hi everyone the micropipetting exercise that is a section in your introductory chapter i'll show you here how it is done it's very very important that you all understand how to operate a micropipette it's such an important tool in labs that you will not get through biochemistry or any biological labs without knowing the use of a micropipette so there are couple different kinds that we use in this lab first is a p200 why is it known as p 200 p for pipette and 200 is because its limit is 200 microliters that's the max volume that it can take and if you see the tip is yellow the top of the micropipette is yellow so it's very easy to know that a yellow tip goes on this micropipe the other kind is a p thousand why thousand now it's the highest volume that it can take is 1000 microliters you can adjust the volume to whatever you need but the maximum volume that you can take with this micro uh pipet 1000 microliters which is equal to 1 ml and if you see it's a blue micropipet so a blue tip goes on it it is very important that we put a tip on the micropipette and fit it snugly before using the micropipe other kinds that we use in this experiment are multi-channel we call the multi-channel because if you see it's not a single channel there are eight channels to this one and 12 channels on this one so these are used not that frequently but during the enzyme kinetics lab we will be using it but more commonly we will be using a p200 so first and foremost you know how to you should know how to hold a micropipette it is very ergonomically designed wrap your fingers around the micropipette like this so that you can operate the piston with a thumb now the experiment will tell you what volume of the particular liquid we need to aspirate so we adjust that volume from this dial if you see it reads 50 right now and if i start going counterclockwise from the top the numbers increase so now i am at 83 84 so i can go up up till 200 but let me go 200 this is 100 microliters we can even you see that line underneath is 100.2 hundred point four so it is so precise that you can go up to the tenth decimal place in a microliter as well so that's how we fix the volume first the other thing to keep in mind are the two stops for the micropipette first level there is a low resistance and it stops there then you put some more pressure and it goes down even more so what are these two stops for i will explain it to you in a bit similarly for the p thousand if you see here it is at the max and if i start rotating the dial the numbers go down so p and the thousands don't usually have a decimal place so whatever it reads that is the volume you are taking if you need to take 800 microliters here is 800 microliters again there are two levels of resistance first is very soft next one is bit harder so if you um when you come to the labs remember this that there are two levels of resistance for a micropipette for a multi-channel it is similar all the channels if i fix a volume it is 50 here if i change the volume all the channels say i come 200 all the channels in this multi-channel will now pick up 100 microliters so on and so forth now i would like to tell you what the two levels of resistance do by picking up some liquid and showing it to you what the two levels do so this is some just some blue water like regular water with some food dye in it nothing alien about it so before picking up the liquid we depress the piston to the first level dip the tip in the liquid and very gently release the thumb if you see my thumb and the liquid fills the tip this is now whatever the reading is here i change it to thousand microliters so there is thousand micro turs about in this micropipette to to dispense this liquid we put the tip next to the um dispensing the container in which you want to dispense the liquid we release to the first level sometimes what happens all of the liquid doesn't come out for that we have to go to the second level this press it again to get rid of all the liquid so that is what the second level of depress the second level of resistance is in each of the micropipettes is to dispense the liquid not pick it up or aspirate the liquid so certain things to keep in mind always use the tip with the micropipette know how to hold it know what the two levels of resistance do and how to adjust the volume on a micropipette so if you do these four things correctly you will know how to use a micropipette and always hold it like this never hold the if especially when the pipette is tip is full with liquid never hold it upside down because there are chances that liquid will go into the barrel and can ruin the machine the mechanism of the pipette we do not touch the tip with our hands because we could be handling corrosive chemicals or microbes sometimes so to dispense the to get rid of the tip we depress the strip dispenser and put it in a beaker so that it's ready for use again now let me show you how to use a multi-channel pipette the basic rule is the same first we add the liquid to a trough so that we can easily aspirate the liquid this is a maximum capacity is 200 but our volume set at this time is 100 microliters so i gently press the piston dip it in the liquid and very gently release the thumb if i do it very quickly there are chances i'll get air bubbles in the tips we don't want that so to release the liquid we do it in a 96 well plate we will be using these 96 wall plates in our experiments so you see there are eight channels here the 96 well plate has eight wells on the x-axis so we gently press the piston and all the liquid comes out and it is good now if we have a p 12 like a 12 channel multi channel because the y axis of the [Music] 96 well plate has 12 wells in it we take if we the experiment demands we take a p uh 12 with a multi-channel with 12 channels same thing press it down let it gently go all the tips have equal amount of liquid in it position it carefully on top of all the wells and again gently press the piston all the way down so that all the liquid is dispensed in the wax so this makes the experiment very fast and accurate now similarly uh as we did not want to touch the tip of a single channel micro pipet we don't want to touch these as well and this again has a tip dispenser so you press it once and all of them come out similarly for a 12 channel you press this once and all of them come out see fun yay hope you are comfortable watch it as many times as you want and try to get the feel of it so that when you come to the labs in fall 21 you're all set for using a microphone thank you