A micropipette has two stops on its plunger. The first measures the liquid and the second pushes out the last drop. Keep the two straight and the rest of the technique follows.
Hold the pipette upright with the tip just below the surface.
Check what your pipette really delivers
A pipette can drift out of calibration with use, and nothing on it shows when it has. A calibration certificate records how the pipette tested before it shipped. At home, a milligram scale and some water tell you whether it still delivers what the dial says.
You need a scale that reads to 0.001 g, distilled water at room temperature and a small container to weigh into. Set the pipette to its maximum, 1000 µL on a 100 to 1000 µL model.
Put the container on the scale and tare it.
Pipette 1000 µL of water into it and write down the reading.
Tare again and repeat until you have ten readings, then average them.
Pick the right pipette and set the volume
Use the pipette whose maximum is closest to the volume you need. A 100 to 1000 µL pipette can't be set below 100 µL, and even at 100 µL Eppendorf rates it at ±3.0 µL against ±0.8 µL for its 10 to 100 µL model.
Turning the volume down, stop exactly on the number. Turning it up, go about a third of a turn past the number, then come back down to it.
Read the display with the pipette held level with your eye, so the number lines up with the marker.
Seat the tip
Push the pipette straight down into a tip in its rack with a slight twist. It should sit snug and straight.
Pounding the pipette into the tips can damage it, and a seal that needs force usually means the wrong tip.
The first stop measures the volume you set; the second blows out the last drop. Reverse pipetting draws up at the second stop and dispenses only to the first. Open the printable diagram
Draw up and dispense
This is forward pipetting, the standard method for water-based liquids.
Wet a fresh tip first. Draw the volume up and push it out two or three times before the transfer that counts. The dry air in a new tip takes up sample as vapor, so the first delivery can run light.
Press to the first stop with the pipette held upright, before the tip touches the liquid.
Dip the tip just under the surface and let the plunger rise slowly. Wait a second for the liquid to finish flowing in.
Dispense against the wall of the receiving tube, with the tip at an angle, pressing smoothly to the first stop.
Wait a second, then press to the second stop. That last push blows out the drop left in the tip.
Slide the tip up the wall as you pull it out, keeping the plunger down until the tip is clear.
Reverse pipetting for thick liquids
Glycerol, detergent and other slightly viscous liquids cling to the tip. Press to the second stop before you draw up, so you take in a little extra.
Dispense to the first stop only. The extra stays in the tip and makes up for what clings to it.
When it drips or draws up short
With water, check the tip first. A tip made for a different pipette may not seal; the tip compatibility guide lists what fits. Then run the scale check above. If it still reads low with a tip that fits, the manual for your model says whether you can service the seals yourself or it needs to go back.
Ethanol and other volatile liquids are different. Eppendorf explains that they evaporate into the air cushion inside the pipette, which expands and pushes liquid out of the tip. Its fix is pre-wetting, at least three times for 70% ethanol and five to eight for stronger mixes. For 100% ethanol or chloroform, Eppendorf says pre-wetting is not enough and a positive displacement pipette is the tool.
Care between uses
Store the pipette upright on a stand. When you finish, set it to its maximum volume, as many lab guides suggest, unless your manual says otherwise.
Check it for liquid inside the nose cone before each session. Liquid in the body means something was drawn up too fast.
Repeat the scale check above now and then, and again if the pipette is dropped.
Pipetting equipment on Amazon
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Micropipette
ONiLAB adjustable micropipette, 100-1000 µL
Single-channel adjustable pipette for 100 to 1000 µL, supplied with a calibration certificate. The tip cone comes off for autoclaving.
The standard method for water-based liquids: press to the first stop, draw up slowly, then dispense to the first stop and on to the second to push out the last drop.
For viscous or foaming liquids. Press to the second stop before you draw up, then dispense to the first stop only.
Weigh ten deliveries of water on a balance and average them. At 20 °C, 1000 µL of water reads about 0.997 g.
