Gradient dilution

Many biological or chemical experiments involve substances with a wide concentration range, in which case gradient dilution is a commonly used technique. As with direct dilution, ensuring accurate pipetting volumes is critical. However, it is easy to make mistakes when multiple pipetting steps are involved.

Common reasons why experiments fail

Although in principle, preparing gradient dilutions is relatively simple, many subsequent experiments fail due to errors in three ways:

Inaccurate pipetting——Due to each dilution in gradient dilution being based on the previous dilution, inaccurate pipetting or pipetting effects may be amplified as gradient dilution progresses

wrong hole——Gradient dilution involves multiple pipetting steps, moving in or out of specific wells in a complex pattern, making it more prone to errors than typical experiments

Mix inconsistently——Incomplete mixing at one concentration level can affect all subsequent concentrations, as each concentration is based on the previous one

Save time and avoid mistakes

To avoid the hassle and errors associated with manual pipetting, consider a liquid handling workstation. Protocols can be easily created using the Opentrons Protocol Library - a free collection of protocols that provides downloadable configurations to meet specific volume requirements. Opentrons Flex custom gradient dilution protocols ensure consistent and accurate dilution results, eliminating the repetitive and labor-intensive work of manual pipetting.

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Gradient dilution vs. direct dilution

Both gradient dilution and direct dilution produce a range of concentrations. However, their main difference lies in their methods:

Gradient dilution: Each concentration is produced by diluting the previous concentration. This method can achieve precise dilution over a larger concentration range. For example, if the reserve solution in the first column is diluted 11 times (with a coefficient of 3 each time), then the dilution factor in the last column is 3 (11)=177147 times.
direct dilution：All concentrations are extracted in different amounts from a single stock solution. This method is easier to set up and there is no risk of errors affecting all subsequent concentrations.

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Opentrons provides open source protocols for OT-2 and Opentrons Flex to help you automate serial dilutions.

Learn more

Feel free to contact us and our team of professional application scientists will help you confirm whether the automated experimental process is suitable for your needs. If you need to check the operation of the experimental process, you can schedule an online demo with us to discuss your experimental needs in depth with the expert team.

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