Download or read book Bioreactors for Stem Cell Expansion and Differentiation written by Joaquim M.S. Cabral and published by CRC Press. This book was released on 2018-09-03 with total page 279 pages. Available in PDF, EPUB and Kindle. Book excerpt: An international team of investigators presents thought-provoking reviews of bioreactors for stem cell expansion and differentiation and provides cutting-edge information on different bioreactor systems. The authors offer novel insights into bioreactor-based culture systems specific for tissue engineering, including sophisticated and cost-effective manufacturing strategies geared to overcome technological shortcomings that currently preclude advances towards product commercialization. This book in the fields of stem cell expansion, bioreactors, bioprocessing, and bio and tissue engineering, gives the reader a full understanding of the state-of-art and the future of these fields. Key selling features: Describes various bioreactors or stem cell culturing systems Reviews methods for stem cell expansion and differentiation for neural, cardiac, hemopoietic, mesenchymal, hepatic and other tissues cell types Distinguishes different types of bioreactors intended for different operational scales of tissue engineering and cellular therapies Includes contributions from an international team of leaders in stem cell research

Download or read book Bioreactor Systems for Tissue Engineering II written by Cornelia Kasper and published by Springer Science & Business Media. This book was released on 2010-10-03 with total page 331 pages. Available in PDF, EPUB and Kindle. Book excerpt: Alternative Sources of Adult Stem Cells: Human Amniotic Membrane, by S. Wolbank, M. van Griensven, R. Grillari-Voglauer, and A. Peterbauer-Scherb; * Mesenchymal Stromal Cells Derived from Human Umbilical Cord Tissues: Primitive Cells with Potential for Clinical and Tissue Engineering Applications, by P. Moretti, T. Hatlapatka, D. Marten, A. Lavrentieva, I. Majore, R. Hass and C. Kasper; * Isolation, Characterization, Differentiation, and Application of Adipose-Derived Stem Cells, by J. W. Kuhbier, B. Weyand, C. Radtke, P. M. Vogt, C. Kasper and K. Reimers; * Induced Pluripotent Stem Cells: Characteristics and Perspectives, by T. Cantz and U. Martin; * Induced Pluripotent Stem Cell Technology in Regenerative Medicine and Biology, by D. Pei, J. Xu, Q. Zhuang, H.-F. Tse and M. A. Esteban; * Production Process for Stem Cell Based Therapeutic Implants: Expansion of the Production Cell Line and Cultivation of Encapsulated Cells, by C. Weber, S. Pohl, R. Poertner, P. Pino-Grace, D. Freimark, C. Wallrapp, P. Geigle and P. Czermak; * Cartilage Engineering from Mesenchymal Stem Cells, by C. Goepfert, A. Slobodianski, A.F. Schilling, P. Adamietz and R. Poertner; * Outgrowth Endothelial Cells: Sources, Characteristics and Potential Applications in Tissue Engineering and Regenerative Medicine, by S. Fuchs, E. Dohle, M. Kolbe, C. J. Kirkpatrick; * Basic Science and Clinical Application of Stem Cells in Veterinary Medicine, by I. Ribitsch, J. Burk, U. Delling, C. Geißler, C. Gittel, H. Jülke, W. Brehm; * Bone Marrow Stem Cells in Clinical Application: Harnessing Paracrine Roles and Niche Mechanisms, by R. M. El Backly, R. Cancedda; * Clinical Application of Stem Cells in the Cardiovascular System, C. Stamm, K. Klose, Y.-H. Choi

Download or read book Stem Cell Manufacturing written by Joaquim M.S. Cabral and published by Elsevier. This book was released on 2016-07-24 with total page 342 pages. Available in PDF, EPUB and Kindle. Book excerpt: Stem Cell Manufacturing discusses the required technologies that enable the transfer of the current laboratory-based practice of stem cell tissue culture to the clinic environment as therapeutics, while concurrently achieving control, reproducibility, automation, validation, and safety of the process and the product. The advent of stem cell research unveiled the therapeutic potential of stem cells and their derivatives and increased the awareness of the public and scientific community for the topic. The successful manufacturing of stem cells and their derivatives is expected to have a positive impact in the society since it will contribute to widen the offer of therapeutic solutions to the patients. Fully defined cellular products can be used to restore the structure and function of damaged tissues and organs and to develop stem cell-based cellular therapies for the treatment of cancer and hematological disorders, autoimmune and other inflammatory diseases and genetic disorders. Presents the first ‘Flowchart‘ of stem cell manufacturing enabling easy understanding of the various processes in a sequential and coherent manner Covers all bioprocess technologies required for the transfer of the bench findings to the clinic including the process components: cell signals, bioreactors, modeling, automation, safety, etc. Presents comprehensive coverage of a true multidisciplinary topic by bringing together specialists in their particular area Provides the basics of the processes and identifies the issues to be resolved for large scale cell culture by the bioengineer Addresses the critical need in bioprocessing for the successful delivery of stem cell technology to the market place by involving professional engineers in sections of the book

Download or read book Development of Miniaturized Bioreactors for Stem Cell Culture written by Mariam Sarvi and published by . This book was released on 2014 with total page 416 pages. Available in PDF, EPUB and Kindle. Book excerpt: Embryonic stem cells (ESCs) are pluripotent cells capable of indefinite self renewal in vitro while maintaining the ability to differentiate into cell types of all three germ layers. ESCs are outstanding options of in vitro cell models for regenerative medicine, functional genomics, human developmental biology and drug discovery study. Stem cell research is among the most promising fields of biotechnology, and which provides the potential of developing novel approaches to repair or replace damaged tissues and cells. The present day, exponentially growing effort of stem cell research emphasizes a major need for convergence of more efficient and appropriate laboratory technologies to sustain the growth, proliferation and differentiation potential of stem cells.Although so far, a variety of conventional bioreactors with different configurations (such as spinner flasks, rotary, perfusion bioreactor, etc.) have been designed and adapted for stem cell expansion and differentiation, bioreactors can be disadvantageous in bench-top research because they need large space, consume huge amount of reagents and need more time to operate and maintain (sterilizing, cleaning, assembling, and disassembling of the bioreactor components). The requirements for costly equipment and generating shear stress due to the fluid flow, and the lack of physical similarity between microenvironments of bioreactor and actual cell microenvironment, make using bioreactors undesirable.In tissue engineering, micro-scale technology is an approach that combines micro-techniques with materials science and surface engineering, and results in a profound exploration of the microenvironment where cells are embedded. These technologies are able to address some of the limitations imposed by conventional tissue engineering methods. Indeed, developing successful novel small-scale technologies for in vitro cultivation of different types of cells can assist in increasing our knowledge on conditions that control stem cell growth and differentiation and organ development. In fact, small scale bioreactors are miniaturized versions of conventional bioreactors, where high- throughput cell based assays can be carried out at low cost compared with their macro-scale counter- parts.The first aim of this thesis was to develop a disposable miniaturized bioreactor through a novel and inexpensive method for effective stem cell proliferation. To this end, an effective surface functionalization method was developed for enhancing the biocompatibility of a PDMS surface that is protein resistant while facilitating cell proliferation (expansion) and maintaining the pluripotency potential of cells. The micro-bioreactor was fabricated in the form of a fixed bed bioreactor with a microchannel reactor bed. The microchannel was functionalized to enable cell adhesion and resistance to protein adsorption. The functionalized surface was found to be biocompatible with cancer and embryonic stem cells (ESCs), and while facilitating cell proliferation.Differentiation of ESCs into a variety of cell types is an important characteristic of these types of cells, which is commonly achieved in vitro by spontaneously self-assembling in low adhesion culture conditions into 3D cell aggregates called embryoid bodies (EBs). Formation of EBs that simulates many of the characteristics of early embryonic development is considered as a vital step to induce differentiation of stem cells. Formation of three dimensional configurations of ESCs as EBs provides possibilities to mechanistically study early differentiation events of pluripotent cells. In fact, EB formation is of paramount importance for in vitro investigation of the embryonic development and differentiation of both the mouse and human ES cells.The second aim of this thesis was to introduce a novel concept of a miniaturized bioreactor made of liquid marble (LM). A novel application of liquid marbles for formation of embryoid bodies (EBs) was then presented. This study showed that the confined internal space of liquid marble, along with the porous and non-adhesive property of the highly hydrophobic liquid marble shell, can facilitate the formation of uniform EBs inside the liquid marbles. The efficiency of liquid marble-born-EBs compared to the liquid suspension (LS) technique as the chosen control method in terms of cell viability and EB uniformity revealed that cells in liquid marble are more viable than those in suspension. Measuring EB size distribution as an indicator of uniformity also confirmed that EBs obtained by the LM are morphologically more uniform and of a narrower size distribution compared to those formed in LS. The feasibility of using liquid marble bioreactors for cardiomyocyte differentiation of mouse ES (mES) cells after formation of EBs inside the liquid marble was further investigated. The results demonstrated that ES cells can differentiate into myocyte cells through the liquid marble as a facile, cost effective, and straightforward method. We proposed for the first time that liquid marbles greatly contribute to ES cardiac differentiation, which provides a new technology platform for ES biology and genetic studies.It is worth mentioning that although the majority of our knowledge in modern biology has been provided by classical two dimensional (2D) cell culture techniques, the most common negative aspects of these systems is deficiency of support from extra-cellular matrix, which represents an important role for cell growth and development. It is now well accepted that cells reside, proliferate, and differentiate in complex 3D microenvironments. The concept of using three dimensional (3D) biodegradable scaffolds as alternatives for extracellular matrix (ECM), which more closely reform cells' native structure, is an interesting area of study in current tissue engineering. Because of their unique function, stem cells need to reside in the specialized, 3D microenvironment that surrounds them in native tissues.The third section of this thesis (chapter 5) focuses on the investigating of the feasibility of forming embryoid bodies using a novel hydrogel as bioreactor embedding material. This hydrogel is porous and biodegradable and is prepared by modifying hydroxypropylcellulose (HPC), with bio-functional methacrylates (MA). Observation of EB formation inside hydrogel implied that the stem cells attached and penetrated to the pores, and proliferated well, while forming uniform EBs. Uniformity of EBs formed inside hydrogel, compared with those formed via liquid suspension (LS) method, as one of the most widely used EB formation techniques. It was observed that porous hydrogel allows the formation of more homogeneous EBs. It was found that cells inside hydrogel-born EBs are more viable compared to those formed in LS method. Expression of germ markers via quantitative PCR and immunostaining confirmed that the hydrogel-born EBs had expressed 3 germ layers with further in vitro differentiation potential. These EBs were allowed to differentiate further in hydrogel, where positive immunostaining of different cardiac markers and observation of beating EBs showed the potential of EBs to further differentiate into cardiac cells lineage.In summary, this thesis first presents a novel, facile and cost effective method via surface bio- functionalization of PDMS bioreactor for better stem cell adhesion and proliferation, and later introduces two novel methods to prepare bioreactor material, namely liquid marble and porous hydrogel (HPC-MA) for formation of embryoid bodies, which is considered as a critical step for in vitro differentiation in ESCs.

Download or read book Cell Engineering and Regeneration written by Heinz Redl and published by Springer. This book was released on 2017-02-16 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: This reference work presents the origins of cells for tissue engineering and regeneration, including primary cells, tissue-specific stem cells, pluripotent stem cells and trans-differentiated or reprogrammed cells. There is particular emphasis on current understanding of tissue regeneration based on embryology and evolution studies, including mechanisms of amphibian regeneration. The book covers the use of autologous versus allogeneic cell sources, as well as various procedures used for cell isolation and cell pre-conditioning , such as cell sorting, biochemical and biophysical pre-conditioning, transfection and aggregation. It also presents cell modulation using growth factors, molecular factors, epigenetic approaches, changes in biophysical environment, cellular co-culture and other elements of the cellular microenvironment. The pathways of cell delivery are discussed with respect to specific clinical situations, including delivery of ex vivo manipulated cells via local and systemic routes, as well as activation and migration of endogenous reservoirs of reparative cells. The volume concludes with an in-depth discussion of the tracking of cells in vivo and their various regenerative activities inside the body, including differentiation, new tissue formation and actions on other cells by direct cell-to-cell communication and by secretion of biomolecules.

Download or read book Stem Cell Systems Bioengineering written by Tiago G. Fernandes and published by Frontiers Media SA. This book was released on 2021-07-02 with total page 214 pages. Available in PDF, EPUB and Kindle. Book excerpt: Dr. Stephanie Willerth has a commercialization agreement with Aspect Biosystems with regards to bioprinting stem cell derived tissues. Dr. Yuguo Lei is a co-founder of CellGro Technologies, LLC, a company focusing on cell expansion technologies. Dr. Tiago Fernandes has no competing interests with regards to this Research Topic.

Download or read book Stem Cells From Mechanisms To Technologies written by Michal K Stachowiak and published by World Scientific. This book was released on 2011-12-19 with total page 394 pages. Available in PDF, EPUB and Kindle. Book excerpt: Advances in stem cell biology and biotechnology have sparked hopes that therapies will soon be available for maladies which were considered incurable before. However, realization of the clinical potential of stem cells will require better understanding of stem cell physiology and the development of advanced technologies for their efficient differentiation in medically relevant quantities. Resolving these issues necessitates synergistic approaches from multiple fields. Systems biology can be employed to dissect the mechanisms regulating the genome and proteome of stem cells during self-renewal and commitment. Microfluidic platforms can be used to recreate aspects of the stem cell niche and obtain a better understanding of the interactions among stem cells and with their environment. The milieu of stem cells and their progeny can be shaped with appropriately designed biomaterials for the engineering of tissues to replace, reconstitute or regenerate damaged organs. To that end, enabling bioreactor technologies will be necessary for the generation of large quantities of stem cells and their derivatives in a robust and cost-efficient manner. This book invites world-renowned experts in the above fields to discuss the latest advances in their respective areas and to provide insights on the future challenges and achievements in the area of stem cells.

Download or read book Stem Cells written by Michal K. Stachowiak and published by World Scientific. This book was released on 2012 with total page 394 pages. Available in PDF, EPUB and Kindle. Book excerpt: Advances in stem cell biology and biotechnology have sparked hopes that therapies will soon be available for maladies which were considered incurable before. However, realization of the clinical potential of stem cells will require better understanding of stem cell physiology and the development of advanced technologies for their efficient differentiation in medically relevant quantities. Resolving these issues necessitates synergistic approaches from multiple fields. Systems biology can be employed to dissect the mechanisms regulating the genome and proteome of stem cells during self-renewal and commitment. Microfluidic platforms can be used to recreate aspects of the stem cell niche and obtain a better understanding of the interactions among stem cells and with their environment. The milieu of stem cells and their progeny can be shaped with appropriately designed biomaterials for the engineering of tissues to replace, reconstitute or regenerate damaged organs. To that end, enabling bioreactor technologies will be necessary for the generation of large quantities of stem cells and their derivatives in a robust and cost-efficient manner. This book invites world-renowned experts in the above fields to discuss the latest advances in their respective areas and to provide insights on the future challenges and achievements in the area of stem cells.

Download or read book Stem Cell Engineering written by David Schaffer and published by CRC Press. This book was released on 2012-11-20 with total page 233 pages. Available in PDF, EPUB and Kindle. Book excerpt: While the potential of stem cells is recognized, their proliferation and differentiation must be more precisely controlled to maximize the production of therapeutically relevant cells and for cell replacement therapies to minimize contamination with residual cells that can give rise to side effects. How can engineers make contributions to address these challenges? With contributions from pioneers and experts, Stem Cell Engineering: Principles and Practices highlights recent advances in the understanding of the cellular and molecular composition of the stem cell niche, as well as approaches to build upon this basic information to direct stem cell differentiation into therapeutically valuable lineages. The growing recognition of stem cells as an important and exciting field will continue to draw investigators with diverse backgrounds—from biology, engineering, and the physical sciences—and thereby enable further progress in these and other new directions. This book discusses advances made during the last decade that have led to increasingly defined culture systems for growing stem cells, starting from co-culture with feeder cells in the presence of serum to growth on synthetic substrates in defined medium. In addition to highlighting many recent advances, it underscores the need for future work.

Download or read book Mesenchymal Stem Cells Basics and Clinical Application II written by Birgit Weyand and published by Springer. This book was released on 2014-07-08 with total page 309 pages. Available in PDF, EPUB and Kindle. Book excerpt: Engineered MSCs from Patient-Specific iPS Cells, by Irina Eberle, Mohsen Moslem, Reinhard Henschler, Tobias Cantz Fate of Intravenously Injected Mesenchymal Stem Cells and Significance for Clinical Application, by Beate Wagner, Reinhard Henschler The Implications of Stem Cell Applications for Diseases of the Respiratory System, by Mei Ling Lim, Philipp Jungebluth, Paolo Macchiarini Potential of Mesenchymal Stem Cell Applications in Plastic and Reconstructive Surgery, by Birgit Weyand, Peter M. Vogt General Principles for the Regeneration of Bone and Cartilage, by Michael Jagodzinski, C. Haasper Adult Mesenchymal Stem Cells Explored in the Dental Field, by K. M. Fawzy El-Sayed, C. Dörfer, F. Fändrich, F. Gieseler, M. H. Moustafa, H. Ungefroren Mesenchymal Stem Cell Therapy and Lung Diseases, by Khondoker M. Akram, Sohel Samad, Monica Spiteri, Nicholas R. Forsyth Mesenchymal Stem Cells as Cellular Immunotherapeutics in Allogeneic Hematopoietic Stem Cell Transplantation, by Claudia Papewalis, Daniela Topolar, Barbara Götz, Stefan Schönberger, Dagmar Dilloo New Cell-Based Therapy Paradigm: Induction of Bone Marrow-Derived Multipotent Mesenchymal Stromal Cells into Pro-Inflammatory MSC1 and Anti-inflammatory MSC2 Phenotypes, by Aline M. Betancourt Interactions Between Mesenchymal Stem Cells and Dendritic Cells, by Grazia Maria Spaggiari, Lorenzo Moretta MSC and Tumors: Homing, Differentiation, and Secretion Influence Therapeutic Potential, by Naomi D’souza, Jorge Sans Burns, Giulia Grisendi, Olivia Candini, Elena Veronesi, Serena Piccinno, Edwin M. Horwitz, Paolo Paolucci, Pierfranco Conte, Massimo Dominici Sources of Mesenchymal Stem Cells: Current and Future Clinical Use, by Michela Pozzobon, Martina Piccoli, Paolo De Coppi Role of the EU Framework in Regulation of Stem Cell-Based Products, by Giovanni Migliaccio, Cristina Pintus

Download or read book Mesenchymal Stem Cell Therapy written by Lucas G. Chase and published by Springer Science & Business Media. This book was released on 2012-12-12 with total page 458 pages. Available in PDF, EPUB and Kindle. Book excerpt: Over the past decade, significant efforts have been made to develop stem cell-based therapies for difficult to treat diseases. Multipotent mesenchymal stromal cells, also referred to as mesenchymal stem cells (MSCs), appear to hold great promise in regards to a regenerative cell-based therapy for the treatment of these diseases. Currently, more than 200 clinical trials are underway worldwide exploring the use of MSCs for the treatment of a wide range of disorders including bone, cartilage and tendon damage, myocardial infarction, graft-versus-host disease, Crohn’s disease, diabetes, multiple sclerosis, critical limb ischemia and many others. MSCs were first identified by Friendenstein and colleagues as an adherent stromal cell population within the bone marrow with the ability to form clonogenic colonies in vitro. In regards to the basic biology associated with MSCs, there has been tremendous progress towards understanding this cell population’s phenotype and function from a range of tissue sources. Despite enormous progress and an overall increased understanding of MSCs at the molecular and cellular level, several critical questions remain to be answered in regards to the use of these cells in therapeutic applications. Clinically, both autologous and allogenic approaches for the transplantation of MSCs are being explored. Several of the processing steps needed for the clinical application of MSCs, including isolation from various tissues, scalable in vitro expansion, cell banking, dose preparation, quality control parameters, delivery methods and numerous others are being extensively studied. Despite a significant number of ongoing clinical trials, none of the current therapeutic approaches have, at this point, become a standard of care treatment. Although exceptionally promising, the clinical translation of MSC-based therapies is still a work in progress. The extensive number of ongoing clinical trials is expected to provide a clearer path forward for the realization and implementation of MSCs in regenerative medicine. Towards this end, reviews of current clinical trial results and discussions of relevant topics association with the clinical application of MSCs are compiled in this book from some of the leading researchers in this exciting and rapidly advancing field. Although not absolutely all-inclusive, we hope the chapters within this book can promote and enable a better understanding of the translation of MSCs from bench-to-bedside and inspire researchers to further explore this promising and quickly evolving field.

Download or read book Bioreactors for Tissue Engineering written by Julian Chaudhuri and published by Springer Science & Business Media. This book was released on 2005-10-18 with total page 394 pages. Available in PDF, EPUB and Kindle. Book excerpt: Developments in tissue engineering for human medicine are increasing rapidly. Advances in stem cell biology, biomaterials science and scaffold design underpin this emerging science. An equally important facet of this field is the rational design and operation of bioreactors to control the nascent tissue growth. For the first time in a single volume, the design, characterisation and operation of the bioreactor system in which the tissue is grown is detailed. Bioreactors for Tissue Engineering presents an overall picture of the current state of knowledge in the engineering of bioreactors for several tissue types (bone, cartilage, vascular), addresses the issue of mechanical conditioning of the tissue, and describes the use of techniques such as MRI for monitoring tissue growth. This unique volume is dedicated to the fundamentals and application of bioreactor technology to tissue engineering products. Not only will it appeal to graduate students and experienced researchers in tissue engineering and regenerative medicine, but also to tissue engineers and culture technologists, academic and industrial chemical engineers, biochemical engineers and cell biologists who wish to understand the criteria used to design and develop novel systems for tissue growth in vitro.

Download or read book Bioreactors written by Carl-Fredrik Mandenius and published by John Wiley & Sons. This book was released on 2016-02-16 with total page 520 pages. Available in PDF, EPUB and Kindle. Book excerpt: In this expert handbook both the topics and contributors are selected so as to provide an authoritative view of possible applications for this new technology. The result is an up-to-date survey of current challenges and opportunities in the design and operation of bioreactors for high-value products in the biomedical and chemical industries. Combining theory and practice, the authors explain such leading-edge technologies as single-use bioreactors, bioreactor simulators, and soft sensor monitoring, and discuss novel applications, such as stem cell production, process development, and multi-product reactors, using case studies from academia as well as from industry. A final section addresses the latest trends, including culture media design and systems biotechnology, which are expected to have an increasing impact on bioreactor design. With its focus on cutting-edge technologies and discussions of future developments, this handbook will remain an invaluable reference for many years to come.

Download or read book Stem Cell Proliferation and Differentiation written by and published by Academic Press. This book was released on 2020-03-24 with total page 258 pages. Available in PDF, EPUB and Kindle. Book excerpt: Stem Cell Proliferation and Differentiation, Volume 138, the latest release in the Current Topics in Developmental Biology series, highlights new advances in the field, with this new volume presenting interesting chapters. Each chapter is written by an international board of authors. Provides the authority and expertise of leading contributors from an international board of authors Presents the latest release in the Current Topics in Developmental Biology series Includes the latest information on stem cell proliferation and differentiation

Download or read book Human Embryonic Stem Cells written by Arlene Chiu and published by Springer Science & Business Media. This book was released on 2003-08-01 with total page 455 pages. Available in PDF, EPUB and Kindle. Book excerpt: A discussion of all the key issues in the use of human pluripotent stem cells for treating degenerative diseases or for replacing tissues lost from trauma. On the practical side, the topics range from the problems of deriving human embryonic stem cells and driving their differentiation along specific lineages, regulating their development into mature cells, and bringing stem cell therapy to clinical trials. Regulatory issues are addressed in discussions of the ethical debate surrounding the derivation of human embryonic stem cells and the current policies governing their use in the United States and abroad, including the rules and conditions regulating federal funding and questions of intellectual property.

Download or read book Cells and Biomaterials in Regenerative Medicine written by and published by . This book was released on 2014 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Engineering and Application of Pluripotent Stem Cells written by Ulrich Martin and published by Springer. This book was released on 2018-02-28 with total page 228 pages. Available in PDF, EPUB and Kindle. Book excerpt: ​This book reviews the latest biotechnological advances with pluripotent stem cells, exploring their application in tissue engineering and medicinal chemistry. Chapters from expert contributors cover topics such as the production of transgene-free induced pluripotent stem cells (iPSCs), expansion, controlled differentiation and programming of pluripotent stem cells, and their genetic instability. Particular attention is given to the application of the pluripotent stem cells for vascularision of engineered tissue and for drug screening. This book will appeal to researchers working in regenerative medicine and drug discovery, and to bioengineers and professionals interested in stem cell research.