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Molecular Biology Unit
The Molecular Biology Unit is designed to prevent any contamination of the final product. The unit consists of three labs: a Master Mix Lab, a Pre-PCR Lab, and a PCR and Post-PCR Lab. The latter is kept under negative pressure to prevent amplified target contamination.
Cell Culture and Immunology Unit
Normal and Cancer Cell Line Maintenance and Propagation
Maintenance and propagation of cell lines are vital processes that assist in developing extended cell lines library at MARC’s cell culture unit. By supplying the proper media that provides the cells with nutrients they need, as well as incubators and biological safety cabinets that offer a sterile environment for growth at an ideal temperature under aseptic conditions which prevent contamination with opportunistic microorganisms, we can tailor the cell culture environment for all cell lines.
Flow Cytometry-Based Cell Cycle Analysis
Flow cytometry is a powerful technique that uses one or several lasers to give a multi-parametric study of individual cells. When cells suspended in fluid injected into the flow cytometer, it quickly moves through laser beam and visible light scattering, or fluorescence is used to evaluate each cell or particle. Fluorescence measurements are made possible by transfecting cells, expressing common fluorescent proteins (such as the Green Fluorescent Protein, or GFP), and then staining the cells with fluorescently conjugated antibodies or fluorescent dyes. Cell cycle analysis is done by staining the DNA of mammalian cells by various DNA binding dyes that bind proportionally to the amount of DNA present in the cell.
Apoptosis and Necrosis Assays
An apoptosis assay measures the biological processes connected to programmed cell death, such as DNA fragmentation, phosphatidylserine (PS) exposure on the cell surface, and caspase activation. Annexin V-FITC staining is used to detect apoptosis by binding to the PS molecules that are exposed to the outside of cell membrane. Propidium iodide is used with Annexin V-FITC to differentiate necrosis from apoptosis. Necrosis and apoptosis assays allow researchers to understand the overall health status of their cell populations.
Cell Line Labeling and Fluorescence imaging
Fluorescence microscopy of live cells has become integral to modern cell biology. We use a range of tools to fluorescently label proteins of interest, including fluorescent protein tags, live cell dyes, and other methods that provide a range of tools to investigate virtually any cellular process under the microscope.
Cytotoxicity Assay
The ability of a drug to harm or kill cells is measured through procedures known as cytotoxicity assays. In basic research and drug development phases, cytotoxicity assays are frequently used to screen libraries for dangerous compounds. We use two assays to measure cytotoxicity (% viability): 1) MTT assay is used to measure cellular metabolic activity as an indicator of cell viability, proliferation and cytotoxicity. 2)Cell Counting Kit-8 (CCK-8)** which allows sensitive colorimetric assays for the determination of the number of viable cells in the proliferation and cytotoxicity assays its sensitivity is higher than MTT.
Cell Migration Assay
Cell migration is a key procedure involved in many biological processes including embryological development, tissue formation, immune defense or inflammation, and cancer progression. We perform cell migration either by using wound healing or transwell cell migration/invasion assays.
Drug Screening
Drug screening aids in exploring candidate drugs to show their effectiveness and determine whether they are likely to have any adverse effects. The effects of a potential innovative drug on cells are investigated in vitro using tissue cultures of human or animal cells.
Cell-Based Potency Assay
Potency tests are essential for guaranteeing the product's consistency and quality. Cell-based potency assays quantitatively determine the biological activity of a biopharmaceutical and biosimilars by comparing the biological response/activity, related to its mode of action, with a control/reference preparation (USP, WHO or in-house reference standard). Our system depends on in-house cell-based potency assays employing several cell lines.
Determination of IC50 and Toxicity Testing
IC (inhibitory concentration) curves are used to determine the concentration of medication needed to reduce the number of viable cells by a predetermined amount. The term "IC50" specifically refers to the concentration at which 50% inhibition of a biological process occurs. IC curves can show the shift in population due to changes in cell growth or mortality. In cancer treatment, if a medication's IC50 concentration is lower, it can be effective at lower doses, reducing systemic toxicity and minimizing damage to healthy cells while targeting cancer cells. Using IC50 concentration allows for potentially eliminating cancer cells with minimal impact on healthy cells. Overall, IC curves and IC50 are essential tools in determining effective medication concentrations for desired outcomes in cell-based experiments and cancer treatment.
Protein Extraction from Different Cell Lines, Gel Electrophoresis and Western Blotting
Protein extraction from cultured cells is the first step in many biochemical and analytical processes (PAGE, Western blotting, mass spectrometry, etc.) . Cultured cells must be effectively disrupted and homogenized in order to provide high protein outputs. Next, gel electrophoresis is used for the separation and analysis of biomacromolecules and their fragments based on their size and charge. Finally, with western blot, researchers can identify specific proteins from a complex mixture of proteins extracted from cells.
RNA Extraction from Different Cell Lines and Gene Expression Analysis Techniques
Total undegraded cellular RNA, primarily free of contaminating DNA, is retrieved from cultured cells. The most advanced technologies are then used to conduct gene expression assays to determine how genes are translated into useful gene products.
Cell-Based ELISA Assays
Cell-based ELISA assays are used in immunogenicity studies and as a screening method. It is used to measure oxidative stress, lipid, inflammatory and apoptosis markers.
Method Validation and Verification of Cell-Based Assays
Method validation and verification are processes that involve carrying out several evaluations to ensure that an analytical test system is appropriate for its intended use and capable of producing useful and reliable analytical results. At our unit, we routinely work with our customers to make sure the assay is optimal for its intended use based on the established assay.
Services
Molecular Biology Unit
PCR, Real-time PCR (including up to six-plex), protein gel electrophoresis, and Western blotting.
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