{"product_id":"ba00209","title":"MTT Assay Kit","description":"\u003ctable style=\"width:100%;border-collapse:collapse\"\u003e\u003ctbody\u003e\n\u003ctr\u003e\u003ctd colspan=\"2\" style=\"padding:12px\"\u003e\u003cstrong\u003eCat. No. BA00209\u003c\/strong\u003e\u003c\/td\u003e\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eSpecifications\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\u003cp\u003e500T\u003c\/p\u003e\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eApplications\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\u003cp\u003eCell growth, cytotoxicity and proliferation assays\u003c\/p\u003e\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eAssay Principle\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\u003cp\u003eMTT reduction to formazan; colorimetric microplate assay\u003c\/p\u003e\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eDetection\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\u003cp\u003eAbsorbance: 490 nm.\u003c\/p\u003e\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eStorage\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\u003cp\u003eShip at low temperature; store at -20 °C protected from light. Solution B may be shipped and stored at room temperature.\u003c\/p\u003e\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eShelf Life\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\u003cp\u003e1 year\u003c\/p\u003e\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eGeneral Information\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\u003cp\u003eMTT is widely used to assess cell growth. The underlying principle is that MTT can be reduced by dehydrogenases in the mitochondria of living cells to form dark-purple formazan crystals, whereas dead cells lack this activity. After the dark-purple formazan crystals are dissolved, their concentration can be determined by measuring the absorbance at 490 nm, and the viability of the cells can thereby be inferred: the more vigorous the cell proliferation, the higher the absorbance; the greater the cytotoxicity, the lower the absorbance.\u003c\/p\u003e\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eAdvantages\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\n\u003cp\u003e1. A unique formazan dissolving solution is used, which dissolves formazan completely and reduces errors.\u003c\/p\u003e\n\u003cp\u003e2. Low background, high sensitivity, wide linear range, and good reproducibility.\u003c\/p\u003e\n\u003cp\u003e3. This product is sufficient for 500 wells (five 96-well cell culture plates) of microplate assays.\u003c\/p\u003e\n\u003cp\u003e4. It can be used for bioactivity assays of biologically active factors, anti-tumor drug screening, cytotoxicity testing, determination of tumor radiosensitivity, etc.\u003c\/p\u003e\n\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eKit Components\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\u003ctable style=\"width:100%;border-collapse:collapse\"\u003e\u003ctbody\u003e\n\u003ctr\u003e\n\u003cth style=\"padding:8px;text-align:left;border:1px solid #d8d8d8\"\u003eComponent\u003c\/th\u003e\n\u003cth style=\"padding:8px;text-align:left;border:1px solid #d8d8d8\"\u003eSize\u003c\/th\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"padding:8px;text-align:left;border:1px solid #d8d8d8\"\u003eSolution A\u003c\/td\u003e\n\u003ctd style=\"padding:8px;text-align:left;border:1px solid #d8d8d8\"\u003e5 ml\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"padding:8px;text-align:left;border:1px solid #d8d8d8\"\u003eSolution B\u003c\/td\u003e\n\u003ctd style=\"padding:8px;text-align:left;border:1px solid #d8d8d8\"\u003e50 ml\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"padding:8px;text-align:left;border:1px solid #d8d8d8\"\u003eManual\u003c\/td\u003e\n\u003ctd style=\"padding:8px;text-align:left;border:1px solid #d8d8d8\"\u003e1 copy\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003c\/tbody\u003e\u003c\/table\u003e\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eProtocol\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\n\u003cp\u003eThe procedure below describes a cell cytotoxicity assay. Other applications are similar or even simpler (e.g., growth curve experiments); the procedure can be modified accordingly and will not be repeated here.\u003c\/p\u003e\n\u003cp\u003e(I) Cell seeding\u003c\/p\u003e\n\u003cp\u003e1. Digest a confluent monolayer of cells by conventional trypsin digestion and collect the cells into serum-containing culture medium.\u003c\/p\u003e\n\u003cp\u003e2. Centrifuge at 200 × g for 5 minutes to collect the cell pellet.\u003c\/p\u003e\n\u003cp\u003e3. Resuspend the cell pellet in culture medium to prepare a single-cell suspension and count the cells.\u003c\/p\u003e\n\u003cp\u003e4. Dilute the cells to 2500–10000 cells\/ml (to be decided according to the growth rate of the cells).\u003c\/p\u003e\n\u003cp\u003e5. Transfer a sufficient amount of the cell suspension to culture dishes (to facilitate sampling with a multichannel pipette). The MTT assay for one 96-well plate requires about 20 ml of cell suspension.\u003c\/p\u003e\n\u003cp\u003e6. Using a multichannel pipette, add 200 µL of cell suspension to the exact center of each well in columns 2–11 of the 96-well plate (for normal cells). For tumor cells, add 100 µL of tumor cell suspension and 100 µL of culture medium (total volume 200 µL). Note: the cells must be added to the exact center of the well; otherwise the cells will aggregate in the corners of the well and affect the assay.\u003c\/p\u003e\n\u003cp\u003e7. Using the multichannel pipette, add culture medium in the same volume as the cell suspension to the wells in columns 1 and 12. The wells in column 1 will serve as the +medium −cells +MTT control (used for zeroing when measuring OD), and the medium added to column 12 serves to reduce the edge effect on the reactions in column 11.\u003c\/p\u003e\n\u003cp\u003e8. Incubate at 37 °C and 5% CO₂ by conventional cell culture methods for 1–3 days so that the cells enter the exponential growth phase.\u003c\/p\u003e\n\u003cp\u003e(II) Drug treatment\u003c\/p\u003e\n\u003cp\u003e9. Dilute the drug in culture medium to 8 test concentrations (if the appropriate concentrations are unknown, a preliminary experiment is needed to determine them); generally, 3 parallel plates are required for one drug.\u003c\/p\u003e\n\u003cp\u003e10. Remove the culture medium from the wells in columns 2 to 11 (10 columns in total; do not disturb the cells) while keeping the medium in the wells of columns 1 and 12.\u003c\/p\u003e\n\u003cp\u003e11. Add 200 µL of fresh culture medium to the wells in columns 2 and 11 (2 columns in total); these wells will serve as the +medium +cells −drug control.\u003c\/p\u003e\n\u003cp\u003e12. Add the 8 concentrations of the test drug to the wells in columns 3 to 10 (8 columns in total), adding one concentration per column.\u003c\/p\u003e\n\u003cp\u003e13. Continue to incubate the 96-well plate at 37 °C and 5% CO₂ by conventional methods for a certain period of time; this period is the drug treatment time and is decided by the user.\u003c\/p\u003e\n\u003cp\u003e14. At the end of the treatment, remove the culture medium from the wells in columns 2 to 11 (10 columns in total, all containing cells) and add 100 µL of fresh culture medium.\u003c\/p\u003e\n\u003cp\u003e15. Change the medium daily so that the number of cells expands 2–3 fold (the time required varies with the cell type).\u003c\/p\u003e\n\u003cp\u003e(III) Viable cell counting\u003c\/p\u003e\n\u003cp\u003e16. At the end of the growth period, remove the culture medium from the wells in columns 1 to 11, then add 100 µL of fresh culture medium and 10 µL of Solution A (containing MTT). Wrap the 96-well plate in aluminum foil and continue culturing at 37 °C and 5% CO₂ for 4–8 hours. Note: Solution A solidifies at low temperatures; before use, allow it to stand at room temperature or warm it in a 20–25 °C water bath until completely dissolved, then mix well before use. MTT is carcinogenic — be sure to wear gloves.\u003c\/p\u003e\n\u003cp\u003e17. Carefully aspirate and discard the medium in the wells (containing Solution A). Since the medium may affect light absorption, it is best to remove it as completely as possible.\u003c\/p\u003e\n\u003cp\u003e18. Add 100 µL of Solution B to each well and shake on an orbital shaker at low speed for 10 minutes to fully dissolve the formazan crystals formed from MTT.\u003c\/p\u003e\n\u003cp\u003e19. Because the product is unstable, immediately measure the absorbance at 490 nm on a microplate reader. Note: use the wells in column 1 (+medium −cells +MTT control) to zero the instrument.\u003c\/p\u003e\n\u003cp\u003e20. Calculate the mean value of the replicates treated in the same way.\u003c\/p\u003e\n\u003cp\u003e21. Plot a curve with the drug concentration on the x-axis and the absorbance on the y-axis. Because the absolute absorbance values of the different treatments differ greatly, they are generally converted into the growth inhibition rate (using the mean of the data from the drug-free wells in columns 2 and 11 as 100%) so that the IC50 can be calculated for comparing the effects of different drug treatments. Note: if a growth curve is being measured, plot time on the x-axis. A normal growth curve is generally S-shaped, and growth-promoting treatments give a steeper slope.\u003c\/p\u003e\n\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eStorage Conditions\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\u003cp\u003eShip at low temperature and store at -20 °C (Solution B may also be shipped and stored at room temperature). Valid for one year.\u003c\/p\u003e\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003ctr\u003e\n\u003ctd style=\"width:24%;vertical-align:top;padding:12px\"\u003e\u003cstrong\u003eIntended Use\u003c\/strong\u003e\u003c\/td\u003e\n\u003ctd style=\"vertical-align:top;padding:12px\"\u003e\u003cp\u003eThis product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications.\u003c\/p\u003e\u003c\/td\u003e\n\u003c\/tr\u003e\n\u003c\/tbody\u003e\u003c\/table\u003e","brand":"Bioss","offers":[{"title":"500T","offer_id":68012803686699,"sku":"BA00209-500T","price":49.0,"currency_code":"USD","in_stock":true}],"url":"https:\/\/www.biossusa.com\/products\/ba00209","provider":"Bioss","version":"1.0","type":"link"}