Next-generation Cas9 protein designed to deliver maximum CRISPR editing efficiency
TrueCut Cas9 Protein is designed to deliver consistently higher editing efficiency across a range of gene targets and cell types. We now offer two versions of our performance-leading Cas9 protein to better meet your genome editing goals—Invitrogen TrueCut Cas9 v2, for most common research applications, and our new Gibco TrueCut Cas9 Protein (CTS-Prototype), for pre-clinical research that requires a Cas9 nuclease that is manufactured with stringent specifications.
TrueCut Cas9 Protein Products
Choose the right Cas9 for your specific genome editing needs—from basic research to preclinical research and process development. No matter which you choose, both versions offer superior performance and value.
Invitrogen TrueCut Cas9
- Consistent high performance in all tested cell lines including standard, primary, stem, and immune (>90% editing in T cells).
- Up to 2X higher editing efficiency in difficult targets than the competition
- High quality: manufactured under strict ISO 13485 quality standards
Gibco TrueCut Cas9
- Consistent high performance in all tested cell lines (>90% editing in T cells).
- Higher stringency manufacturing in compliance with standards for Ancillary Materials for Cell and Tissue-Based Products including USP <1043>, Ph.Eur. 5.2.12, and ISO 20399 -1, -2, -3, following the principles of 21 CFR Part 820 in a FDA–registered manufacturing site.
- Traceability documentation and extensive safety testing, including bioburden, endotoxin, mycoplasma, residual host nucleic acids and protein
- Aseptic manufacturing using single-use technology
- Sterile filling in a Class A/B cleanroom
- Large pack sizes: 2.5 and 5.0 mg at a 10 mg/mL concentration
Truly stunning performance, reliable results, and more choices—your research demands nothing less
Achieve functional knockout in up to 90% of transfected human primary T-cells
Figure 1. T cell receptor knockout using TrueGuide synthetic gRNAs. Human T cells were isolated and activated using Invitrogen Dynabeads magnetic beads (Cat. Nos. 11344D and 11141D, respectively) and then transfected with TrueCut Cas9 Protein v2 and TrueGuide Synthetic gRNA for T cell receptor alpha (TRAC) or beta (TRBC) regions using the Invitrogen Neon Transfection System. Following transfection, editing efficiency was measured by the Invitrogen GeneArt Cleavage Detection assay (A), or by measuring % T cell receptor negative (TCR–) cells using the Invitrogen Attune NxT Flow Cytometer after the cells were stained with a PE-labeled T cell receptor–specific antibody conjugated to PE (B).
Consistently outperforms the competition
Figure 2. Invitrogen CRISPR genome-editing tools consistently outperform products from other suppliers. Genome editing of multiple gene targets was performed with TrueCut Cas9 Protein v2 and corresponding TrueGuide Synthetic gRNAs. Delivery was achieved using optimized transfection protocols and Invitrogen Lipofectamine CRISPRMAX Transfection Reagent in two cell lines, (A) A549, a human lung carcinoma cell line and (B) MDA-MB231, a human breast cancer cell line. The graphs compare the same experiments using products and recommended protocols from another manufacturer. With the Invitrogen system, cleavage efficiency is improved for low-efficient loci (PRKCG-T1 and CMPK1-T1) and shows consistently greater efficiency when compared to products and protocols from other suppliers, even for challenging loci.
Robust editing efficiencies in cancer cell lines
Figure 3. Robust editing efficiencies in cancer cells. A wide range of cancer cell lines and gene targets were tested to determine editing efficiencies using the modified single TrueGuide Synthetic gRNA complexed with TrueCut Cas9 Protein v2 (Cat. No. A36497). Here, the Cas9 RNP complex was delivered into three different cancer cell lines using Lipofectamine CRISPRMAX Cas9 Transfection Reagent (Cat. No. CMAX00003) and, at 72 hours posttransfection, cells were harvested and tested for cleavage efficiency using either the GeneArt Genomic Cleavage Detection Assay (GCD) or Ion Torrent Next-Generation Sequencing (NGS). Panel ( A) A549, an epithelial lung carcinoma cells, (B) U2OS, osteosarcoma line, (C) MDA-MB231, epithelial (breast) adenocarcinoma line
Reliable editing efficiencies across a broad range of cell lines and with different delivery methods
Figure 4. Consistent editing efficiency across a wide range of cell types. TrueCut Cas9 Protein v2 has been tested using lipid-mediated delivery or the Neon electroporation transfection system across a wide range of cell types. Here we demonstrate high efficiency editing in four representative cell types including adherent cell lines (A549, HepG2), suspension cells (THP1) and primary immune cells (primary human T cells). Cell lines were transfected with TrueCut Cas9 Protein v2 (Cat. No.A36497) and the TrueGuide Positive Control crRNA (Cat. No.A35517) and TrueGuide tracrRNA (Cat. No. A35506).
User guide and protocols
Getting started with CRISPR? Need proven CRISPR validated protocols? Our CRISPR Validated Protocols collection has been developed by our expert R&D team. We have matched each of the validated and optimized protocols with the best products to help reduce trial and error and to accelerate your rate of discovery. These step-by-step CRISPR protocols have been optimized for maximum efficiency, viability, and reproducibility across a broad range of cell types and gene targets. We truly want you to succeed with your CRISPR experiment the first time and have confidence in your data.
Cas9 (CTS-Prototype) Performance
Figure 5. Both TrueCut Cas9 products exhibit exceptional performance with >90% editing efficiency in human primary T cells. Human peripheral blood mononuclear cells (PBMC) were thawed into CTS Optimizer medium (Cat. No. A1048501) (with L-glutamine and 2% human serum) in presence of IL2 (Cat. No. CTP0023) and T expander CD3/CD28 Dynabeads (Cat. No. 11131D). On day three, 2 x 105 cells were electroporated with the Neon Transfection System (Cat. No. MPK5000) with 1.0, 2.5 or 10 μg of the Invitrogen TrueCut Cas9 Protein v2 (Cat. No. A36499) or Gibco TrueCut Cas9 (CTS-Prototype) (Cat. No. A45220P) complexed with 240 ng, 600 ng or 2,400 ng, respectively, of TrueGuide modified sgRNA (Cat. No. A35534) against the TRAC locus. Electroporation was done using program 5 pulse setting -1700V, 20 ms, 1 pulse. On day six, cells were stained with Fixable Viability Dye (FVD) Violet (Cat. No. 65-0863-14) and anti-TCRa/b antibodies (Cat. No. 12-9986-42) and analyzed on the Attune NXT flow cytometer. Gating was performed to include only viable cells.
FAQ for Cas9 (CTS-Prototype)
Do you have a GMP Cas9?
We do not currently sell a GMP Cas9 product. However, we manufacture TrueCut Cas9 Protein (CTS-Prototype) in compliance with standards for Ancillary Materials for Cell and Tissue-Based Products including USP <1043>, Ph.Eur. 5.2.12, and ISO 20399 -1, -2, -3, following the principles of 21 CFR Part 820 in a FDA–registered manufacturing site utilizing aseptic manufacturing principles in qualified cleanroom facilities. We have early access to this CTS material now which is termed “prototype.”
What’s the difference between TrueCut Cas9 Protein (CTS-Prototype) and TrueCut Cas9 Protein v2?
Both products are the same Cas9 protein, but TrueCut Cas9 Protein (CTS-Prototype) is manufactured with a new production process and formulated to meet the standards for Ancillary Materials for Cell and Tissue-Based Products. This has the following added benefits:
- Traceability documentation—including Drug Master Files*, and/or Regulatory Support Files are available†
- Extensive safety testing—including bioburden, endotoxin, mycoplasma, residual nucleic acids and proteins.
- Aseptic manufacturing—entire process completed using single-use technology
- Sterile filling—the finished product is vailed in a Class A/B (Class 100 and ISO 5 equivalents) Cleanroom
- Large pack sizes—2.5 and 5.0 mg with a 10 mg/mL concentration
*Drug Master File available after completion of real time stability and product validation in 2021. †Country-specific Regulatory Support File process will be initiated upon request; will require additional documentation time.
Can I use the prototype for clinical manufacturing?
The prototype is marked for “Research Use Only” and is acceptable for preclinical work. Our goal is provide this material to scientists as early as possible to aid in preclinical research and process development. CTS products have a particularly long development cycle –namely due to the real-time stability data requirement, which extends the timeline by at least 13 months. During this period, we are making the same material available, without full validation and real-time stability information, to aid in preclinical development.
CRISPR has a lot of patents. Does Thermo Fisher Scientific have clinical rights?
Thermo Fisher Scientific’s Limited Use Label licenses associated with our Cas9 products address the use of TrueCut Cas9 Protein (CTS-Prototype) for discovery research, preclinical research, and clinical research activities that fall within safe harbor statutes of 35 U.S.C. §271(e)(1). The scope of activities that are covered by 35 U.S.C. §271(e)(1) needs to be determined by the customer.Anything beyond this scope puts onus on the customer to obtain any required third-party rights.
|Activity Assay, in vitro||>90% in vitro of uncut reference DNA converted to cleavage products|
|Endotoxin by LAL||<10.0 EU/mg|
|Purity by HPLC-DAD||≥ 95%|
|Purity by SDS-PAGE||≥ 95%|
|Aggregation by SEC-HPLC||≤ 5%|
|pH||7.0 - 7.8|
|Residual Host Protein||<10.0 ng/mL|
|Residual DNase||<LOQ* DNaseALERT|
|Residual RNase||<LOQ* RNaseALERT v.2|
|Residual Host Genomic DNA||<LOQ* qPCR|
|Residual Plasmid DNA||<LOQ* qPCR|
*LOQ = Limit of Quantification
Review more in these publications
Order Invitrogen TrueCut Cas9 Protein v2
Order Gibco TrueCut Cas9 Protein (CTS-Prototype)
TrueGuide Synthetic Guide RNA
Incorporate the latest in gRNA design research coupled with our extensive in-house experience. Our proprietary gRNA design algorithms select gRNAs for maximum editing efficiency without compromising specificity.
Can’t find what you are looking for? Contact us at CRISPR@thermofisher.com
For Research Use Only. Not for use in diagnostic procedures. Product is a prototype and performance characteristics of this product have not been established.