Lisa Grufman, Senior product manager
Inger Salomonsson, Research engineer
Micro-purification in biotechnology: what it is and why it matters
In biotechnology and life sciences, precision and efficiency are critical. Micro-purification, the use of highly precise purification techniques to isolate biomolecules at micro or nano scales, plays an important role in sample purity and quality. This level of accuracy is essential for downstream analyses and applications. Whether you're working on advanced or basic applications, high-throughput (HT) processes or simpler workflows, with automatic or manual injections, having the right tools can make all the difference.
In this article, we introduce key products and solutions from Cytiva for successful micro-purification, from sample injection to fraction collection. These solutions are designed to deliver high performance, reliability, and flexibility, helping you achieve the highest data quality from your work.
From structural biology to monoclonal antibody analysis with ÄKTA pure™ micro chromatography system
ÄKTA™ pure micro chromatography system is designed to meet the demands of high-performance microscale purification. Its optimized flow path minimizes peak broadening between the UV monitor and the fraction collector, as well as between the injection valve and the column. With minimal delay volumes, the resolution delivers strong results even for low sample volumes (Figure 1).
Figure 1. Comparison of signals detected in the flow path by the UV monitor versus UV detected in the final fraction for: a) ÄKTA pure™ micro (orange) and b) ÄKTA pure™ 25M (blue). Column: Superdex™ 200 Increase 3.2/300 column.
As the leading fast protein liquid chromatography (FPLC) system for microscale purification, ÄKTA pure™ micro offers exceptionally high performance when paired with our protein purification columns and resins. Its capabilities have been demonstrated in structural biology as highlighted in this article.
ÄKTA pure™ micro is a versatile system, ensuring that every peak matters, providing insights needed to achieve a pure monoclonal antibody (mAb) with reduced levels of aggregates, fragments, and charged variants. This article describes how the system was used in a laboratory-scale process to assess critical quality attributes (CQAs) of a mAb using Superdex™ 200 Increase 3.2/300 and Capto™ HiRes 5/50 chromatography columns.
Increasing throughput and reproducibility with autosamplers
Working with many samples at the same time and continuously purifying them often requires multiple manual interventions, which can introduce errors into the process. One of the standout benefits of an autosampler is its ability to handle multiple samples seamlessly.
The autosampler models, ALIAS Bio and ALIAS Bio Cool, can handle up to 2 x 384-well plates, 2 x 96-well plates, or 108 vials for sample volumes up to 5 mL, making them a good choice when increasing throughput in micro-purifications.
Using ALIAS partial injection enables reproducible injections, reduces sample consumption, and provides flexibility in injection volume. The performance of partial injections equals that of manual injections (Figure 2).
Figure 2. Comparison of a) ÄKTA™pure™ micro with ALIAS injection and b) ÄKTA pure™ micro with manual injection. Sample volume: 5 µL; flow rate: 0.05 mL/min; column: Superdex™ 200 Increase 3.2/300 .
Achieving accurate peak collection with fraction collectors
In the work referenced above, the F9R fraction collector has consistently demonstrated its effectiveness. Designed for use with microtiter plates and/or tubes, the F9T fraction collector employs nozzles that produce extremely small droplets, enabling highly accurate peak collection (Figure 3). One important note: avoid collecting very small fractions to minimize the risk of evaporation.
Figure 3. Micro purification using ÄKTA pure™ micro system with Capto™ HiRes S 5/50 column and F9-T fraction collector. The fractions were collected in a 96-well plate, the absorbance measured at 280 nm and the absorbance from the plate was added to the chromatogram as a histogram curve showing minimal effect of the delay between the UV monitor and the fraction collector.
Proven performance in micro-purification workflows
Our solutions have consistently delivered high performance across a variety of applications. For example, Evotec, an R&D biotech company, used our technology to efficiently screen and identify optimal detergent conditions for membrane proteins, increasing process efficiency and productivity. By adding an ALIAS autosampler to their setup, Evotec was able to screen 48 conditions in a single day in the cold room.
Selecting the right tools and solutions is critical for achieving accurate, consistent, and efficient micro-purification workflows, enabling reliable results across diverse applications.
Related resources:
- Webinar with Evotec: Accelerating protein purification with parallelized automated workflows
- Purify app: Design your system today with the Purify app. Configure your next ÄKTA™ chromatography system, select columns and resins.
- Purify club: Join us today for a world of knowledge at your fingertips.