MSACL 2019 EU Abstract

Maximize the Output of Routine Proteome Analyses by Using Micro Pillar Array Column Technology

Christof Mitterer(1), Robert van Ling (1), Geert Van Raemdonck(1), Jeff Op de Beeck(1), Kurt Van Mol(1), Bo Claerebout(1), Natalie Van Landuyt(1), Wim De Malsche(2), Gert Desmet(2), Paul Jacobs(1)
(1) PharmaFluidics NV, Ghent, Belgium (2) Vrije Universiteit Brussel, Brussels, Belgium

Warning: Undefined variable $headshot in /var/www/html/view_abstract/view_abstract_in_program.php on line 704

Christof Mitterer (Presenter) PharmaFluidics NV

Relevant Financial Disclosures (within past 24 months) Salary PharmaFluidics NV Discussing Products? YES, presenter will present educational content related to products/services of one or more ineligible company(ies) listed above. Conflict mitigation IN PROCESS.

INTRODUCTION
As an alternative to the conventional packed bed nano LC columns that are frequently used in bottom-up proteomics research, PharmaFluidics off¬ers micromachined nano LC chip columns known as micro pillar array columns (μPAC™). The inherent high permeability and low ‘on-column’ dispersion obtained by the perfect order of the separation bed makes μPAC™ based chromatography unique in its kind. The peak dispersion originating from heterogeneous flow paths in the separation bed is eliminated (no A-term contributions) and therefore components remain much more concentrated during separation resulting in unprecedented separation performance. The freestanding nature of the pillars also leads to much lower backpressure allowing an high operational flow rate flexibility with exceptional peak capacities.
RESULTS
Complementary to its landmark 200 cm long column which is ideally suited to perform comprehensive proteome research, a 50 cm long μPAC™ column is now available which can be used in a more routine research setting. With an internal volume of 3 μL, this column is perfectly suited to perform high throughput analyses with shorter gradient solvent times (30, 60 and 90 minute gradients) and it can be used over a wide range of flow rates, between 100 and 2000 nL/min. Recently performed experiments with 500 ng of HeLa cell digest indicate that an increase in protein identifications up to 50% and a gain of 70% in peptide identifications can be achieved when comparing the 50 cm µPAC™ column to the current state-of-the-art in packed bed columns. The conventional packed bed columns (2 different vendors) used for this benchmarking experiment were 15 cm in length and were packed with sub 2 µm porous silica particles. LC pump pressures needed to operate these classical columns at a flow rate of 300 nL/min range between 200 and 300 bar, whereas only 40 bar was need to operate the 50 cm µPAC™ column at the same conditions.
CONCLUSION
μPAC™ technology clearly offers several benefits regarding robustness, high operational flexibility and excellent separation performance compared to conventional packed bed column technology. When aiming for comprehensive proteome analysis with deep coverage, the 200 cm long μPAC™ column which delivers unprecedented separation performance is the best choice. However, the true benefits of using a long 200 cm μPAC™ column will only come into full play when working with long solvent gradient times (>120min). With an internal
column volume of approximately 3 μL and an increased operational flexibility (flow rates up to 2000 nL/min), the
50 cm μPAC™ column serves those who are looking for increased separation performance in daily routine proteome
analysis settings where shorter gradient times (<120min) and increased throughput are desired.