Cell Free Protein Expression, Purification, and Quantification

URL: https://cloud.ginkgo.bio/protocols/cfps-expression-purification-quantification Status: Ginkgo Certified Price: $159/sample Turnaround: up to 12 days Throughput: Up to 88 constructs per run (1 column reserved for controls), 96-well format

Overview

End-to-end automated cell-free expression, Strep-tag purification, and quantification of StrepII-tagged proteins, combining A280 yield with LabChip purity/size assessment. Linear DNA templates are expressed in CFPS reactions for 8 hours, purified with magnetic beads on the Agilent Bravo, quantified by A280 on the BMG PHERAstar, then characterized for purity and apparent molecular weight on the Revvity LabChip.

Input

• DNA Input: Linear DNA sequence (.xlsx template)
• Tag Orientation: N-terminal or C-terminal fusion (validated with C-terminal tags; success is protein-dependent)
• Format: Up to 88 constructs per run, 1 column of wells for controls

Output

• Expression Confirmation: Verification of the target protein at the expected molecular weight
• Baseline Titer: Initial quantitative yield measurement (mg/L)
• Initial Purity: Percentage of target protein vs. impurities, delivered with virtual gel images

Automated Workflow

Phase 1 - CFPS Reaction Setup & Incubation

1. Retrieve plates from storage (HRB TundraStore)
2. Stamp DNA templates into CFPS reaction mix (Agilent Bravo 96)
3. Seal plate (Agilent PlateLoc)
4. Incubate shaking at 29-30 deg C (Thermo Cytomat)

Phase 2 - Mag Bead Purification & Quantification

1. Dispense reagents and load samples (Biotek / Agilent)
2. Bind sample to beads (Bravo Shaker)
3. Wash beads (Agilent Bravo 96)
4. Elute purified protein (Agilent Bravo 96)
5. Transfer eluate to read plate (Agilent Bravo 96)
6. Read A280 & fluorescence (BMG PHERAstar)

Phase 3 - LabChip Purity and Size Assessment

1. LabChip assessment (Revvity LabChip)
2. Seal plate (Agilent PlateLoc)
3. Store at 4 deg C (HRB TundraStore)

Ordering

• Number of Proteins: configurable
• Number of Replicates: configurable
• File Upload: CSV, Excel, FASTA, TXT, PDF, ZIP
• Additional Details: free-text field for special requirements

Use Cases

• Purified yield plus purity/size profiling in one cell-free run
• Characterizing constructs before scale-up production
• Comparing titer and purity across sequence variants