AssayMAP. Mitu Chaudhary Product Marketing Manager Bay area product specialist Robert (Bob) Boice

Transcription

1 AssayMAP Quantitative, Automated, Analytical Scale Protein Sample Preparation Mitu Chaudhary Product Marketing Manager Bay area product specialist Robert (Bob) Boice LRIG Bay Area, December 2012

2 AssayMAP Biomolecule Sample Prep Solutions Samples Automation AssayMAP platform = Automated Sample Prep Workflows Chemistry Analysis The image cannot be displayed. Your computer may not have enough memory to open the image, or the image may have been corrupted. Restart your High throughput & time to results High precision & reproducibility Miniaturization (reduced sample, ligand, etc.) Open access ( scientist-friendly ) automation Gold standard chemistries

3 AssayMAP Cartridges Chromatography column on a micro scale. 3

4 AssayMAP Bravo Liquid Handler Best in Class Compact Liquid Handling Accessorize the Deck to Any Application AssayMAP Head with Probe Syringe 4

5 Simplified User Interface with Application-specific Forms AssayMAP Platform = AssayMAP cartridge, Bravo with AM head, VWorks with forms Page 5

6 Portfolio Summary- Biologics Characterization Workflows Quan5ta5on Biomanufacturing Process Elute Target PA-W PG SA-W Pep5de Mapping TR-W RP-S Elute Target Digest Target Pep5de Cleanup Affinity Capture N- glycan analysis RX CU from Cleave glycan label glycan Glycan cleanup

7 7 Protein A

8 Protein A for Ab purification Sample Analysis 8

9 Protein A Performance Characteristics µg Eluted PA- W Identity Linear (Identity) µg higg Loaded Quantitative Capture Quantitative Elution pure concentrated sample

10 Protein A: Reproducibility & Precision µg load N = 96 %CV = 1.26 A µg higg Loaded N = 8 µg higg loaded %CV

11 Customer testimonial for Protein A Novo Nordisk, Seattle Purification Tips Purification Plates Magnetic Beads AssayMAP Bravo* Cost $150/plate $750/plate $1200/plate $170/plate % Recovery 20-30% 10-15% 20-30% ~80% Automatable Yes difficult Semi Fully Notes >20µg input -- >20µg input <5 µg * Unoptimized, beta cartridges with beta hardware!

12 12 Streptavidin

13 Streptavidin- SA-W Bravo Work Flow Immobilization - Single pass - Irreversible - Generates an affinity cartridge Use of the affinity cartridge - Reusable - Bind, wash and elute - Purify and quantify target molecule Anything you can biotinylate can be a capture ligand!

14 Streptavidin (SA-W) Elution of FLAG-BAP from biotinylated αflag SA-W. 14

15 15 Trypsin

16 On-Cartridge Trypsin Digestion 16

17 Trypsin Cartridge (TR-W) TR-W comparisons 200 DAD: Signal A, 215 nm/bw:4 nm DAD: Signal A, 215 nm/bw:4 nm 200 Urea 1.11 pre-digest Urea 1.11 digest urea 1.11 pre-digest pm.dat urea 1.11 digest pm.dat Urea Analyte: BSA on RP-HPLC Flow Rate: 2 µl/min. Load: 67 µg mau mau TR-W comparisons 200 DAD: Signal A, 215 nm/bw:4 nm DAD: Signal A, 215 nm/bw:4 nm 200 TFE 18.1 pre-digest TFE 4.50 digest tfe 18.1 pre-digest pm.dat tfe 4.50 digest pm.dat Minutes 160 TFE Analyte: BSA on RP-HPLC Flow Rate: 2 µl/min. Load: 67 µg mau mau Minutes 0

18 18 Reverse Phase

19 Reverse Phase for Peptides Peptide Digest 19

20 Reverse Phase (RP-S) µg Insulin in Flow Through or Elution Quantitative Binding and Elution to ~400 µg Elution Flow through µg Insulin loaded Complete elution 1.8 in 2 column vols (10 µl) 1.6 A µL/min 5µL/min 10µL/min 20µL/min Cumulative Elution Volume (µl) 20

21 Reversed- Phase PepIde Cleanup Analysis on Agilent 1260 HPLC with an Agilent Poroshell 120 SB-C18 column 21

22 N-Glycan Sample Preparation 22

23 GlykoPrep N- Glycan Kit- CollaboraIon with Crude sample Pure sample Analysis by HPLC, CE, LC/MS Purify with Protein A Denature & immobilize Digest with PNGaseF Fluorescent label Cleanup & elute PA-W SA-W RX CU Conven5onal method- manual, 3 days, 10 s of samples per run AssayMAP method- automated, <3 hours, 100 s of samples per run

24 RelaIve Peak Area Reproducibility 12 consecuive runs α- 1 Acid Glycoprotein 40% 35% % Total Area for Major Peaks Peak 3 CV = 1.6% 30% 25% 20% 15% 10% 5% 0% Well Number Peak 4 CV = 1.5% Peak 1 CV = 2.6% Peak 6 CV = 3.2% Peak 2 CV = 2.7% Peak 5 CV = 3.1% Peak 1 Peak 2 Peak 3 Peak 4 Peak 5 Peak 6

25 AssayMAP Cartridge Portfolio Affinity Purification Protein A Streptavidin Protein/Peptide Analysis Trypsin Reverse Phase N-Glycan Analysis- GlykoPrep 25

26 AssayMAP technology adoption Several active customers in Americas, Europe and Asia. Purification is the most commonly used application. Glycan analysis sample preparation and digestion increasingly being adopted. Technology collaborations with key industry leaders in all geographies. Collaborations include sample preparation for biomarker/ proteomics experiments (Broad Institute, among others).

27 Summary of AssayMAP Bravo Por\olio High throughput microchromatography plaworm 5 µl packed bed cartridge with any µm par5cle resin Best in class 96- well liquid handler Single pass, fully quan5ta5ve binding & elu5on Posi5ve displacement, bidirec5onal flow control Integrated workflows involving purifica5on, quan5ta5on & reac5on Mul5ple applica5ons in biopharmaceu5cal development Affinity purifica5on N- glycan sample prep Pep5de mapping 27