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Sample Preparation

It is important to ensure samples are compatible and within the limits of the analytical technique/assay. Furthermore, while it is ideal to minimize the handling and manipulation of the original sample to reduce error/variability to the analytical workflow; there are many instances when sample preparation is needed. It is particularly important when dealing with complex samples and/or trace-level targeted analytes.

SPE Guide | Avantor

Solid Phase Extraction

Solid phase extraction (SPE) is one of many sample preparation techniques typically employed to reduce the complexity of the sample and/or pre-concentrate targeted molecules of interest. It utilizes a sorbent bed/porous material (solid phase) in a disk, cartridge (may also be described as a column), or a 96-well plate format (higher sample throughput configuration). The analytes of interest are separated/pre-concentrated (extracted) from other sample matrix compounds utilizing fundamental chromatography principles.

Sample clean up – Integration of an SPE method into your workflow can help improve the selectivity and sensitivity of your LC/GC data. Furthermore, it can ensure a longer column lifetime and increase the time frame between scheduled instrument maintenance.

Sample pre-concentration – SPE can ensure your targeted analytes are within the detection and/or quantification limits of the analytical equipment and assay. Once your target analytes are extracted from the sorbent, a ‘blow down’ step increases the concentration of the analyte by evaporating the excess solvent. If required, the solvent environment can be modified to suit your initial LC/GC separation conditions.

Silica-based materials

Most analysts are familiar with using silica-based chromatography columns and can adopt the same chromatography approaches when developing an SPE sample preparation method. Where reversed phase, strong and weak ionic interactions, and mixed-mode interactions (a combination of reversed-phase and ionic interactions) can be employed depending on the chemistry/bonded phase of the silica-based material.

Polymer-based materials

Polymeric resins (when available) are alternatives to silica-based SPE sorbents. They typically have a wider pH compatibility range and different chromatography properties that are commonly employed for ion exchange interactions, although other chemistries are available.

The SPE Process

The main stages of SPE:

  1. Conditioning: To activate the sorbent a highly eluotropic solvent is applied to the sorbent bed, this ensures that the sorbent is fully wetted, and that the analyte will have access to the pore structure. 
  2. Equilibration: Excess conditioning solvent is removed by flushing the sorbent with a low eluting strength solvent, typically it will be the same solvent as is used to load the sample.
  3. Loading: The sample is loaded onto the sorbent. The sample is loaded in a state that will optimize the retention of the sample onto the sorbent, which may require altering the pH of the loading solvent. It is important at this stage that the solubility of the sample in the loading solvent is considered carefully. At this stage some of the matrix components will be flushed through the cartridge as they will not retain on the sorbent. The nature of matrix components that are flushed from the cartridge will depend on the chemical nature of the matrix as well as the chemistry of the sorbent.
  4. Washing: A wash solvent is applied which is not strong enough to elute the compound(s) of interest from the sorbent, however it will remove some of the matrix components. This process can be optimized using a process called elution profile where differing eluotropic strength solvents are applied to determine the strength of the solvent required to elute the compound of interest. At this stage the impact of pH can be very important and may result in the use of two wash solvents at different pHs.
  5. Elution: The analyte of interest is eluted from the sorbent using a suitable solvent. Considerations should be made for the resulting eluent and making sure that there is compatibility with either a blow-down step or the chromatographic procedure that will be applied directly after, as well as ensuring that the recovery is kept close to or ideally at 100%.

This short video demonstrates how SPE can be used in practice.


This guide will help you to choose the right stationary phase SPE product for your analyte of interest.