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AFFINITY
OVERVIEW
Affinity applications are used for IgG isolation, recombinant protein purification, anti-drug antibody assay, therapeutic drug monitoring, and more. Because we understand that the needs of a laboratory are often individualized and specific, IMCStips® are available in a variety of resins designed to optimize multiple testing processes. The combination of our patented dispersive technology and customized resin options facilitate consistent, high recoveries of significant analytes.
BENEFITS
- Rapidly yields small but valuable quantities of pure recombinant protein in a single chromatography step
- Consistent, high recoveries resulting in less than 10 percent variance across 96 samples
- Process 96 samples in minutes
- Templated workflows that combine complex methods
- Delivered with Hamilton Microlab STAR™ pre-programmed software scripts with an intuitive user interface to control sample volumes, aspirate/dispense cycles, and wash steps
HIGH PERFORMANCE AND CONSISTENCY
1 mL IMCStips with 50 µL of Protein A resin was used at increasing aspirate/dispense cycles to improve the final yield of IgG as indicated by protein quantification (UV-Vis Abs @ 280 nm) and also by the depletion of the heavy and light chains in the flow through media. The eluants are after 20x aspirate/dispense cycles followed by a final elution volume of 400 mL in NaOAc pH 3.5. The sample was neutralized with 1/10 volume of 1 M Tris pH 8. Each cycle is a duplicate run with the exception of the final 20x flow through and eluants which are in triplicates.
1 mL IMCStips with 50 µL of Protein A resin was used for the enrichment of mAb from cell media. It was tested over three days using a different number of tips and two different aspirate/dispense cycles.
Affinity IMCStips for Tecan
Affinity purification with IMCStips® utilizes patented dispersive solid-phase extraction (dSPE) technology. The loose resin is dispersed during repeated aspirate and dispense cycles, resulting in maximum analyte binding to the resin in a shorter amount of time. IMCStips are designed for seamless compatibility with the Tecan Fluent® automation workstation, enabling you to rapidly process up to 96 samples in under one hour. The combination of our patented dispersive technology, unrivaled scientific support, and customized resin options facilitates consistent, high recoveries of significant analytes. We provide the scripts so you can Just Click Go!
AUTOMATED AFFINITY PURIFICATION WITH IMCSTIPS ON A TECAN FLUENT
Sample deck layout for automated purification of 96 samples using IMCStips on a Tecan Fluent.
Protein Recoveries Following Automated Affinity Purification Using IMCStips on a Tecan Fluent
Cell culture media was processed with Affinity IMCStips on a Tecan Fluent. The starting sample volume was 850 μL at a concentration of 337 μg/mL (or 286 μg of starting protein). Measurements were performed using Octet. 150 μL of samples were eluted with 100 mM glycine, pH 3.0, and neutralized with 15 μL of 1 M Tris HCl, pH 8.5.
Affinity IMCStips for Dynamic Devices
Increased binding across a wide range of concentrations
The results above demonstrate the automated affinity purification of polyhistidine-tagged GFP using 1 mL IMCStips on the Dynamic Devices Lynx LM1200. The IMCStips were packed with three different resin beds (25 μL, 50 μL, and 100 μL) to purify a range of polyhistidine-tagged GFP concentrations. The graphs below indicate (A-C) the percent GFP bound and (D-F) the recovery profile over 30 binding cycles for 25 μL, 50 μL, and 100 μL resin beds.
The results above demonstrate automated affinity purification of polyhistidine-tagged GFP using 5 mL IMCStips on the Dynamic Devices Lynx LM1200. (A) The binding profile of various loading concentrations of GFP over 50 mixing cycles. Higher protein amounts require an increase in binding cycles. (B) Elution volumes were tested on tips bound with 15 mg of GFP. Recovery percent plateaus at 2 mL and most of the bound fraction is recovered with 2 elutions. (C, D) Recovery of GFP from bacterial cell lysate at different GFP load amounts, using one or two elutions. Each elution used 2 mL.