How to Use a Micropipette: Proper Pipetting Technique

Get ready to make a new best friend: the micropipette. Why is it your new best friend? Because the vast majority of things you’ll be doing in a lab will involve pipetting at some point or another, so learning how to use a micropipette is crucial and learning to love (or at least respect it enough to do it properly) is crucial. This, then, is a brief guide in proper pipetting techniques. I’ve written out a very thorough guide below and found a pretty decent tutorial video. If you want the quick and dirty, watch the video. For more precise instructions, read on.
 

What is a Micropipette?

how to use a micropipetteMicropipettes measure very small quantities of liquids. In a lab, a micropipette is used to work with small volumes of acids, bases, DNA and RNA solutions, volatile compounds, and so forth. How precise do these micropipettes get? Very. While pipettes come in different sizes (and most labs will have 4-5 micropipettes sizes that can all handle different amounts), the smallest ones will measure 1/10th of a microliter. That is 1/10,000,000 liters.

Given how precise micropipettes can be, you should be aware that there is a lot of room for operator error. In other words, it’s easy for you to mess up a measurement so precise. Read on to learn how to use a micropipette and the mistakes you’ll need to avoid to keep from messing up your experiment.

How to Use a Micropipette

Your goals in learning proper pipetting technique are simple: 1. Measure a liquid precisely, 2. Discharge the measured liquid fully, 3. Don’t ruin your equipment. Everything here addresses one of these goals. I’ve got a brief video that covers much of the basics, but be sure to read below to get all of the details.

Step 1: Choose the Right Micropipette

Do you need to measure 5 μl of a liquid? Awesome. Don’t use a p1000 micropipette. As a general rule, choose the smallest volume pipette that will measure your desired amount. Pipettes come with the following volumes:

  • .5-10 μL (called a p10 micropipette)
  • 2-20 μL (called a p20 micropipette)
  • -200 μL (called a p200 micropipette)
  • 100-1000 μL (called a p1000 micropipette)

So if you need to measure out 100 μL, use the p200 rather than the p1000 because the p200 will be more precise because it deals with smaller amounts.

Step 2: Set the micropipette

The micropipette will have a visual window that has three numbers on it. Here’s how it works: The number on top will be the biggest number that the pipette can measure. For example. The p10 can only measure up to 10 μL, so the top number will be a “tens” place for microliters. You would never want this top number to read 2, because p10 should only be used to measure up to a one in the tens place (and a zero in the ones place), and having a 2 in the tens place would indicate you were in the 20’s- well out of the p10’s range. Another example: the top number on the p1000 will be the thousand’s place. So just like with the p10, you would never want the top number on the p1000 to show a 2. You wouldn’t want the three numbers to show 105, either, as that would indicate 1,050 μl which is still 50 μl beyond what it should measure.

After you’ve got the right micropipette, make sure you put on a disposable tip. There are several different sizes, so make sure yours is correct and fits on appropriately.

Step 3: Draw in your liquid

Your micropipette will have a plunger on top that when you depress, readys the pipette to take in your set amount. When you depress the plunger, you will feel a “first stop” where it feels like you’ve hit the bottom. If you press downward some more, you will pass the first stop and go to the “second stop.” The second stop is useful for expelling the liquid (we’ll get to that later). For now, do the following to draw up your liquid:

  1. Depress the plunger to the first stop
  2. Hold your liquid you are drawing from at about eye level
  3. Put the tip of the micropipette into the liquid, not too far under the surface
    1. Expert Level: for extra precision, try to keep the tip at the same level under the liquid as you draw
  4. Release the plunger (not too quickly) in order to draw up the liquid
    1. Pro tip: If you’re drawing in a thick liquid, make sure you release the plunger slowly and after you’ve released the plunger completely, wait for about 2 seconds. You will likely see the liquid still rising during this time
  5. For each new sample you are drawing up, eject off the old disposable tip and put on a new one
    1. This is to not only avoid cross-contamination (as you may be working with several chemicals/concentrations), but it’s also to help out consistency. A wet pipette tip will work differently than a dry on, so when accuracy matters be consistent. Some people will draw up a sample and eject it several times to wet the disposable tip well before drawing up their actual sample.

DON’T:

  • Don’t lay your micropipette down when it has liquid in it. This will get the liquid beyond the disposable tip and into the pipette itself, which could contaminate all other samples the micropipette will be used for down the road!!!
  • Don’t press down the plunger when your tip is already in the liquid. Bubbles are a source of inaccuracy and you want to avoid them at all costs

Step 4: Expel your sample

Place your micropipette tip right near the edge of the container you will be expeling the sample into and press to the first stop, pause, then press to the second stop. This second press is called a “blow out” and should help expel the last little bit of sample in your tip. This can be crucial to do, especially when you’re working with the smaller tips where capillary action can be strong enough to make it tough to get out that last little bit. The blow out itself, combined with having your tip close to the wall of your new container, should help expel and draw out your sample completely. Also, placing your tip next to the wall will help avoid creating bubbles in your new container.

Step 5: Set your micropipette back to its highest volume

This may be a bit counter-intuitive. After all, if someone comes along behind you and turns the nob the wrong way, they’ll have violated one of the cardinal rules about pipetting- don’t over turn your micropipette. The reason why you want to leave your micropipette at a high volume is because the plunger is spring operated. Storing it in a relaxed state will help with the longevity of your micropipette.

Congratulations, you now know how to use a micropipette!

For a more detailed investigation on what micropipettes are and what other types of pipettes are out there, check out Wikipedia’s article on them.

For a more technical read on micripipette structure and technique, check out UCLA’s guide or a very thorough walkthrough done by Accutek

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