Vision Research Protocols. P. ElizabethRakoczy, ed. Totowa, NJ: Humana Press, 2001. Pages: 340. Price: $99.50. ISBN 0‐89603‐837‐8.

Vision Research Protocols. P. Elizabeth Rakoczy, ed. Totowa, NJ: Humana Press, 2001. Pages: 340. Price: $99.50. ISBN 0-89603-837-8. Who's afraid of molecular biology? Unfortunately, many ocular aficionados, both in clinical practice and in basic research, are. In the words of the editor, P. Elizabeth Rakoczy, Vision Research Protocols “was written for those scientists, optometrists and ophthalmologists who are interested in eye research, but that have not been trained in molecular biology.” With these words in mind, the goal of the book is to bring current techniques in molecular biology to clinicians and vision research practitioners with a constant emphasis on molecular techniques as they specifically relate to eye research. This goal is important, laudable, and ambitious. The book is organized into 14 chapters. Chapters 1 through 4 provide a foundation for the subsequent chapters in the book. These chapters introduce basic molecular biology techniques, transfection of deoxyribonucleic acid (DNA) into cultured cells, in situ hybridization, and immunocytochemistry for the localization of specific proteins by both light and electron microscopy. Chapter 5 discusses methods to inhibit gene expression both in cultured cells and in vivo using antisense oligodeoxynucleodies. Chapter 6 introduces the exciting concept of using catalytic ribonucleic acid (RNA) molecules (ribozymes) to specifically destroy mRNA transcripts expressed from mutant genes responsible for dominant genetic eye disorders, while leaving the messenger ribonucleic acid (mRNA) expressed from the nonmutant allele intact. Chapters 7 through 10 are concerned with viral-mediated gene therapy approaches, and these chapters specifically present methods to prepare and deliver recombinant adenoviruses, adeno-associated viruses, and retroviruses to ocular tissues. Protocols concerning transgenic and knockout mouse models to study ocular development and disease are discussed in Chapters 11 and 12. A short introduction to mutation analysis in human ocular genes is presented in Chapter 13, and Chapter 14 is devoted to an approximately 50-page review of inherited diseases affecting the retina “from the perspective of the clinician.” Although I have 15 years of experience in applying molecular techniques to study eye development, I attempted to evaluate Vision Research Protocols from the perspective of a molecular biology novice. It is my firm belief that molecular biology is the glue that will eventually bind eye researchers and clinicians alike to a better understanding of normal and abnormal ocular physiology and ultimately lead to more specific and effective diagnostic and therapeutic approaches for patients with ocular disease. In the preface, the editor states “in spite of the constant media attention, most people remain confused about the importance and application of these new discoveries. It seems that only the high priests of this new technology, armed with PhDs in molecular biology, understand the meaning, importance, and consequences of these new breakthroughs.” The editor's goal is apparently to bring an understanding of these technologies to all those who may benefit from this knowledge either in their own vision research programs or in their ability to explain disease processes or treatments to patients in a clinical setting. Before reading this book, one must consider what can reasonably be accomplished in a text of only 320 pages. One can not expect the kind of breadth or detail presented in manuals such as John Wiley & Sons Current Protocols in Molecular Biology, but that certainly is not the editor's intention. Also, the sheer weight of the enormous four volumes of the Current Protocols tome is enough to intimidate the molecular biology novice back to the days of leeches and bloodletting. What the book does accomplish is a brief introduction to how basic molecular biology, molecular genetics, and gene therapeutic approaches can be applied to ocular disease research. The length of the text demands a rather superficial coverage of each topic presented. For example, the first chapter, entitled Basic Molecular Biology Techniques, provides protocols for the isolation of nucleic acids including genomic and plasmid DNA, as well as total RNA and mRNA; Southern, Northern, and Western blots; polymerase chain reaction and reverse transcriptase PCR and subcloning in 30 pages including references. Although this opening chapter effectively introduces these techniques, omissions (such as the isolation of DNA from agarose gels, the isolation of proteins, or solvents used to dissolve antibiotics) that could easily have been remedied by the inclusion of references to molecular biology manuals, would have made the chapter stronger. Readers interested in localizing mRNA or protein expression in vertebrate tissues should find Chapters 3 and 4 particularly helpful. I found the chapters (Chapters 5 and 6) on antisense DNA technology and the inhibition of gene expression by ribozymes particularly well written and interesting, and I am an admitted novice in these areas. The pros and cons of particular viral vectors in mediating gene therapy protocols are effectively presented and discussed in Chapters 8 through 10. Chapter 11, on transgenic mice, rightfully points out the fact that many inbred mouse strains have inherent genetic abnormalities that can exclude them as good models of ocular disease. This fact is often not appreciated until after months of effort have evaporated. The concentration of DNA injected into mouse embryos, however, was mistakenly reported as 200 to 400 copies of DNA per microliter rather than 1 to 2 ng of DNA per microliter. The volume of DNA injected into a mouse pronucleus is on the order of 1 to 2 picoliters, and the number of molecules injected depends entirely on the size of the transgenic construct. A similar factual error was found in Note 11 on the knockout animal model chapter (Chapter 12) where the author states, “It is more convenient to use male chimeras to establish germline transmission. Females can be used, but the process is necessarily slower.” Although it is correct that it is usually preferable to use male chimeras to achieve germline transmission of embryonic stem cell clones, this has little to do with the process being slower in females. Most embryonic stem cell lines are male in origin and cannot go through the female germline without losing the Y chromosome through a nondisjunction event. In contrast, the few female embryonic stem cell lines can likewise not go through the germline of male chimeras. Despite this error, the chapter does do a good job of introducing technical aspects of homologous recombination in embryonic stem cells. The final chapter also does a fine job of bringing the readers back to clinical relevance and what is known about the genetics of several different retinal dystrophies and diseases. This chapter is as useful to educate the focused basic researcher on a wide variety of diseases impacting the retina as other parts of the text are to educate clinicians on basic molecular biology. Vision Research Protocols does an admirable job of introducing a wide range of topics in molecular biology and molecular genetics to novices in these areas. Although some basic knowledge in genetics is helpful to the reader for following the protocols, the techniques presented are solid and basic. The topics are not necessarily cutting edge technology. In particular, I think that it would have been appropriate to discuss conditional gene targeting strategies because these technologies are clearly at the forefront of current transgenic and knockout publications. However, given the size of the text, the editor has put together a heroic and largely successful effort to bring the tools and concepts of molecular biology to those who might otherwise have remained unenlightened.