Overview Service Features FAQs Scientific Resources Related Services
Overview
Creative Biolabs is dedicated to providing several viable and cost-effective methods for pluripotent stem cells (iPSC) reprogramming. We employ advanced iPSC reprogramming factor delivery strategy by virus, iPSC reprogramming factor delivery by episomal vectors, as well as other iPSC reprogramming methods (mRNA, protein) to help you obtaining the desired iPSC.
Typically, during human disease modeling and cell-based therapies, the capacity to reprogram somatic cells to induce iPSCs has provided great value. Nowadays, iPSC derivatives have already reached the stage of clinical testing in human patients. Using different techniques to produce induced iPSCs needs that scientists choose the suitable reprogramming method for generating iPSCs, thus the resulting iPSCs enable to be transitioned towards clinical applications effectively. Generally steps in a reprogramming experiment include tissue selection, proceeding through iPSC generation, possible transgene excision to produce iPSC cells that are ready for use in a translational setting.
Fig 1. Schematic workflow of iPSC Reprogramming.
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Retro-and lentivirus: Retro- and lentiviral delivery vectors have been used to generate iPSCs almost every lab, because this viral transduction is easily implemented in most biomedical research labs. Using this delivery system, the reprogramming efficiency was 0.01–0.02% in human cells with iPSC colonies occurring between 25 and 30 days post-infection. As retroviruses only infect dividing cells, lentiviral delivery system was more popular so that both dividing and non-dividing cells could be infected in the hopes of improving reprogramming efficiency.
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Lentiviral plus cre-lox mediated transgene excision: To date, the first production of transgene free iPSCs were performed with lentiviral vectors containing loxP sites in the 5′and 3′LTR of the viral vectors. The existence of loxP sites gave a substrate to remove most of the transgene sequences by Cre-mediated recombination.
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Adenovirus: Adenovirus is a non-integrating virus that does not infect replicating cells. The first known research reporting the production of iPSCs with an adenoviral vector was reprogrammed mouse tail tip fibroblasts to iPSCs. However, the reprogramming efficiency was in the range of 0.001–0.0001%, because the expression window of reprogramming factors is too narrow to induce the expression of endogenous factors required to transition somatic cells.
Episomal Vectors for iPSC reprogramming are vectors developed to offer the preeminent system for performing transgene-free and virus-free induced iPSCs in a feeder-free environment. Until now, a great number of different somatic cell types have been successfully reprogrammed by using episomal iPSC reprogramming vectors. This advanced system possesses several advantages:
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Absolute security during your iPSC project development
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Wide range of application for a great number of somatic cell types
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Optimized for feeder-free reprogramming
It has proven that direct expression of reprogramming factors as proteins also enable to generate footprint-free iPSCs. Therefore, this approach may be another good choice for the generation of iPSCs appropriate for studies in translational medicine.
Fig 2. Schematic diagram of the generation of human iPSCs from somatic cells of a healthy or diseased individual.
For iPSCs creation, expression of reprogramming factors as mRNAs is also an available zero-footprint technology. However, the strong immunogenic response elicited in cells by the introduction of synthetic mRNA became a bid problem to reprogramming with mRNA. Scientists have solved this problem through modified the RNA bases by substituting 5-methylcytidine for cytidine and pseudouridine for uridine, as well as added the interferon inhibitor B18R into a cell culture media. These modification extremely reduced cell death which derived from the strong antiviral response that is formed when cells come into contact with mRNA containing standard ribonuclease bases in a common cell culture medium.
Reprogramming by Minicircle Vectors
Minicircle vectors are circularized vectors and their plasmid backbone has been released leaving only the eukaryotic promoter and cDNA(s) that are to be expressed. A minicircle vector composed of Lin28, GFP, Nanog, Sox2, and Oct4, and enables to be used to reprogram human adipose stem cells. Usually, the reprogram steps contain three transfections of the minicircle vector: an initial electroporation, followed by sorting of GFP+ cells, then two lipid-based transfections.
With professional scientists devoted themselves in iPSC reprogramming, Creative Biolabs is dedicated to providing the first class iPSC reprogramming services for our customers. Our services will contribute greatly to the success of your iPSC programs. Please contact us for more information and a detailed quote.
Features of Our Services
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Comprehensive services - We perform all aspects of iPSC reprogramming, from cell isolation and cultivation, reprogramming, expansion, and cryopreservation of potential iPSC clones to downstream validation (e.g., pluripotency and karyotyping) using state-of-the-art technology.
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High reprogramming efficiency – Based on our optimized protocols and advanced delivery systems, we can ensure high reprogramming efficiency, increasing the likelihood of obtaining iPSC lines with minimal passage.
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Speed and cost-effective - Our service is designed to save your time and resources, by leaving the complex and lengthy process of iPSC generation and validation to our experienced team.
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Exceptional quality control - Each step of our process is under rigid quality control guidelines, ensuring the generation of high-quality, purity, and genetically stable iPSC lines.
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Comprehensive analysis - Every iPSC line we produce comes with a detailed analysis, including documentation of morphology throughout the process, growth rate, and results of pluripotency and other selected tests.
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Customization - We offer flexible service packages tailored precisely to the specific needs of our clients. You can choose to start with primary cells, or established cell lines, and choose the specific validation tests that best suit your needs.
By utilizing our iPSC reprogramming services, researchers and companies can accelerate research and development work and contribute to breakthroughs in regenerative medicine, disease modeling, and drug discovery. Our team remains at your disposal for the duration of the project, providing timely updates and addressing any issues or clarifications you may need.
FAQs
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Q: What types of cells can be reprogrammed into iPSCs using this service?
A: We can reprogram a range of cells including skin fibroblasts, blood cells, or Urine cells and more. The choice of somatic cell source typically depends on the availability and ease of collection. Please contact us with specifics about your source cells for more detailed information.
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Q: How are iPSCs stored once reprogramming is complete?
A: Upon completion, iPSCs are cryopreserved and stored in liquid nitrogen. We will ship the cryovials to you under carefully controlled conditions, ensuring cell viability.
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Q: Can the iPSCs be differentiated into any cell type?
A: In theory, iPSCs have the capacity to differentiate into any cell type in the body. Certain specific lineages, however, may be more challenging than others and may require specific differentiation protocols.
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Q: What kind of support do you offer after the service is done?
A: We supply extensive post-service support, including guided protocols for culture and maintenance of the generated iPSCs. If you confront difficulties or have any concerns, our team of experts will be readily available to provide professional advice and solutions.
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Q: How long does the reprogramming process take?
A: The entire iPSC reprogramming process typically takes around 4-6 weeks. This timeline may vary based upon the cell type provided for reprogramming and the nature of the customer's specific requirements.
Scientific Resources
References
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Rao, M. S., (2012). “Assessing iPSC reprogramming methods for their suitability in translational medicine.” Journal of cellular biochemistry, 113(10), 3061-3068.
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Dash, B. C., (2015). “Induced pluripotent stem cell-derived vascular smooth muscle cells: methods and application.” The Biochemical journal, 465(2), 185.
For Research Use Only. Not For Clinical Use.
