IMCS Poster Presentation Downloads
Automated High-Throughput Plasmid DNA Purification with 5 mL IMCStips on Dynamic Devices Lynx Platform
Abstract: As the demand for scalable, efficient plasmid DNA (pDNA) purification workflows rises in biotechnology and molecular biology, there is a growing need to move beyond traditional spin-column methods. While reliable, these methods require extensive manual handling, are limited by centrifugation capacity, and involve labor-intensive steps, creating bottlenecks impractical for high-throughput applications.
Here, we present a fully automated, large-volume plasmid purification protocol using 5 mL DDS (MidiPure) IMCStips on the Dynamic Devices Lynx platform, optimized for laboratories requiring streamlined workflows and consistent, high-quality results. This novel protocol integrates all steps of post-lysate clearance, enabling high-throughput, hands-free processing of large bacterial cultures yielding high-purity pDNA suitable for downstream applications.
The MidiPure IMCStips on the DDS are a silica-based purification method for plasmid DNA that can be tailored to fit purification scales ranging from midipreps to maxipreps. In this work, several resin amounts were employed to determine resin binding capacities. For larger-scale applications, the employed protocol was benchmarked against traditional midi spin columns from Company A. IMCStips containing 150 mg of silica resin recovered 117 ± 4 µg of pDNA from 200 mL of bacterial lysate at 109 OD per sample well. Two IMCStips with 150 mg of resin produced yields (233 ± 3 µg) comparable to one traditional midi spin column (251 ± 9 µg), while significantly reducing hands-on time and eliminating the need for labor-intensive centrifugation. Purity and integrity assessments via NanoDrop™ measurements consistently showed 260/280 and 260/230 ratios in the optimal range, with TapeStation analysis and sequencing further confirming the intactness of the plasmid across different plasmid sizes (3-8kbps).
This automated approach on the Lynx platform offers a robust alternative to traditional manual methods, delivering high-throughput, reproducible results suitable for downstream applications such as gene therapy, cloning, and sequencing. The fully automated workflow minimizes human error and processing time, establishing it as an efficient and scalable option for laboratories aiming to enhance productivity without compromising pDNA quality.
Poster # 1216 - A
Automating lower throughput repetitive and tedious protein purifications with a variable span pipetting system
Abstract: Recombinant protein purifications are essential procedures in biotech and biopharma laboratories. For high-throughput production laboratories, dedicated high-end liquid handlers process hundreds of samples per hour. However, for lower throughput operations where sample numbers are decreased, processes are run using manual methods or through a combination of magnetic beads, centrifuge tubes or vacuum manifolds. Such processes involve repeated manual pipetting and intermittent manual transfers of labware from one piece of equipment to the next. Here, we demonstrate pipettes packed with protein A conjugated agarose resins for hands-free purification of monoclonal human immunoglobulins from cell supernatants using an INTEGRA ASSIST PLUS. By incorporating IMCStips on this lower budget, compact automated pipetting system, end users can purify samples from different labware formats such as microfuge tubes and plates. Protein A resin (50 µL) packed in a 1-mL wide bore tip was used to purify monoclonal IgG (titer of 0.5 g/L) from cell supernatant (900 µL). Cell cultures were spun down, stored frozen, and thawed prior to purification with IMCStips. Up to six microfuge tubes were processed in parallel using a 6-channel VOYAGER pipette with 13.5 mm variable spacing on the ASSIST PLUS pipetting robot. Aspiration and dispensation flow rates, repeated pipetting cycles, and pipetting heights were adjusted to address foaming or residual liquids and eliminate any carryover. Two wash steps were implemented using 1x PBS, and both wash solutions were analyzed by absorbance. Protein concentrations in the eluates (150 µL + 10 µL neutralizing buffer) were determined with spectrophotometer at absorbance at 280 nm. Antibody recoveries were over 80% with relative standard deviation of 10% or less. For increased throughput, we used the ASSIST PLUS to transfer up to 48 microcentrifuge tube samples to a 96-well plate which we then processed using INTEGRA's VIAFLO 96, following a similar protocol including a sample binding, two wash steps, and followed by an elution step into a 96-well plate. Key advantages of the pipette-based purification program were the reduction in hands-on time, maximizing resin and sample contact time, and no additional equipment. Preparation time was similar to centrifuge-based method, but the centrifuge and magnetic bead methods required multiple manual interventions whereas the pipette-based workflow was walk-away. .
Poster# 1191 - D
Comparative IMCStip-Based Protein A Purification on the Tecan Fluent: Impact of Resin and Elution pH on Antibody Aggregation
Introduction
Affinity chromatography is a cornerstone technique in protein purification, and the choice of resin can significantly influence the performance of the process. In this study, we compare two commercially available affinity resins - MabSelect PrismA and Praesto™ Jetted A50 HipH - in 1-mL Tecan IMCStips. MabSelect PrismA has become an industry standard for antibody purification, known for its high binding capacity and specificity. Praesto resin is recognized for its ability to elute antibodies at a higher pH, which may offer advantages in optimizing recovery and maintaining product quality. Additionally, this work extends the use of IMCStips the Tecan Fluent automated liquid handling platform, facilitating a fully automated antibody purification and buffer exchange process. Here, using the two Protein A resins, we demonstrate that increasing the pH of the elution buffer results in similar recovery up to pH 3.6 followed by a significant difference in recovery for elution pHs between 3.9 – 4.9. Furthermore, we compare the monomer percentage of purified antibody showing resin-specific and pH-specific effects.
Methods
All automated purifications were performed on a Tecan Fluent system. Monoclonal antibodies (mAbs) were captured from clarified cell culture supernatants using MabSelect PrismA and Praesto™ Jetted A50 HipH resins, both of which were evaluated across a range of elution pH values (from pH 2.5 to pH 4.9). Final purified and buffer exchanged antibody samples were analyzed for recovery and monomer percentage using A280 measurements, size-exclusion chromatography (SEC) and SDS-PAGE.
Preliminary Data
The automated method established on the Tecan Fluent enables the purification and buffer exchange of 96 samples in under 60 minutes, resulting in efficient and reproducible antibody purifications. Our results show that MabSelect PrismA has a higher binding capacity compared to Praesto Jetted A50 HipH. Using two different cell supernatant samples containing mAbs, both resins achieve similar recovery across a range of low pH values, with comparable yields up to pH 3.6. At low pH elution conditions, the monomer percentages of the two antibodies tested were notably influenced by the resin used. At higher pH elution conditions (3.9 and above), the Praesto resin yields significantly higher recovery and monomer percentages varied depending on the resin used.
Novel Aspect
This study highlights the advantages of comparing two different Protein A resins for antibody purification and emphasizes the need to forgo a “One Size Fits All” approach for Protein A purifications. Our data shows that recovery and monomer content are influenced by both the specific resin used for purification and the elution pH. This work paves the way for automating and optimizing antibody purification screening strategies to improve both yield and monomer content.
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