VIDEO OF SPONTANEOUS ACTIVITY OF ACR-2 (ICR) IN MOUSE NEURONS AND ASTROCYTES STIMULATED BY 10mM GLUTAMATE. (Chapter 4, part 3)
Courtesy of Professor Jean-Yves Chatton and Marc Bricquet, Department of Neuroscience, University of Lausanne, Switzerland.
One of the most tantalizing challenges that emerged after the advent of these indicators was the quest to develop red-emitting calcium indicators. Initiated in 1985, these endeavors faced repeated setbacks. This book embarks on a unique exploration, tracing the path of calcium research from its early days to the development of the cutting-edge modern fluorescent calcium indicators that continue to be invaluable in research today.
Intriguingly, the book also sheds light on the fascinating journey of red-emitting calcium indicators. It offers a comprehensive overview of their history, from their inception to the year 2014, as discussed in Martin Oheim’s review article, which emphasized the need for effective red-emitting calcium indicators. Within these pages, readers will find a thorough account of the design, synthesis, properties, and exceptional application of the first authentic CYTOSOLIC (Not in Organelles) red-emitting calcium indicator, denoted as ICR. Notably, ICR is a patent-free creation, which means that scientists are free to utilize it for their unique research and screening requirements.
a) Chapter 1
Most of the information in the first part of chapter 1 on Measurement of Calcium before 1980 were obtained from chapter 9 of Current Trends in Neurobiology, vol 3, John Wiley and Sons(1980) by Roger Tsien and his colleague Tim Rink after the invention of the first modern fluorescent synthetic calcium indicator, Quin-2.
Most of the information on the second part of chapter 1 on Transition from the old methods to the new modern synthetic fluorescent calcium indicators were obtained from personal experience and communication with Roger and his Nobel Prize Biography. The parts from his Nobel prize biography can be seen as quotes.
b) Chapter 2
Chapter 2, which describes the beginning and commercialization of the technology of modern fluorescent calcium indicators, describes my involvement from the development at the beginning with the introduction of the first modern fluorescent calcium indicators and the beginning of their commercialization at Molecular Probes in Eugene, Oregon.
Further expansion of the commercialization from my company, Teflabs, in Austin, Texas and development of modified versions of the original calcium indicators by Molecular Probes, Eugene Oregon, and AATBioquest, in Sunnyvale, California are also described.
To bring the technology to date, new indicators like MaPCa are also described. The chapter describes only useful calcium indicators.
c) Chapter 3
Chapter 3 is actually divided into 3 parts: Please display portions of each part and sub-part in the book after each introduction but not the whole thing.
- Why red fluorescent calcium indicators is further, which is the part first divided into three sub-parts. The first sub-part discusses why I decided to write this book on red-emitting fluorescent calcium. It discloses the information obtained by reading the review article by Oheim et al as shown in the quote and then proceeds to show the experience I got from making ACR-1 and the results I got from Scientists who tested it and the beautiful results they obtained. I set the wavelength parameters of a good red-emitting fluorescent calcium indicator also in this sub-part.
The second sub-part discusses the quest for
red-emitting fluorescent calcium indicators,
and it is obtained from the review article as
well as what prompted me to make ACR-1. The
third sub-part discusses the criteria for a good
red-emitting fluorescent calcium indicator and
was obtained by examining the properties and
applications of all the red-emitting fluorescent
calcium indicators reported by Oheim et al and
of course, my ACR-1. It is these criteria that gives
me the audacity to call ACR-2 (ICR) an ideal
indicator.
- The second part discloses a summary of all the red-emitting fluorescent calcium indicators and their flaws or shortcomings after my examination. They are based on their wavelengths as a supposed red indicator, how successfully they have been utilized as a red indicator and how they have been discontinued due their lack of usefulness.
- The third part discloses red-emitting fluorescent calcium since the article by Oheim et al was written. Only useful red-emitting fluorescent calcium indicators are discussed. GECIs are not red-emitting fluorescent calcium indicators but are discussed because they are becoming useful in imaging and are the specialty of Oheim et al. Halo tags which are red-emitting fluorescent calcium indicators with protein tags for targeting are also discussed because they are the latest group of specialty compounds for imaging.
d) Chapter 4
Chapter 4 is divided into three parts. Portions of each part can be displayed after the discussion of each part.
The first part is design and synthesis of ACR-2 (ICR). This part was obtained from my lab notebook and as such I will be happy to help any chemist who encounters problems with their synthesis. All the have to do is email me at koniyaw@gmail.com and I will be happy to reply with explanations and suggestions. I have tried to be as thorough as possible and hope the chemists do not encounter problems. Portions can be displayed from the book.
The second part displays the physical properties of ACR-2 (ICR) as reported by Dr. Joe Kao in the book. He obtained these results from the potassium salt of ACR-2 (ICR) made from crude hydrolysis of the final step of the synthesis. Some of the numbers like the dynamic range will increase if the salt is purified further. An HPLC purification may double the dynamic range.
The third part is Applications and results from various cells, results from four scientists. These are unedited results from the four scientists delivered by email. They seem to be comprehensive. Dr.Dylan Meyer is at Vertex Pharmaceutical. Dr. Weaver is at Vanderbilt University. Dr. Schubert is at Dresden and Dr. Jean Yves Chatton is at University of Lausanne, Switzerland. Dr. Mario Schubert is at the Technical University, Dresden Germany. The first two are no longer at the addresses connected with their results. Dr Schubert’s results has detailed protocols and will not be addressed. Dr Chatton’s results have protocols in the email, but the results were sent as a link to an AVI movie. Details of the movie could not be shown in a book, so we took still pictures. The website is therefore the best place to see the live version of what is in the book.
e) Chapter 5
Chapter 5 is a condensed biography and therefore should be viewed as such.