Next-generation Cas9 proteins designed to deliver maximum CRISPR editing efficiency
TrueCut Cas9 Proteins are designed to deliver consistently high editing efficiency across a range of gene targets and cell types. We offer two performance-leading Cas9 proteins to better meet your genome editing goals—Invitrogen TrueCut Cas9 Protein v2, for most common research applications, and Invitrogen TrueCut HiFi Cas9 Protein, for applications that are sensitive to off-target effects.
TrueCut Cas9 proteins must be combined with a CRISPR guide RNA (gRNA), to produce a functional, target-specific editing complex. Learn more about our TrueGuide Synthetic gRNAs, predesigned for highly efficient gene knockout, or visit the TrueDesign Genome Editor to design custom gRNAs and DNA donors for precise genome engineering.
TrueCut Cas9 Protein Products
Choose the right Cas9 protein for your specific genome editing needs. TrueCut Cas9 (CTS-Prototype) is also available for pre-clinical research that requires a Cas9 nuclease that is manufactured with higher stringency specifications. Learn more about our TrueCut Cas9 Protein (CTS-Prototype).
Invitrogen TrueCut Cas9
Exceptional Editing Efficiency—Consistently high on-target editing efficiency in all tested cell lines, including standard, primary, stem, and immune (>90% editing in T cells)
Superior Performance—Up to 2X higher editing efficiency in difficult targets than the competition
High Quality—Manufactured under ISO 13485 quality standards
Invitrogen TrueCut HiFi
Improved Specificity—Significantly reduce off-target effects in a broad range of cell types including primary, stem, and immune cells
High Editing Efficiency—Preserved high on-target editing efficiency relative to TrueCut Cas9 v2 (wild type)
High Quality—Manufactured under ISO 13485 quality standards
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).
Robust editing efficiencies in cancer cell lines
Figure 2. 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
Achieve higher specificity in human primary T-cells
Figure 3. Superior off-target profiles in human primary T-cells when using TrueCut HiFi Cas9. 21 gRNAs targeting four therapeutic genes (CD52, TRAC, TRBC, and PD1) were co-transfected with TrueCut Cas9 v2 (Cat. No. A36497), TrueCut HiFi Cas9 (Cat. No. A50575), and other supplier's HiFi Cas9 using the Neon Transfection System. Target-enriched GUIDE-Seq (TEG-Seq), an Ion Torrent-adapted NGS method, was used for off-target detection. The red dots are on-target events and normalized to 100%. Gray dots are off-target events plotted against the corresponding off-target to on-target ratio, which represents the risk probability of each off-target event.
Increased specificity in human iPSCs
Figure 4. Increased specificity in human iPSCs with TrueCut HiFi Cas9. Two gRNAs targeted SNPs in the hemoglobin β subunit gene (HBB) associated with sickle cell anemia and another gRNA targeted a commonly studied off-target site in HEK4 were co-transfected with TrueCut Cas9 v2 (Cat. No. A36497), TrueCut HiFi Cas9 (Cat. No. A50575), and other supplier's HiFi Cas9. Read per millions (RPM) for all on- and off-target events are plotted in the bar graph.
Maintained high on-target editing efficiency in T-cells
Figure 5. TrueCut HiFi Cas9 maintained >80% of on-target editing efficiency in T-cells. Ion Torrent NGS-based Targeted Amplicon-seq was used for screening. Normalized on-target activity of wt-Cas9, Supplier I high-fidelity Cas9, and TrueCut HiFi Cas9 for indels and HDRs in T-cells.
Preserved high editing efficiency in human iPSCs
Figure 6. TrueCut HiFi Cas9 maintained high on-target editing efficiency in human iPSCs. On-target indel and HDR activity of TrueCut HiFi Cas9 with two gRNAs in iPSCs. One gRNA targeted SNPs in the hemoglobin ß subunit gene (HBB1) and another one targeted the BCL11A gene. Ion Torrent NGS-based Targeted Amplicon-seq was used for screening.
TrueCut Cas9 v2 consistently outperforms the competition
Figure 7. 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.
Superior off-target profiles with TrueCut HiFi Cas9 compared to the competition
Figure 8. TrueCut HiFi Cas9 reduced 80% more off-target events with high risk probability (>10%) compared to the competition in T-cells.. 21 gRNAs targeting four therapeutic genes (CD52, TRAC, TRBC, and PD1) were co-transfected with TrueCut Cas9 v2 (Cat. No. A36497), TrueCut HiFi Cas9 (Cat. No. A50575), and other supplier's HiFi Cas9 using the Neon Transfection System. Total number of off-target events grouped according to risk probability.
Reliable editing efficiencies across a broad range of cell lines and with different delivery methods
Figure 9. 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 Manual: TrueCut Cas9 Proteins
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.
Order Invitrogen TrueCut Cas9 Protein v2
Order Invitrogen TrueCut HiFi Cas9 Protein
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.
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For Research Use Only. Not for use in diagnostic procedures. Product is a prototype and performance characteristics of this product have not been established.