{"id":11553,"date":"2025-03-17T13:21:45","date_gmt":"2025-03-17T13:21:45","guid":{"rendered":"https:\/\/www.caltagmedsystems.co.uk\/information\/?p=11553"},"modified":"2025-03-18T15:53:48","modified_gmt":"2025-03-18T15:53:48","slug":"crispr-cas9-protein","status":"publish","type":"post","link":"https:\/\/www.caltagmedsystems.co.uk\/information\/crispr-cas9-protein\/","title":{"rendered":"KACTUS: CRISPR Cas9 Protein"},"content":{"rendered":"\n

Highly-active GMP-grade CRISPR Cas9 protein<\/h2>\n\n\n\n

Due to the ease of design and operation, the CRISPR-Cas system has become the most widely used gene editing technology today. At KACTUS, they have developed a highly-active, wild-type Cas9 nuclease based on their SAMS\u2122 technology platform<\/a>. Their Cas9 nuclease has been codon-optimised and process-optimised to reach the best yield and purity. The addition of the nuclear localisation signal (NLS) makes it suitable for gene editing work in eukaryotic cells.<\/p>\n\n\n\n

KACTUS’ GMP-grade, recombinant CRISPR Cas9 protein meets the highest quality and performance standards for gene editing research and manufacturing. Their GMP-grade Cas9 enzyme is manufactured in compliance with cGMP standards and accompanied by comprehensive quality documentation for regulatory support, making it eligible for CGT process development and CMC manufacturing. Finally, with a well-established global supply chain, they are capable of stably supplying our Cas9 enzyme in bulk quantities worldwide.<\/p>\n\n\n\n

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View Cas9 Products ><\/strong><\/a><\/div>\n<\/div>\n\n\n\n

Why choose KACTUS’ Cas9 protein?<\/h2>\n\n\n\n
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\"Bioactivity<\/figure>\n\n\n\n
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GMP Production<\/h3>\n\n\n\n

Analytical Method Validation<\/h3>\n\n\n\n

Batch Consistency<\/h3>\n\n\n\n

Their GMP-grade Cas9 is manufactured in compliance with GMP standards, ensuring the highest levels of quality and safety. They offer both research-grade and GMP-grade versions to seamlessly transition from preclinical study to clinical\/commercial production.<\/p>\n\n\n\n

KACTUS validate all analytical methods according to pharmacopeia standards, ensuring consistent performance and reliability of their CRISPR Cas9 protein. This rigorous validation process guarantees that their product meets the most demanding quality benchmarks in the industry.<\/p>\n\n\n\n

Their rigorous quality control ensures every batch of our CRISPR Cas9 protein is manufactured with consistent quality and activity, providing you with a reliable product that delivers solid, consistent results every time. This consistency is critical for reproducible research outcomes and regulatory submissions.<\/p>\n\n\n\n

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High editing activity<\/h3>\n\n\n\n

QC & Stability Testing<\/h3>\n\n\n\n

Quality Documentation<\/h3>\n\n\n\n

Their CRISPR Cas9 protein demonstrates high editing activity, both\u00a0in vitro\u00a0<\/em>and\u00a0in vivo<\/em>, enabling precise and efficient genome editing. This ensures that your gene editing experiments are both effective and reliable, reducing the need for extensive optimisation.<\/p>\n\n\n\n

KACTUS conduct comprehensive sterility and stability testing on their CRISPR Cas9 protein to ensure that it remains effective and safe over time. This confirms the protein\u2019s longevity and reliability, even under varying storage and usage conditions. This documentation is also available to support regulatory filings.<\/p>\n\n\n\n

They provide comprehensive quality documentation to support your IND and BLA filings, This includes animal-free origin statements (TSE\/BSE), certificates of analysis, batch records, and other required documents to facilitate smooth regulatory submissions and approvals. Learn more about their GMP production \u2192<\/a><\/p>\n<\/div>\n\n\n\n

Product Performance Data<\/h2>\n\n\n\n

KACTUS has conducted extensive performance and analytical testing on their Cas9 protein to ensure high editing activity and quality. Their GMP-grade Cas9 protein demonstrates strong editing activity across various cell types, comparable to industry standards. They showcase successful gene knockouts in diverse cell lines, including 293T, Jurkat, and primary T cells, with editing efficacies greater than 85%. Their team has also conducted comprehensive stability testing on the Cas9 enzyme to underscore its reliability for use clinical and commercial applications.<\/p>\n\n\n\n

High Editing Activity in Multiple Cell Types<\/h3>\n\n\n\n
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(A)\u00a0BCL11A<\/em>\u00a0Knockout in 293T Cells<\/figcaption><\/figure>\n\n\n\n
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(B)\u00a0BCL11A<\/em>\u00a0Knockout in Jurkat Cells<\/figcaption><\/figure>\n\n\n\n
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(C)\u00a0TRAC<\/em>\u00a0Knockout in Primary T Cells<\/figcaption><\/figure>\n<\/div>\n\n\n\n

<\/a>Figure 1. Gene knockout in 293T (A), Jurkat (B), and T cells (C) with different batches of KACTUS GMP-grade Cas9 protein. Results show greater than 85% editing efficacy across all three cell types, comparable to a leading supplier. Briefly, 75 pmol of Cas9 protein was electroported along with 225 pmol of sgRNA into various cells using the Lonza 4D Nucleofector system. The cells were cultured for three days followed by TIDE analysis. Gene knockouts: (A) knockout\u00a0BCL11A<\/em>\u00a0in 293T cells; (B) knockout\u00a0BCL11A<\/em>\u00a0in Jurkat cells; (C) knockout\u00a0TRAC<\/em>\u00a0in Primary T cells.<\/p>\n\n\n\n

Comparable activity to leading suppliers<\/h3>\n\n\n\n
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Figure 2. Comparison of gene knockout efficiency using Cas9 nucleases from KACTUS and two leading suppliers in UCAR-T cells. Gene knockout efficiency was assessed at 72 hours and 120 hours post-electroporation. Results indicate that KACTUS Cas9 nuclease performs comparably to leading suppliers, effectively knocking out genes related to immune rejection reactions on donor T cells.<\/p>\n\n\n\n

Figure 3. Gene knockout efficiency of BCL11A<\/em> in hematopoietic stem cells (HSCs) for the treatment of \u03b2-thalassemia using Cas9 nucleases from KACTUS and two leading suppliers. Results demonstrate that KACTUS Cas9 nuclease achieves gene knockout efficiency comparable to that of the leading suppliers, validating its effectiveness for therapeutic applications.<\/p>\n<\/div>\n\n\n\n

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<\/a>Batch consistency and long-term stability<\/h3>\n\n\n\n

KACTUS’ rigorous quality control ensures every batch of their CRISPR Cas9 protein is manufactured with consistent quality and activity, providing you with a reliable product that delivers solid, consistent results every time. This consistency is critical for reproducible research outcomes and regulatory submissions.<\/p>\n\n\n\n

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<\/a>Figure 4. Consistent purity and activity performance was observed across three manufacturing batches of KACTUS GMP-grade Cas9. Three batches of KACTUS GMP-grade Cas9 were stored at -20\u00b0C for long-term stability testing. Purity and\u00a0in vitro<\/em>\u00a0cleavage activity were analysed at 3, 6, 9, 12, 15, 18 months, respectively. Results show their GMP-grade Cas9 has a good stability and consistent activity across time and different manufacturing batches.<\/p>\n\n\n\n

Product Specifications & Quality<\/h2>\n\n\n\n

Product Specifications<\/h3>\n\n\n\n

Both their GMP-grade\u00a0and\u00a0research-grade\u00a0Cas9 protein adhere to the following product specifications.<\/p>\n\n\n\n

Parameter<\/th>Specification<\/th><\/tr><\/thead>
Express System<\/td>E. coli with CRISPR Cas9 gene of S. pyogenes<\/td><\/tr>
Concentration<\/td>9.5-12.5 mg\/mL<\/td><\/tr>
Form<\/td>Liquid<\/td><\/tr>
Storage Buffer<\/td>30mM Tris-HCl, 0.3M NaCl, 50% Glycerol, 0.1 mM EDTA, pH 7.4<\/td><\/tr>
Storage<\/td>-20\u00b15\u00b0C for 18 months<\/td><\/tr>
Purity (Bis-Tris)<\/td>\u2265 95.0%<\/td><\/tr>
Purity (SEC-HPLC)<\/td>\u2265 95.0%<\/td><\/tr>
Activity<\/td>> 85.0%<\/td><\/tr>
Residual RNase<\/td>Negative<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n

Quality Control Criteria<\/h3>\n\n\n\n

Each batch of GMP-grade\u00a0Cas9 Protein is tested for and meets the quality control criteria defined in the table below. Their research-grade\u00a0Cas9 protein demonstrates similar performance but does not undergo lot-specific testing for all QC criteria.<\/p>\n\n\n\n

Assay<\/th>Acceptance Criteria<\/th><\/tr><\/thead>
Appearance<\/td>Clear Liquid<\/td><\/tr>
Loading Amount<\/td>Not less than the amount of identification<\/td><\/tr>
Concentration<\/td>9.5-11.5mg\/mL<\/td><\/tr>
Identification<\/td>Corresponding to Reference Spectrum<\/td><\/tr>
Purity (RP-HPLC)<\/td>\u2265 95.0%<\/td><\/tr>
Purity (SEC-HPLC)<\/td>Monomer \u2265 95.0%, Aggregates \u2264 5.0%<\/td><\/tr>
Purity (NR-CE)<\/td>\u2265 85.0%<\/td><\/tr>
Purity (R-CE)<\/td>\u2265 90.0%<\/td><\/tr>
Bacterial Endotoxin<\/td>\u2264 10.0 EU\/mg, (LOQ=0.1EU\/mL)<\/td><\/tr>
Activity<\/td>\u2265 85.0%<\/td><\/tr>
Residual DNase<\/td>Negative (LOD=0.33U\/\u03bcL)<\/td><\/tr>
Residual RNase<\/td>Negative (LOD=2.2\u00d710\u207b\u2077U\/\u03bcL)<\/td><\/tr>
Residual Host Cell Protein<\/td>\u2264 100.0 ng\/mL (LOQ=3ng\/mL)<\/td><\/tr>
Residual Host Cell DNA<\/td>\u2264 3.0 ng\/mL (LOQ=0.03ng\/mL)<\/td><\/tr>
Sterility<\/td>No growth<\/td><\/tr>
Residual Nickel Salt<\/td>\u2264 10.0 ppm (LOD=10.0ppm)<\/td><\/tr>
pH<\/td>7.4\u00b10.5<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n

GMP-Grade Production<\/h2>\n\n\n\n

Their GMP-Grade Cas9 Protein\u00a0is produced following cGMP standards, ensuring the traceability of raw materials, manufacturing process and QC release. This Cas9 protein is available in long-term bulk supply with batch-to-batch consistency to ensure suitability for industrial applications.<\/p>\n\n\n\n

Documentation Package<\/h3>\n\n\n\n

Cas9 nuclease, GMP-grade\u00a0comes with customisable regulatory documentation including Data Sheet, MSDS, COA, TSE\/BSE statement, CoO. KACTUS has an integrative documentation system consisting of a digital quality management system, specification program, criteria support documents, and quality reports. Their product release process involves 15+ contaminant detection steps, 30+ quality tests, 4+ report reviews, and 4+ product release checkpoints. The documentation package is customisable based on relevant regulatory filings.<\/p>\n\n\n\n

Quality Management System<\/h3>\n\n\n\n