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«Episode 57, “Two Protons Walk Into an H Bar” Dramatis personae: - Ben Tippett - Brian Cross - Nicole Prent - Jacqueline Townsend Copyright Ben ...»

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Episode 57, “Two Protons Walk Into an H Bar”

Dramatis personae:

- Ben Tippett

- Brian Cross

- Nicole Prent

- Jacqueline Townsend

Copyright Ben Tippett

The Titanium Physicists Podcast

www.titaniumphysics.com

Ben: Never be afraid. There's nothing which is known which can't be understood. There's nothing

which is understood which can't be explained. For over 50 episodes now, my team and I have

brought you to the very frontier of knowledge in physics and astronomy. And still our mission goes on. To present you with your birthright: an understanding of the Universe. I've traveled the world seeking out a certain type of genius. Masters of not only their academic disciplines but also at explaining their research in understandable ways. And I've bestowed upon these women and men the title of Titanium Physicists. You're listening to the Titanium Physicists Podcast and I'm Ben Tippett. And now... allez physique!

01:12 [Intro song; Tell Balgeary, Balgury Is Dead by Ted Leo and the Pharmacists] 01:46 Ben: Only two women have won the Nobel Prize in physics. This shouldn't be taken as an indication that women haven't made outstanding discoveries in physics in XX century but rather of the sexism women in physics have often faced. For example, Madame Wu who was one of the greatest experimentalists of her day and designed and ran an experiment showing that our Universe is not symmetric under parity transformations. You remember, we covered this topic in episode 52 when we were talking about things going to the mirror world. Anyway, in 1957 Madame Wu was denied the Prize in favor of two of her colleagues who worked on the theoretical side of the problem. But like I said, only two women have been awarded this Prize in physics. You know the first one - it's Marie Curie but the second one was also brilliant. Her name was Maria Goeppert-Mayer and she shared the Prize in 1963 with J. Hans D. Jensen and Eugene Wigner for their work on the structure of atomic nuclei. Now, Maria Goeppert-Mayer's PhD thesis was kind of crazy - to my mind, at least - and crazy good. We've talked a lot on our show about quantum mechanics and atomic transitions. It takes a very specific wavelength photon energy to get an electron in one orbital to jump to a higher orbital. And upon dropping back down to the lower orbital, the electron will release a photon of the exact same wavelength. That atoms only interact with very specific colors of photons is the heart of the field we now call molecular spectroscopy which is recognizing which atoms or molecules are present in a cloud of gas based on which colors are absorbed or emitted. So you take a gas and you shine some light on it and then you look at the spectrum and the missing lines in the spectrum tell you what atoms are in it. Okay, but like I said - the atom will only absorb one specific energy of photon. It'll leave the rest untouched. Now, Maria Goeppert-Mayer's theory said that this wasn't exactly true. She said that if two photons with exactly half the required energy showed up at the same time, they'd get absorbed instead. It's like/ imagine if there was a parking meter that only accepted dimes. Maria Goeppert-Mayer proposed that putting two nickels in at the same time would also sometimes work. Or it's like two kids trying to sneak into a restricted movie by sitting on each other's shoulders and wearing an overcoat. It's a crazy argument but the effect works - these two little kid photons need to be really, really, really close to the atom at the same time to be absorbed.

Like, an attosecond apart. An attosecond, incidentally, is the amount of time that takes for light to travel the length of three hydrogen atoms. It's a really small distance. But, it works. So, today on the Titanium Physicists Podcast, we're gonna be talking about how this effect gets used to see inside things without perturbing them. Using lasers! It's called multiphoton microscopy and I'm so excited to be doing a show on biophysics. Now, the desire to see through a system without perturbing it is a lot like watching a play. The characters on the stage are meant to be as human as possible but somehow they're ignoring hundreds of faces that are staring at them and watching them and judging them. And somehow, in this most contrived of circumstance, the audience is meant to learn about the intimate workings of this human soul. So who better to invite on the show than a Broadway actor? Our guest today got a degree in Economics from Brown University and then moved to New York City, the Big Apple, to become an actor. In the last few years he's appeared in a variety of roles, both on and off Broadway. Welcome to the show, Brian Cross!

Brian: Hey!

Ben: So, Brian is currently featured on an off-Broadway play Desire, opening September 10th, at 59 East 59 Street Theater. Okay, so Brian.

Brian: Yes?

Ben: For you today, I've assembled two brand new Titanium Physicists. I call them my Biophysics Team. Arise, Doctor Nicole Prent!

Nicole: Choo choo! It's me!

Ben: Dr Nicole got her PhD from the illustrious University of Toronto in Biophysics, specializing in non-linear microscopy. She's currently teaching physics at Camosun College in a beautiful Victoria, British Columbia. Now arise, Doctor Jacqueline Townsend!

Jacqueline: Whoo-ooo-hooo!





Ben: Dr Jacqueline got her PhD from the University of Pittsburgh in Biophysics. She's currently faculty, teaching physics at Colorado State University! Alright everybody, let's start talking about light and cells and stuff. So, Brian, are there any questions to start off?

06:17 Brian: What is biophysics?

Jacqueline: Biophysics is basically a really broad field where we take a lot of sophisticated physics tools and principles of physics like thermodynamics and fluorescence and spectroscopy and all of these tools that come out of really high-end physics and turn those back around to look at life, to look at how proteins work, how enzymes catalyze reactions within the cells, how our muscles move.

You know, all of the really physical and chemical processes that go about to enable life to be. You know, to look at those we have to go back and use really some sophisticated tools that have come out of the field of physics.

Brian: Hm. So in short/ Nicole: It's taking fundamental physics and putting it into a biological sample. You know, obviously makes it more complicated.

Brian: So, in a way it's a study of the way in which the physical, non-living phenomena on a very small scale affect and inform living phenomena?

Jacqueline: Yeah, that's a good way to put it.

Ben: So, today's topic involves the broader field of microscopy. Essentially, we wanna look inside stuff. And on the face of it, looking inside stuff should be fairly simple, right? Like, a microscope what is it? We take some light and we shine it under a small sample of bacteria or whatever and the light's filtered through and then our eyes perceive it. And we go "oh, look, gross, I was drinking that".

In some broader sense, though, if you want to go past that, we want to be able to discriminate between different things inside the system, right? I mean, one squiggly bit is going to look like another squiggly bit just in your microscope. So, essentially at this point in the game we want to move past that and we wanna talk about different techniques we've used to move past that to store or filter light in such a way that we can tell the difference between different things inside the cell.

08:20 Jacqueline: So I know a lot of the previous episodes on this show've been/ were about astrophysics and microscopy has followed sort of analogous paths to telescopes in terms of/ you know, when Galileo first invented the telescopes, you know, you could only see things very nearby, you know.

And over last 150 years as the telescopes have gotten better, we can see further and further out into the Universe and into different galaxies and, you know, we have all those spectroscopic tools that let us see what types of elements are on different, you know, star systems and all that sort of thing. And with microscopy we've gone from - again, really, really basic - microscopes that just have lenses, you know, ground glass very similar in nature to the early telescopes, to developing tools that instead of letting us look further and further out now let us look further and further in. We are able to look at not just the cell itself but within cells. We're developing more and more sophisticated techniques that let us selectively look at different molecules within cells. And a lot of how we selectively look at different parts of cells has in the past mostly relied on staining things. And stains are molecules that, well, interact with specific other types of molecules in a very rough sort of way. You know, if you spill a glass of wine on your carpet, you get a stain on your carpet. If you dump a bunch of iodine in an onion cell - and a lot of you've probably done this in high school - you put iodine on an onion cell and then you look at it under a really basic microscope and you can see the outline of the cell. Because the molecule binds to specific parts of the cell and not others. And that early, early methods let us start developing a sense of what different parts of the cells were and there's a lot of limitations to those as stains only let us work at, you know, very very rough classes of molecules. Maybe they only let us look at lipids or only at proteins or only at cellulose. And so it's a very limited technique. It was best we had for a very long time but we're really moving a lot beyond that these days. So the types of stains that we've used historically have a lot of limitations to them. They only bind to very general classes of molecules and normally let us see very rough structures within the cell. And in recent decades we've been coming up with better and better ways of looking at finer and finer resolution images. Both in terms of the types of microscopy that we have available to us and also in terms of the ways that we actually visualize biological molecules. One of the other limitations of stains is that they kill the cells. So you can't look at something that's alive and also stained. You know, and dead cell are obviously not exhibiting a lot of very interesting biological behaviors because when we study biology, we wanna study things that are still alive. And so we're gonna kind of talk through some of the different types of imaging and get back around to how did we solve that problem of looking at very specific structures within living cells and the techniques that enable us to do that.

11:49 Brian: So staining is kind of foreign process to me. So just to get an understanding of exactly what that is/ Um, when you stain a cell, so you take the onion cell example. You look at it under a microscope and you're looking at the stained version of what is a dead onion cell, right?

Jacqueline: Right.

Brian: Are you seeing the cell or are you seeing it the way you'd see the negative of a photo? You're seeing, like, parts of the cell that are highlighted because of the staining but not the cell itself? Does that question make sense?

Jacqueline: Yeah, that's not quite right but that's not a terrible analogy. So when you're adding the iodine to an onion cell, you're highlighting certain structures and not others.

Brian: Okay.

Ben: The stain molecules will bind to some of the different types of molecules in it. They'll bind preferably to, like, a lipid or something. That way they'll, you know, glow red.

12:40 Jacqueline: So, iodine binds to starch molecules really well. But there's a lot more in that cell than just starch.

Brian: So the limit of staining, as a technique of seeing cells, is one - it only works on certain kinds of cells and two - even on the cells it works on, you're only highlighting a limited number of things within that cell.

Jacqueline: Different stains will only work on specific types of cells and on those cells it only highlights certain types of molecules. Plus, it only highlights the very general class of molecules. So, say, if you're using a stain that binds to protein, you know, there's thousands of different types of protein molecules within a cell but it will just show you all of them. So it's not very good at differentiating things - it's very limited, you know. Plus, with the fact that it kills the cells. And a lot of times we wanna look at live cells.

Brian: If only there were a better way.

13:38 Nicole: If only there were a better way. [laughs] Jacqueline: It was a big advantage. Because if you look at that onion cell with no staining it's practically invisible.

Nicole: So, it was definitely a big step over "no stain".

Ben: There's a general principle behind how staining works that's consistent in all the other techniques we'll see today, which is that it makes part of the cell that you're interested in more visible. Discriminates between that part of the cell and anywhere else. And so you can tell where the parts that are starchy are or where the proteins are, right? That alone is helpful information but if you want to understand how, say, cells work in more detail, you need an ability to discriminate between very, say, specific proteins. You wanna know where some very specific proteins are or where those proteins go. And so the common element in all the things we're gonna be talking about today is that we are doing things that make certain parts glow, essentially. Should we talk about fluorescence, a little bit more? Um, what is it? Nicole, what's fluorescence?



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