The Pipette: A Tool for Liquid Precision
How Does a Pipette Work? The Science of Air Displacement
Most modern pipettes operate on the principle of air displacement. Imagine using a medical syringe. When you pull the plunger up, you create a vacuum, and liquid is drawn into the barrel. A pipette works similarly, but with a crucial twist: a cushion of air sits between the plunger and the liquid. This design protects the internal mechanism of the pipette from the chemical being used.
The process has three key steps:
| Step | Action | What Happens Inside |
|---|---|---|
| 1. Aspiration | Press and release the plunger with a disposable tip attached. | Pressing down expels air. Releasing the plunger creates a partial vacuum. Atmospheric pressure pushes the liquid up into the tip. |
| 2. Dispensing | Press the plunger down again to the first or second stop. | The air cushion is compressed, pushing the liquid out of the tip and into the target vessel. |
| 3. Ejection | Press the ejector button or plunger to the final stop. | The used plastic tip is ejected into a waste container to prevent cross-contamination[1]. |
The volume is controlled by a precision piston and a dial that adjusts how far the plunger moves. The relationship between the plunger movement and the volume of air displaced is what makes the measurement accurate. The formula governing this is based on the ideal gas law, simplified for constant temperature: $P_1V_1 = P_2V_2$. Initially, the air inside has a certain pressure ($P_1$) and volume ($V_1$). When you create a vacuum by releasing the plunger, the pressure decreases ($P_2$), causing the volume of the air cushion to increase, which in turn draws in a precise volume of liquid.
Types of Pipettes: Choosing the Right Tool
Just as you wouldn't use a tablespoon to measure a cup of flour, scientists use different pipettes for different tasks. The main categories are defined by their volume range, precision, and operation method.
Volumetric Pipettes are the most precise. They have a single graduation mark and are designed to deliver one specific volume (e.g., $25.00 mL$) at a calibrated temperature. They are often used for preparing standard solutions in analytical chemistry.
Graduated or Serological Pipettes have multiple marks along their length, allowing them to measure variable volumes. You might use one to transfer $3.5 mL$, then $7.2 mL$ from the same solution.
Micropipettes are the adjustable workhorses of modern biology and chemistry labs. They handle small volumes, typically from $0.5 \mu L$ to $10,000 \mu L$ (10 mL). They are further divided into:
| Type (by Volume) | Common Range | Typical Use |
|---|---|---|
| P2 | $0.2 \mu L$ to $2 \mu L$ | Molecular biology, DNA samples |
| P20 | $2 \mu L$ to $20 \mu L$ | Adding enzymes or primers in PCR[2] |
| P200 | $20 \mu L$ to $200 \mu L$ | Protein assays, bacterial cultures |
| P1000 | $200 \mu L$ to $1000 \mu L$ | Buffers, growth media, larger samples |
Precision and Accuracy: What's the Difference?
When talking about pipettes, accuracy and precision have distinct scientific meanings. They are both crucial for trustworthy experiments.
Accuracy is how close a measured volume is to the true, intended volume. If you set a pipette to $100.0 \mu L$, an accurate pipette will deliver exactly $100.0 \mu L$.
Precision (or repeatability) is how close repeated measurements are to each other. If you pipette $100.0 \mu L$ five times and get $99.8 \mu L$, $99.9 \mu L$, $99.8 \mu L$, $100.0 \mu L$, and $99.9 \mu L$, the pipette is very precise, even if it's slightly off from the true value.
A good analogy is throwing darts. High accuracy means your darts are clustered near the bullseye. High precision means your darts are clustered tightly together, even if that cluster is not on the bullseye. The goal in pipetting is to have both high accuracy and high precision. This is ensured through regular calibration, where the pipette's performance is checked against known weights of distilled water (since $1$ gram of water $ = 1$ mL at a specific temperature).
From Recipe to Reaction: Pipetting in Action
Let's follow a concrete example. Imagine you are a food scientist developing a new vitamin-enriched juice. You need to test how much vitamin C remains after pasteurization. Your protocol requires you to prepare a standard solution of known vitamin C concentration to compare against.
The Task: Prepare $50 mL$ of a $0.1$ molar (M) ascorbic acid (vitamin C) solution. You have solid ascorbic acid powder ($C_6H_8O_6$) and distilled water.
Step 1: The Calculation. First, you need to know how much powder to dissolve. The molar mass[3] of ascorbic acid is approximately $176.12 g/mol$. You need $0.1$ moles per liter, but you only want $0.050$ liters ($50 mL$).
$Mass = (0.1 \ mol/L) \times (0.050 \ L) \times (176.12 \ g/mol) = 0.8806 \ g$
So, you need to measure out $0.8806$ grams of ascorbic acid powder using an analytical balance.
Step 2: The Dissolution. You transfer the powder into a clean $100 mL$ volumetric flask[4]. You add some distilled water and swirl to dissolve the powder completely.
Step 3: Precise Dilution. This is where the pipette shines. You cannot just pour water up to the mark; you must be exact. You use a graduated pipette or a micropipette with a large volume tip to carefully add distilled water until the bottom of the liquid's meniscus[5] rests exactly on the $50 mL$ calibration line etched on the flask's neck. This ensures the final volume is precisely $50.00 mL$, giving you an exactly $0.1$ M solution.
Step 4: The Test. Now, to test your juice sample, you might use a P200 micropipette to transfer exactly $100.0 \mu L$ of your juice into a test tube. Then, you use a P20 micropipette to add exactly $10.0 \mu L$ of a chemical reagent that changes color in the presence of vitamin C. The precision of these volumes is critical. If you added $105 \mu L$ of juice instead of $100 \mu L$, your calculated vitamin C concentration would be wrong, leading to incorrect conclusions about the pasteurization process.
Important Questions
Why can't I use the same pipette tip for different liquids?
What do the two "stops" on a micropipette plunger do?
How do I read the volume on a micropipette?
Footnote
[1] Cross-contamination: The unintentional transfer of material (e.g., chemical, biological cells) from one sample to another, which can compromise experimental integrity.
[2] PCR (Polymerase Chain Reaction): A laboratory technique used to make millions of copies of a specific segment of DNA, crucial for genetic testing and research.
[3] Molar Mass: The mass of one mole of a substance (element or compound), usually expressed in grams per mole (g/mol). It is numerically equal to the atomic or molecular weight.
[4] Volumetric Flask: A type of laboratory glassware with a flat bottom, a long neck, and a single calibration mark. It is designed to prepare solutions of an extremely precise volume.
[5] Meniscus: The curved surface of a liquid in a container, caused by surface tension. When measuring volume, you read the value at the bottom of the curve for concave meniscuses (like water).