Tech Snapshot® captures today’s cutting-edge tools and technologies that will help drive drug discovery tomorrow. This installment was written by IMCS and Kemp Proteins.

Four Reasons Why Automated Dispersive Solid Phase Extraction (dSPE) in Pipette Tips is the Future


What is dSPE, and what are dSPE tips? dSPE stands for dispersive solid-phase extraction. From its name, the technique relies on the dispersion of a solid-phase (i.e., functionalized chromatography resin) to facilitate batch extraction. dSPE pipette tips such as IMCStips® contain loosely packed resin stored between upper and lower porous filters. A blue ring, called a disperser, is placed within the disposable pipette tip to promote turbulent mixing. The resins within the pipette tip, each with their unique properties, influence the application’s scope and parameters. For instance, by using affinity resins, recombinant proteins can be purified inside a dSPE tip using the conventional bind, wash and elute workflow reminiscent of traditional chromatography columns. With dSPE, the difference lies in the fact that repeated aspiration and dispense in the tips (binding cycle) facilitate a bi-directional flow that maximizes analyte binding to the resin. A wide range of applications has benefitted from dSPE tips: from isolating small molecules and cleaning up samples to enriching phosphopeptides in cell lysates. These publications have also demonstrated that dSPE tips are faster alternatives to spin column chromatography, magnetic bead-based affinity purification or vacuum manifold purification. IMCStips were most recently shown to enable affinity purification using commercially available resin and a pipetting robot, effectively facilitating the automation and scalability of high throughput purifications.    

As the demand for high throughput applications continues to rise, there is a great need for a micro-purification strategy that accommodates fully automated workflows. Such an approach requires compatibility with standard robotic liquid handlers, low sample carryover, reliability, high protein yields and less labor time—features fully met by dSPE in a pipette tip.

The following are the reasons why these tips are the future:

Reason #1: Loose Resin in dSPE Pipette Tips Facilitate Higher Analyte Recoveries

As mentioned earlier, dSPE tips contain loose resin. The repeated aspiration and dispense in sample wells enable turbulent mixing that is further enhanced by the disperser. Resin dispersion maximizes the surface area, making this technique more effective than traditional unilateral flow. Extractions with dSPE tips lead to higher analyte recoveries in less time than conventional methods such as spin column chromatography. The figure above shows results from an experiment that demonstrates increased protein recovery from IMCStips compared to the traditional spin column method. Spin columns and IMCStips used in this experiment contained equal amounts of the same affinity resin. The identical samples and sample volumes were processed three (3x) or five (5x) times to mimic the 3x or 5x aspiration and dispense (or binding cycles) of automation with a pipette tip. The ability to control pipetting speed and to reset the resin bed by dispersion during pipetting steps negate any channel effects that may limit analyte interactions to the resin in traditional chromatography products.

Reason #2: Well-characterized Binding Kinetics Enable Tailored Workflows

Since dSPE tips are miniaturized chromatography columns, methods and models to characterize binding kinetics are within reach. Binding profiles for IMCStips are available, enabling users to select resin bed amounts and binding cycles based on available binding data. For instance, the figure above shows that the binding rate of polyclonal human antibody (huIgG) to affinity IMCStips is dependent on protein concentration and ligand densities distributed throughout the porous resin. Based on the curve above, reducing the binding cycles for lower titers (< 0.5 mg/mL) with smaller resin beds will provide faster processing. Pushing for greater yields at higher titers (> 0.7 mg/mL) will require more binding cycles equivalent to longer residence times. This automated approach cuts down the purification time of 96 samples to somewhere between 10 to 30 minutes.

Reason #3: dSPE tips Demonstrate Consistency and Reproducibility when Paired with Automation

Ideal execution is leveraging the precise liquid aspiration and dispense speeds of automated pipetting systems, and the beauty of this technology lies in its compatibility with automation systems. Allowing the liquid handler to facilitate repeated pipetting throughout the purification workflow enables consistency and reproducibility. For instance, an automated affinity purification workflow with IMCStips has been widely used by Kemp Proteins for their high throughput micro-purification services, giving clients access to highly purified micro-quantities of their protein. This workflow provides consistent results with high accuracy, precision and repeatability for candidate protein screening. To learn about how Kemp Proteins implemented IMCStips and automation to their processes, watch the webinar, Levaraging Automated High-throuhput Clonal Selection: How MicroScale Purifications Prevent Bottlenecks and Increase Success.

Reason #4: Tip-based Sample Preparation Has a Wide Variety of Applications

The applications with dSPE tips such as IMCStips are endless. Historically, dSPE tips have mostly been used for small molecule isolation and sample clean-up followed by analyte detection. Other examples include contaminant detection in pork and wine and catecholamine extraction in urine. With the foray of IMCStips in the biomacromolecule purification market, expanded applications include affinity purification, peptide desalting with reverse-phase resins, ion-exchange chromatography, phosphopeptide enrichment and even automated multi-attribute method (MAM)

To learn more about IMCStips, please visit Learn how Kemp Proteins uses IMCStips by watching this webinar.