closely linked to toxicology since they embody essential aspects of the overall assessment of product safety. Some authorities consider certain parts of these subjects to be outwith the scope of the GLP requirements but for the reasons stated this is contrary to our own view. We have tried where possible to define in SOP format for use in our own laboratories the unit operations involved in these disciplines and they form the basis of this volume. Some relevant material from previous volumes has been brought together in updated form and is also presented here for completeness. Dr I P Sword Managing Director Inveresk Research International Musselburgh EH21 7UB Scotland ix Introduction GENERAL 1. The Food and Drug Administration of the US Government published its Good Laboratory Practice Regulations for Non-Clinical Laboratory Studies in the Federal Register (22 December 1978). The Regulations are the culmin ation of a number of years of investigation into the standards to which safety evaluation studies were performed in laboratories in the USA."

The XIIIth International and the VIIth European Congress of Clinical Chemistry took place at the Netherlands Congress Centre in The Hague, from June 28th to July 3rd 1987. The Organizing Committee and the Scientific Committee for these combined congresses aimed to present the state-of-the-science as well as the state-of-the-art for those fields of clinical chemistry which show a strong progress and which will most probably inflict a great part of all clinical chemists. "Clinical Chemistry, an Overview" comprises almost all papers which were presented during the congress in 5 plenary lectures and 97 lectures during 24. symposia. The invited speakers, being experts in their fields of clinical chemistry, succeeded very well in presenting an overview over the newest developments in connection to the knowledge already known, thereby demonstrating the progress made in clinical chemistry during the last years. The Editors take great pleasure in thanking once more the members of the Scientific Committee and of the International Scientific Advisory Board in creating an excellent scientific programme for this congress. The Editors also take great pleasure in thanking all those whose afforts have made possible the publication of this book. We are most grateful to all speakers who also prepared a manuscript for publication. The Editors also appreciate the most helpful and encouraging attitude of Plenum Press Publishers Corporation."

9. REFERENCES 90 CHAPTER 9: OPPORTUNITIES FOR REPLACEMENT, REDUCTION AND REFINEMENT: HUMAN VIRAL VACCINES 93 1. INTRODUCTION 93 2. TESTS FOR EXTRANEOUS MICROORGANISMS 94 3. MUMPS VACCINE . . . 94 4. HEPATITIS B VACCINE 95 4. 1. Test for freedom from live hepatitis virus 95 4. 2. Potency test 95 5. INFLUENZA VACCINE 96 5. 1. Potency test 96 5. 2. Abnormal-toxicity test 96 6. MEASLES VACCINE. . . . . 96 6. 1. Tests for extraneous agents 97 6. 1. 1. Test for Mycobacterium tuberculosis 97 6. 1. 2. Test for extraneous viruses 97 6. 2. Abnormal-toxicity test 98 7. POLIOMYELITIS VACCINE. . . 99 7. A. Inactivated poliomyelitis vaccine 99 7. A. l. Production. . . . . . 99 7. A. 2. Test for residual live virus 99 7. A. 3. Tests for extraneous agents 100 7. A. 3. l. Test for Mycobacterium tuberculosis . . . . . 100 7. A. 3. 2. Test for extraneous viruses 100 7. A. 4. Tumourigenicity test. . . . . 100 XII 7 . A. 5. Potency test . . . . . . . 101 7. A. 6. Test for abnormal toxicity 102 7. B. Live, oral poliomyelitis vaccine 103 7. B. l. Test for extraneous agents 103 7. B. 2. Test for neurovirulence 105 7. B. 3. Test for abnormal toxicity 106 8. RABIES VACCINE 106 8. 1. Production 106 8. 2. Test for residual live virus 107 8. 3. Tests for extraneous microorganisms 107 8. 3. 1. Test for Mycobacterium tuberculosis 107 8. 3. 2. Tests for extraneous viruses 108 8. 3. 2. 1. In dogs 108 8. 3. 2. 2.

Enzyme-linked immunospot assay (ELISPOT) has been known for some time as a unique state-of-the-art technique for studying the cytokine-secreting activity of immune system cells, and it appears to be one of the fast growing applications in biomedical research, becoming an indispensable tool in vaccine development, HIV research, transplantation studies, and cancer and allergy research. The second edition of Handbook of ELISPOT: Methods and Protocols, only the second book in the field which is entirely dedicated to ELISPOT assay, shares the detailed techniques that have been developed since the release of the popular first edition. Straight from the labs of seasoned experts, this book covers setting and performing ELISPOT assays, ELISPOT for veterinary research, advanced ELISPOT techniques, image and data analysis, as well as vaccine development and diagnostics. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters include introductions to their respective chapters, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Handbook of ELISPOT: Methods and Protocols, Second Edition serves as a compilation of a technical reference and a troubleshooting guide for researchers, both experienced and novice, worldwide in order to advance the usage of this key tool.

The focus of this symposium was on the present and future capabilities of flow cytometry for both medical and biological applications in cancer. This technology began with quite modest instrumentation, with limited capabilities to answer biological questions. Today, both the clinical workhorses and the powerful multi-laser, multi-detector, sorting machinery, coupled with sophisticated computers and storage devices and the increasing storehouse of markers and dyes, are taking us to the limit and beyond in finding answers to the cause and cure of cancer. In the past, both normal hematopoietic tissue and leukemias have been the tissue samples of choice in the application of flow cytometry, and some of the most recent applications with these tissues are presented here. However, the book also discusses the increasingly sophisticated disaggregation techniques which allow investigators the possibility to train their lasers on solid tumors. Not only can we use flow cytometry with associated fluorescent markers to understand the biology of cancer, but also the wide array of existing and developing markers provides us with important diagnostic tools in the detection of cancer early in either the malignant or relapse process. And the field comes full circle, with the use of the technology for gene mapping and other genetic studies to unlock the basic malignant process.

Radiophannaceuticals labeled with short-lived radionuclides are utilized to unravel biochemical processes, and to diagnosis and treat diseases of the living body are-developed through extensive evaluation in ~iological models. 'fhC first attempt to compile information was a volume entitled ANIMAL MODELS IN RADIOTRACER DESIGN that was edited by William C. Eckelman and myself in 1983. The volume had a focus on the animal models that investigators were using in order to design radiotracers that displayed in vivo selectivity as measured by biodistribution and pharmacokinetic studies. A concern in the early days of nuclear medicine was species differences. Often a series of labeled compounds were evaluated in a several different animal models in order to gain confidence that the selected radiotracer would behave appropriately in humans. During the past 12 years there have been remarkable advances in molecular genetics, molecular biology, synthetic radiopharmaceutical chemistry, molecular modeling and visualization, and emission tomography. Biological models can now be selected that are better defined in terms of molecular aspects of the disease process. The development of high resolution PET and SPET for clinical applications facilitates the development of new radiopharmaceuticals by the use of models to quantitatively evaluate drug effects, and progression of disease, and hence to arrive at better diagnosis and treatments for animals and humans. With these advances there is an effective use of biological models, and the refinement of alternatives for the development of new radiophannaceuticals.

This volume provides guidance and answers to frequently asked questions in infectious diseases, thus facilitating improved patient care, prudent and cost effective management and investigation of these disorders. Other more complicated but less common conditions are also reviewed. Uniquely, this volume directly discusses several controversies regarding infectious diseases from the 21st century.

Volume57 in the internationally acclaimed "Advances in Clinical Chemistry" contains chapters submitted from leading experts from academia and clinical laboratory science. Authors are from a diverse field of clinical chemistry disciplines and diagnostics, ranging from basic biochemical exploration to cutting-edge microarray technology.
Written by authors representing the diverse field of clinical chemistry and diagnostics, reviews in Advances in Clinical Chemistry cover a range of cutting-eduge research ranging from basic biochemical exploration to microarray technology."

Volume56 in the internationally acclaimed "Advances in Clinical Chemistry" contains chapters submitted from leading experts from academia and clinical laboratory science. Authors are from a diverse field of clinical chemistry disciplines and diagnostics, ranging from basic biochemical exploration to cutting-edge microarray technology.
Written by authors representing the diverse field of clinical chemistry and diagnostics, reviews in Advances in Clinical Chemistry cover a range of cutting-eduge research ranging from basic biochemical exploration to microarray technology."

Despite the many milestones in cystic fibrosis (CF) research, progress towards curing the disease has been slow, and it is increasingly difficult to grasp and use the already wide and still growing range of diverse methods currently employed to study CF so as to understand it in its multidisciplinary nature. Cystic Fibrosis: Diagnosis and Protocols aims to provide the CF research community and related researchers with a very wide range of high-quality experimental tools, as an easy way to grasp and use classical and novel methods applied to cystic fibrosis. Volume I: Approaches to Study and Correct CFTR Defects focuses on the cystic fibrosis transmembrane conductance regulator (CFTR) and its expression, biogenesis, structure, and function in terms of the defects causing CF. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Comprehensive and practical, Cystic Fibrosis: Diagnosis and Protocols will provide readers with optimal working tools to address pressing questions in the best technical way, while helping all of us, as a research and clinical community, to move faster hand-in-hand toward unravelling the secrets of this challenging disorder and cure it.

Despite the many milestones in cystic fibrosis (CF) research, progress toward curing the disease has been slow, and it is increasingly difficult to grasp and use the already wide and still growing range of diverse methods currently employed to study CF so as to understand it in its multidisciplinary nature. Cystic Fibrosis: Diagnosis and Protocols aims to provide the CF research community and related researchers with a very wide range of high-quality experimental tools, as an easy way to grasp and use classical and novel methods applied to cystic fibrosis. Volume II: Methods and Resources to Understand Cystic Fibrosis focuses on pathophysiology, Omics approaches, and a variety of key resources recently made available for CF research. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Comprehensive and practical, Cystic Fibrosis: Diagnosis and Protocols will provide readers with optimal working tools to address pressing questions in the best technical way, while helping all of us, as a research and clinical community, to move faster hand-in-hand toward unravelling the secrets of this challenging disorder and cure it.

Volume55 in the internationally acclaimed "Advances in Clinical Chemistry" contains chapters submitted from leading experts from academia and clinical laboratory science. Authors are from a diverse field of clinical chemistry disciplines and diagnostics, ranging from basic biochemical exploration to cutting-edge microarray technology.
Leading experts from academia and clinical laboratory science Volume emphasizes novel laboratory advances with application to clinical laboratory diagnostics and practical basic science studies"

Recently, important new findings in the polyamine field and a variety of new experimental systems have revolutionized the study of these ubiquitous cellular components, essential for normal growth and development. In "Polyamines: Methods and Protocols," leading researchers contribute an extensive collection of up-to-date laboratory techniques for the further pursuit of polyamine study. The volume delves into vital subjects such as neoplasia studies with animal models and human patients, therapeutic roles for polyamine inhibitors and analogs, polyamine metabolism and oxidative damage, polyamines as regulators of critical ion channels, as well as polyamine transport systems and polyamine-responsive genes. Written in the highly successful "Methods in Molecular Biology " series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and expert notes on troubleshooting and avoiding known pitfalls.

Comprehensive and cutting-edge, "Polyamines: Methods and Protocols" provides a key resource for all scientists pursuing the study of this dynamic and significant aspect of cellular biology."

This volume includes a comprehensive theoretical treatment and current state-of-the art applications of the quartz crystal microbalance (QCM). It discusses interface circuits and the study of viscoelasticity and micromechanics as well as surface roughness with the QCM. Coverage also details the broad field of analytical applications of piezoelectric sensors.

Research in the pharmaceutical sciences and medicinal chemistry has taken an important new direction in the past two decades with a focus on large molecules, especially peptides and proteins, as well as DNA therapeutics. In Drug Design and Discovery: Methods and Protocols, leading experts provide an in-depth view of key protocols that are commonly used in drug discovery laboratories. Covering both classic and cutting-edge techniques, this volume explores computational docking, quantitative structure-activity relationship (QSAR), peptide synthesis, labeling of peptides and proteins with fluorescent labels, DNA-microarray, zebrafish model for drug screening, and other analytical screening and biological assays that are routinely used during the drug discovery process. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters include introductions to their respective topics, lists of the necessary materials, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Thorough and accessible, Drug Design and Discovery: Methods and Protocols serve as a vital laboratory reference for pharmaceutical chemists, medicinal chemists, and pharmacologists as well as for molecular biologists.

Volume 53 in the internationally acclaimed "Advances in Clinical Chemistry" contains chapters submitted from leading experts from academia and clinical laboratory science. Authors are from a diverse field of clinical chemistry disciplines and diagnostics, ranging from basic biochemical exploration to cutting-edge microarray technology.

Leading experts from academia and clinical laboratory science
Volume emphasizes novel laboratory advances with application to clinical laboratory diagnostics and practical basic science studies