Most importantly, I thank my advisor, W.E. Moerner. It is difficult to explain how wonderful it has been to study under him. W.E. is a real scientist’s scientist: he fundamentally cares about good science and presenting results in a clear and honest manner. He always impressed me with his understanding of sciences outside his field and his scholarship, as I doubt that there is any paper I have read that he has not. W.E. always knows where some obscure piece of equipment is in the lab, and what type of power cable it requires. W.E.’s humor and generosity have been invaluable during my time in his lab, not to mention his scientific guidance. I could not have asked for a better Ph.D. advisor.
I joined the Moerner lab because W.E. seemed to run a fun and exciting research program, and I have not been disappointed. Other members of the Moerner Lab have been instrumental in my education and research. Kallie Willets mentored me when I first arrived at Stanford. Kallie was fun to work with and I am very grateful for the time and energy she dedicated to helping me get a solid footing in the lab by teaching me the right way to do things (and clean up afterwards).
After Kallie graduated, it was entertaining (to say the least) to get to know my officemate Dave Fromm. Dave was always willing to discuss problems I was facing in my experiments, and often suggested perfect solutions. (He was also always willing to discuss his adventures and funny stuff he found on the internet.) Dave and Jim Schuck regularly played darts over my head … literally. In general, this was entertaining and helpful to my overall spirit, and I appreciate the fun times with Jim and Dave. In those early years, I also enjoyed the company of (and scientific input from) Nick Conley, Anika Kinkhabwala, Adam Cohen, Stefanie Nishimura, Jaesuk Hwang, Kit Werley, So Yeon Kim, Andrea Kurtz, Marcelle Koenig, and Jian Cui.
In the later years of my tenure in the Moerner Lab, I have benefited from another batch of amazing people. Nick is one of the most motivating collaborators I have had the pleasure of working with; he is always excited about results, and his mind wanders to great places (not to mention that his skills as an organic chemist were very helpful to me)! I also had the opportunity to work with Hsiao-lu Lee, who was always generous with her time and expertise in cell culture. I am thankful to have those two wonderful coauthors. Alex Fürstenberg has been a fun (and very tolerant) officemate, and is always a great person to ask about anything photophysical. Mike Thompson is hard working and smart, but most importantly he laughs at more than 83% of my jokes. Julie Biteen is opinionated and usually right, and has been fun to bounce ideas off. All the other members of the Moerner Lab (Shigeki, Randy, Majid, Steve, Jianwei, Whitney, Lana, Yan, Sam B, Quan, Matt, etc.) are exceptional people and have made Stanford a wonderful place.
Marissa Lee started joined the lab in 2008, joining my project. I have enjoyed mentoring her and passing on as much as possible of what Kallie, Dave, Jim, Stefanie, Nick, Hsiao-lu, So Yeon, Jaesuk, Adam, and Anika taught me over the years. I wish her luck in her time at Stanford. Several summer students worked with me to get a taste of research. I thank Jennifer Alyono, Daniel Lau, Nathan Hobbs, and John Servanda for their help taking spectra.
Of course, I must also acknowledge Bob Twieg and his students at Kent State University. As a physical chemist, there is nothing better than an excellent collaboration with a group of top-notch synthetic chemists. W.E. and Bob have worked together since their IBM days in the 1980s and 1990s, and I had the fortune to benefit immensely from that bond between labs. Nearly every compound mentioned in this Dissertation was synthesized by the Twieg lab, and the back-and-forth (or push– pull?) design process between the labs should serve as an example to what all collaborations should strive for. Bob’s students have made great compounds over the years, and I thank all of them for being super collaborators: Meng, Hui, Zhikuan, Na, Reichel, Ryan, Alex, and Jarrod.
Friends have made grad school a blast. I met the Moilanens immediately, and enjoyed marathon training and adventures with David and Hailey. Ben Spry was a great help studying for placement exams, and I enjoyed driving to San Jose with Ben so he could buy a Camaro. William Childs and Charles McCrory—after I finally decided to like them—were indispensible: grad school will be filled with fond memories of coffee, lunch, and arguments because of Wm and Charles. So many other friends made my time at Stanford wonderful: Nichole, Kate, Alicia, Jen, Drew, Ashley, John, Zalatan, Chad, Matt, Griffin, Kendall, Daniel, Adrienne, Adam, Avisek, Eric, Ethan, Kevin, Emily, Ken, Dan, Scott, and everyone else! It has been fun having Jordan and Maria in California, and so many other non-Stanford friends that I cannot possibly name them all. I have had positive interactions with several faculty members, and I thank Bob Waymouth, Chris Chidsey, Dick Zare, Bianxiao Cui, Steve Boxer, Justin DuBois, Vijay Pande, Bob Pecora, and Ed Solomon. I also must recognize the members of the Stanford staff who contributed to my work and enjoyment, namely: Roger Kuhn, Todd Eberspacher, Brian Palermo, Patricia Dwyer, Grace Baysinger, Steve Lynch, and all the Conways—Marc, Daragh, and Mariette.
I feel that I must also acknowledge those in my past who influenced me and led me down the path of science. My earliest memories of enjoying the natural world were at Audubon’s Mast Landing Camp, playing and learning about nature with Aaron and Ira and Matt. In the third grade, Mrs. Solari recognized and encouraged my inclination toward science, as have many teachers since. I thank Dr. Root, who mentored me for my 7th-grade science fair project; Mr. Plummer for dealing with 8th graders; Mr. Glick for the astronomy and recycling clubs and supporting me throughout high school; Mr. Herrick, for being the best physics teacher I never had; Mr. Gauger for insisting that Heisenberg’s uncertainty principle can explain why things still jiggle at zero Kelvin; John Anderson for arguing with me; Don Cass for teaching my first college chemistry class and making it so exciting; Tony Planchart for teaching biochemistry in a way that convinced me to be a chemistry major; Helen Hess for fun classes biology and biomechanics; Michael Rubinstein for his entertaining exploration of polymer physics; Royce Murray for teaching analytical chemistry; Max Berkowitz for stat mech classes; and Charles Schroeder, Eric Shaqfeh, and Steve Chu for a great summer research experience. I should offer a special bit of gratitude to Sergei Sheiko, whose lab I worked in as an undergrad, and who helped make my time at UNC spectacular.
This Dissertation is dedicated to my family: the Lords, the Cyrs, and the Hearns. My parents have always encouraged my interests, without pushing me too hard. I wouldn’t be half the person I am without their support. My brother Jackson has been a life-long companion, so I was very pleased when he moved to California and we could play together like when we were growing up. My grandparents Lord funded my education, which I greatly appreciate. I probably get some of my curiosity from my pépère Cyr. My first year at Stanford, I met Brenna Hearn and married her a few years later. She has made my life wonderful, and I thank her for her support throughout grad school. I cannot thank Brenna enough for her companionship, so I’ll stop there.
Looking back at this, I wish that I had made it 50 times longer and cut out the rest of the dissertation.
LED illumination is awesome for epifluorescence. No mechanical shutters, no changing mercury lamps every 200 hours, no hot lamphouses, no worries about letting it cool down before turning the lamp back on, less wasted electricity, immediately ready to use after turning it on, etc.
We have a Lumencor SpectraX on our Nikon TE2000 scope and we love it. It contains multiple LED that are independently triggerable. For high-speed imaging, we bought one new Chroma quad-band dichroic and emission filter set, as well as 4 separate single-band emission filters for our emission filter wheel (although this latter set is not absolutely necessary).
The amazing thing is to be able to run color sequences at the frame rate of the camera (because the SpectraX accepts TTL triggering of each line independently). It is beautiful to see the rainbow of light flashing out of the scope at 20+ frames per second!
We haven’t run into any issues with brightness: the SpectraX is bright enough for all our cell imaging experiments. Typically, we run it at 20% power. That said, I’m aware that the very bright peaks in an arc lamp spectrum (e.g. UV, 435, 546) aren’t there in the LED spectra. So for FRAP or something, you may not be able to bleach as fast.
And, of course, a fancy illuminator like the Spectra X is not cheap. But for run-of-the-mill epi imaging, white-light sources like the Lumencor Sola might be a good option. Another downside is that the fans on the Spectra X are audible, but not annoying. Despite that minor issue and the cost, I highly recommend LED illumination (and the Spectra X, specifically).
I recommend you demo a few LED sources from a few companies (e.g. ScopeLED, Lumencor, Sutter, etc.) and make sure it will fit your needs.
* Make sure your camera supports TTL triggering of an external shutter.