in this video, we're gonna begin our discussion on specific activity. But before we talk about specific activity, were first going to talk about activity in general. And so activity is just a quantitative measure of both the function and the amount of a specific target protein of interest. And so the greater the activity, the more of the target protein we're gonna have and the more function that we're going to get from the target protein. And so when biochemist are interested in purifying a target protein, they normally take the time to develop an essay for the target protein. And so an essay is any test or experiment that will measure both the presence as well as the activity of a target protein. And so this essay can be performed at any point during our protein purification process on just a small portion of the sample. So we don't need to use a lot of our sample, and we do this ass A to check for the activity off the target protein just to make sure that our target protein this even still there after we performed a purification technique. And so it's important to note is that if two different samples have the same exact amount of activity that means that they're going toe, have equal amounts of the target protein and so down below. In our example, we're gonna clarify this idea of activity. And so, in our example, we're going to compare the activity of the red protein and both of the flasks below. So notice we have flash number one on the left and flash number two on the right. And these marble looking things are representing proteins. And so notice that flash number one has a lot more proteins than flashed number two, but specifically for the red protein, which is our target protein of interest. In this case, notice that flashed number one has 12345 of the red proteins and flashed Number two has 12345 of the red proteins as well. So, uh, even though flash number one has more proteins in general, flash number two and flash number one have equal amounts of target protein of the red target protein. And so this question over here asks which flask has greater activity? Is it flashed number one, flash number two, or do both flask have an equal amount of activity. And so recall from our lesson up above that activity eyes a measure of both the function and the amount of a target protein. And if two samples have equal amounts of target protein, then that means that they're gonna have the same activity. And so what that means is that flat number one and flats number two, because they have the same. They have equal amounts of target protein, the red target protein. That means that they're also gonna have equal amount of activity. So here, option C is gonna be the correct choice. We can, uh, market as being the correct choice and cross off all the other options. So what you can see here is that activity will tell us the amount of the target protein. But what activity will not tell us is how pure our target protein is. So notice that the red protein and flash number one is not very pure at all, because there are so many other different types of proteins and the red protein and flash number two, even though it's a lot more pure than the red protein and flash number one, the activity of both of these proteins is still exactly equal. And so how do biochemist determine if their target protein is pure? And that's exactly where specific activity comes into play on. We're going to talk about specific activity and our next video, so I'll see you guys there.

So in our last lesson video, we said that activity is a measure of the amount of a target protein. But the activity alone does not tell a biochemist if their protein is pure. And so here we have a question, and it asks, How can biochem is actually tell if they've successfully purified a protein? Well, it turns out the answer has to do with the specific activity. And that's because protein purity can be measured by a target protein specific activity. And when it comes down to it, all the specific activity is is just a ratio. It's the ratio of the target protein activity over the total amount of all of the protein that's present. And so, looking at our example below notice, we have an equation for the specific activity. And really, all the specific activity is is just the ratio of the target protein activity or the amount of the target protein over the total amount of all of the protein that's present. And so it's important to note that specific activity actually increases during purification. And the answer to our question here, the way that biochemist actually can tell if they've successfully purified of protein is when the specific activity becomes both maximal and constant. And so when the specific activity is at its maximum level and when that specific activity does not increase anymore because it's constant, biochemists know that they have a pure protein. And so let's take a look at our example down below to clear up this idea of specific activity, and we're going to compare the specific activity of the red protein of interest and both of the flash shown below. And so notice the question below is asking which flask has a greater specific activity. Is it flashed Number one Flash number two Or do both flash have equal specific activity. Now we already know from our previous lesson video that both flask one and flashed to have equal amounts of activity. And that's because they both have a total of five of the red target proteins of interest. And so because the activity is the same between both flasks, the only thing that's gonna be different is the total amount of protein, and it's clear to see that flash number one has a lot more total protein than flash number two, and so a bigger number in this denominator is actually gonna make the specific activity smaller. It's gonna make it a fraction and a smaller number. And the denominator is gonna make the specific activity much, much larger. And because we're looking for the flash that has a greater specific activity, it's clear to see that flask, too, has the greater specific activity, so we can indicate that be here is correct and the other ones are incorrect and weaken. Scratch them all. Now again, we know that specific activity increases during purification, so ah, larger specific activity means aim or purified protein. So because flashed to has a greater specific activity, the red protein in flash number two is more pure than the red protein is in flash number one. And that's pretty clear to see here visually. Now, over here on the right, we're gonna change gears a little bit and talk about how specific activity relates to these protein purification techniques that we covered in our previous lesson videos. And in the first column of our chart. Here we have the purification technique or step, and first we have the crude cellular extract that results from protein extraction. Then we have the sample after salting out uh, ion exchange size exclusion and affinity chromatography, and then the sample after a second round of affinity chromatography. And what you'll notice is that with each protein purification step, the volume of the sample is decreasing. The total amount of protein is decreasing and the target protein activity is also decreasing. And so recall that the target protein activity is the amount of the target protein. And that might seem like something that we don't want to happen. If we're trying to purify a target protein, why would we want the amount of the target protein to decrease? Well, it turns out that's something that we don't want to decrease. But it's inevitable with protein purification and we're going toe lose some of our target protein. But it's worth losing some of the target protein to purify the target protein, and we know that the protein is being purified because of the specific activity and noticed that the specific activity is actually increasing with each purification step. And again, a greater specific activity means aim or purified protein and notice towards the bottom that the specific activity is maximal and it's also constant, since it doesn't increase even after a second round of affinity chromatography. And so that means that we have a pure target protein at that point. Now, notice that the target protein activity is really the top number here of our activity in our equation and the total amount of protein here is really the bottom number here of our equation. And of course, the specific activity is going to be the specific activity over here. So, essentially, if we take this yellow column right here and we divide that number by the total protein column here, we'll get the number, the specific activity number in this column here. And so really, the major take away that I want you guys to get from this video is that the larger the specific activity is the Mawr purified of protein is and so this concludes our lesson on specific activity, and we'll be able to get some practice utilizing these concepts and our next practice video. So I'll see you guys there