| Description | The Cell Cycle and Apoptosis Kit (Cell Cycle and Apoptosis Kit) is a test kit that uses the classic propidium iodide (PI) staining method to analyze the cell cycle and apoptosis. Propidium iodide is a fluorescent dye for double-stranded DNA. When propidium iodide binds to double-stranded DNA, The Cell Cycle and Apoptosis Kit (Cell Cycle and Apoptosis Kit) is a test kit that uses the classic propidium iodide (PI) staining method to analyze the cell cycle and apoptosis. Propidium iodide is a fluorescent dye for double-stranded DNA. When propidium iodide binds to double-stranded DNA, it can produce fluorescence, and the fluorescence intensity is proportional to the content of double-stranded DNA. After the DNA in the cells is stained with propidium iodide, the DNA content of the cells can be measured using a flow cytometer, and then based on the distribution of DNA content, the cell cycle and apoptosis can be analyzed. After staining with propidium iodide, if the fluorescence intensity of G0/G1 phase cells is 1, then the theoretical fluorescence intensity of G2/M phase cells containing double genomic DNA is 2, and the fluorescence intensity of S phase cells undergoing DNA replication is between 1 and 2. Apoptotic cells, due to the condensation of the cell nucleus and the fragmentation of DNA (DNA fragmentation), cause some genomic DNA fragments to be lost during staining, so the iodine staining of apoptotic cells presents a significantly weak color, that is, the fluorescence intensity is less than 1, and a so-called sub-G1 peak appears on the fluorescence graph of flow cytometry, which is the peak of apoptotic cells. When cells undergo apoptosis, due to the concentration of cytoplasm and chromatin, apoptotic bodies are produced, causing changes in the light scattering properties of the cells. In the early stage of cell apoptosis, the ability of the cell to forward-angle light scattering significantly decreases, while the ability of lateral light scattering increases or remains unchanged. In the late stage of cell apoptosis, the signals of forward and lateral light scattering both decrease, so the changes in cell light scattering can be measured using a flow cytometer to observe the apoptosis situation. This kit is usually applied to the detection of the cell cycle and apoptosis of cultured adherent or suspended cells. If used for the cell cycle and apoptosis detection of cells from tissues, the tissue must be digested into a single-cell state before detection can be performed. P1373478Component20 T50 T100 TStorageQuantity Per TestP1373478ADyeing buffer solution10 mL25 mL50 mL-20℃.500 µL per 1 × 10⁶ cells.P1373478BPropidium Iodide staining solution (20X)100 µL250 µL500 µL-20℃.Store in the dark.5 µL per 1 × 10⁶ cells.P1373478CRNase A (50X)0.2 mL0.5 mL1 mL-20℃.10 µL per 1 × 10⁶ cells.Note: The recommended number of cells to stain per test is 1 × 10⁶ cellsInstructions for use1.Collect cells and wash twice with ice-cold PBS; resuspend in ice-cold PBS at a density of 0.1–1.0 × 10⁷ cells/mL.2.In the cell suspension, anhydrous ethanol was added dropwise while shaking until the final ethanol concentration reached 70%.3.Mix gently and fix overnight at 4 °C.4.Wash cells twice with ice-cold PBS, then resuspend in dyeing buffer solution of 2 × 10⁶ cells/mL.5.Add 5 µL of Propidium iodide and 10 µL RNase A to 500 µL of the cell suspension.6.Resuspend cells gently and thoroughly, then incubate at room temperature for 30 min in the dark.7.Analyze samples by flow cytometry.Note: Please bring your own PBS and absolute ethanol.Precautions 1. Fluorescent dyes all have the problem of quenching. Please try to avoid light during storage and use to slow down fluorescence quenching. 2. Iodophor is irritating to the human body. Please take appropriate protective measures. 3. Iodophor is a known mutagen, so this solution needs to be treated with activated carbon before being discarded. 4. When fixing cells with 70% ethanol, make sure it is thorough; otherwise, uneven staining will result in unclear or inaccurate results. It is recommended to fix the cells overnight at -4℃ with 70% ethanol. 5. After cell fixation, ensure that the cells are in a single-cell suspension; cell adhesion may affect our results. 6. For your safety and health, please wear laboratory coats and disposable gloves during operation. 7. This product is only for scientific research and cannot be used for clinical diagnosis or treatment, nor for food or medicine. It must not be stored in ordinary residences. 8. For your safety and health, please wear laboratory coats and disposable gloves during operation... Read More | The content of this cell is too long for an XLSX file (more than 32767 characters). Please use the CSV format for this export | Product content： Component O66550510 preps O665505 50 preps Blocking Buffer 100 ml 500 ml Antibody Pretreat Solution（ HRP/Mouse ） 1 ml 5 × 1 ml Dilution Buffer 100 ml 500 ml Wash Buffer（ 10× ) 100 ml 500 mlProduct Introduction:The one-step rapid WB assay kit (Product content： Component O66550510 preps O665505 50 preps Blocking Buffer 100 ml 500 ml Antibody Pretreat Solution（ HRP/Mouse ） 1 ml 5 × 1 ml Dilution Buffer 100 ml 500 ml Wash Buffer（ 10× ) 100 ml 500 mlProduct Introduction:The one-step rapid WB assay kit (mouse) is the latest Western Blot detection kit developed by Kangwei Century, which can obtain high-quality Western Blot results in about 1 hour. It is easy to operate, has high detection sensitivity, low background, does not require the addition of secondary antibodies, and has strong system stability. The conventional Western Blot indirect detection process (blocking, primary antibody binding, and secondary antibody binding) requires a long time, a complex experimental process, and requires multi-step condition optimization. After transferring the protein on the gel to the carrier membrane, incubate it with the blocking solution in the reagent kit for 5 minutes, and then incubate the carrier membrane with the primary antibody treated with antibody reaction solution. After washing three times (5 minutes each time), luminescence or colorimetric detection can be performed. This reagent kit is designed for use in experimental systems where the target protein primary antibody is derived from mice.Notes:1. The customer prepares their own mouse source primary antibody.2. Before using Blocking Buffer blocking solution, Antibody Pretreat Solution (HRP/Mouse) antibody reaction solution (mouse), and Wash Buffer (10 x) rinse solution, please mix thoroughly.3. If there is precipitation in the rinsing solution when stored at 2-8 ℃, please restore it to room temperature, dissolve the precipitation, and use it normally. The 1x rinsing solution can be stored at room temperature for one month.4. It is recommended to stain the membrane with reagents such as spring red after the transfer is completed, and cut off any excess parts on the membrane to increase the efficiency of the reagents.5. The optimal dilution amount for primary antibody and antibody reaction solution HRP (mouse) needs to be determined through preliminary experiments.6. The antibody reaction solution HRP (mouse), antibody dilution solution, and antibody dosage can be increased or decreased proportionally according to the size of the membrane.7. The antibody dilution solution containing the first antibody can be recycled and reused once. It is recommended not to reuse antibodies with poor specificity and affinity. If the recovered antibody is used within 1-2 days and stored at 2-8 ℃ for long-term storage, please freeze it at -20 ℃ to avoid repeated freeze-thaw cycles.8. If there is a high background, please adjust the amount of antibodies and increase the number of times the film is washed.9. All reagents in the reagent kit should be stored at 2-8 ℃ to avoid freezing and thawing.Operation steps:This product is suitable for the sealing and antibody incubation steps after membrane transfer, taking a 5 cm x 8 cm membrane as an example:1. Preparation of rinsing solution: Dilute 10 ml of Wash Buffer (10 x) with distilled water to 100 ml, which is 1 x Wash Buffer. Set aside. Use 8-10 ml for each film wash.2. Sealing: After the membrane transfer is completed, immerse the membrane in 10 ml Blocking Buffer and seal at room temperature for 5 minutes.3. Rinse: Pour off the sealing solution, add 8-10 ml of 1 x Wash Buffer, and rinse at a high speed on a shaker for 1 minute.4. Prepare antibody incubation solution while washing the membrane: Take Antibody Pretreat Solution (HRP/Mouse) 100 µ Add mouse derived primary antibody 3-10 into the centrifuge tube µ g. Suck and beat the gun head until thoroughly mixed, and incubate at room temperature for 5 minutes. Add to 10 ml Dilution Buffer and mix well. Note: 1) The dosage of primary antibody can also be adjusted according to the dilution of the antibody. Taking the final dilution of antibodies at 1:1000 as an example, take 100 µ Add HRP (mouse) antibody reaction solution into the EP tube and add 10 µ Add the first antibody to 10 ml of antibody diluent, mix well, and incubate at room temperature for 5 minutes. 2) If the membrane area is small, the amount of antibodies, reaction solution, and diluent can be reduced proportionally.5. After completing step 3, pour out the rinsing solution and add the antibody incubation solution mixed with primary antibody, Antibody Pretreat Solution (HRP/Mouse), and Dilution Buffer to the membrane (ensuring that the incubation solution completely submerges the surface of the membrane). Incubate at room temperature on a shaker at around 60 rpm for 40 minutes.6. Discard (recover) the antibody incubation solution and rinse 3-5 times with the prepared 1 x Wash Buffer, each time for 3 minutes. 7. Conduct subsequent testing. It is recommended to use ECL or DAB methods for testing.Application examples:Example 1 Antigen is 293T cell lysateA: Normal WB control: beta actin mouse monoclonal antibody (CW0096) 5 µ Incubate at room temperature for 40 minutes, wash the film and dilute the secondary antibody sheep anti mouse HRP (CW0102) 1:10000. Incubate at room temperature for 40 minutes and expose ECL (CW0049).B: One step method WB: beta actin mouse monoclonal antibody (CW0096) 5 µ Incubate at room temperature for 40 minutes and expose ECL (CW0049).Example 2 Antigen is E. coli multi label protein lysateC: Normal WB control: GST mouse monoclonal antibody (CW0084) 2.5 µ Incubate at room temperature for 40 minutes, wash the film and dilute the secondary antibody sheep anti mouse HRP (CW0102) 1:10000. Incubate at room temperature for 40 minutes and expose ECL (CW0049).D: One step method WB: GST mouse monoclonal antibody (CW0084) was incubated at room temperature with 2.5ug for 40 minutes, and ECL (CW0049) was exposed... Read More | Products content Products IntroductionThis kit is a dedicated sample preparation solution for microbiome analysis and is suitable for the purification and enrichment of genomic DNA of pathogenic microorganisms such as bacteria and fungi from mixed samples such as swabs, blood, sputum, alveolar Products content Products IntroductionThis kit is a dedicated sample preparation solution for microbiome analysis and is suitable for the purification and enrichment of genomic DNA of pathogenic microorganisms such as bacteria and fungi from mixed samples such as swabs, blood, sputum, alveolar lavage, etc. During the purification process, differential lysis of the host cells and subsequent enzymatic digestion can effectively remove most of the host DNA while providing a comprehensive coverage of the bacterial and fungal DNA loci to a higher level. By differential lysis of host cells and subsequent enzymatic digestion, this kit can effectively remove most of the host DNA while maximizing the full coverage of bacterial, fungal and other pathogenic microbial DNA sites, thus obtaining microbiome DNA enrichment products with a higher coverage. Microbial DNA purified with this kit is suitable for a variety of downstream applications, including whole genome sequencing analysis, 16S rDNA-based high sensitivity microbiome analysis, and macrogenomic birdshot sequencing analysis. Self-contained reagents and consumablesSterile pipette tips with aerosol barrier to prevent cross-contamination anhydrous ethanol Microcentrifuge tubes (2 ml/1.5 ml) PBS buffer (required for some samples only)Pre-experiment Preparation and Important Notes1. Add 1.25 ml Proteinase K Storage Buffer to Proteinase K and store at -20℃. Do not leave the prepared Proteinase K (20 mg/ml) at room temperature for a long time, and avoid repeated freezing and thawing to avoid affecting its activity.2. Dissolve Lysozyme (100 mg) in 10 ml Enzymatic Lysis Buffer to a final concentration of 10 mg/ml, dispense into sterile tubes and store at -20℃. Do not leave the prepared Lysozyme (10 mg/ml) at room temperature for a long time and avoid repeated freezing and thawing to avoid affecting its activity.3. Thaw Buffer GB1 and Buffer GB2 at room temperature or 2-8°C before use and mix thoroughly. Thawed Buffer GB1 and Buffer GB2 can be left at 2-8°C for 1-2 weeks without affecting their activity, and should be stored at -20°C for long term storage. To ensure optimal performance, do not freeze or thaw more than three times. If less than one bottle of Buffer GB1 and Buffer GB2 is required for a single extraction, ensure that it is used under sterile conditions such as an ultra-clean bench and avoid microbial contamination and growth in the remaining buffer.4. Before first use, anhydrous ethanol should be added to Buffer GW1 and Buffer GW2 according to the instructions on the vial label and labeled.5. Check Buffer GL for crystallization or precipitation before use, and if crystallization or precipitation occurs, redissolve Buffer GL in a 56°C water bath.6. If the downstream experiments are sensitive to RNA contamination, 4 µl of DNase-Free RNase A (100 mg/ml) can be added before adding Buffer GL. RNase A is not provided in the kit, but can be ordered separately from CW0601S.7. This kit is designed for the isolation of DNA from intact microbial cells. To ensure optimal recovery of microbial DNA, samples should be fresh. If storage or transportation is required, this should preferably be done at 2-8°C and not frozen or thawed, as freezing and thawing can damage the integrity of the microbial cells and therefore result in the loss of exposed microbial DNA during host DNA removal.8. To avoid false results due to contamination, keep the work area clean, wear protective clothing, and set up controls for quality control. Use appropriate measures to handle sample materials to minimize the risk of cross-contamination. During the extraction process, use DNA-free pipette tips and consumables, and cap reagents immediately after use to prevent contamination. procedure1. Sample pre-treatment: 1a: For swab samples, swirl the swab portion of the swab in 0.5 ml PBS for at least 20 s. Squeeze the swab several times against the wall of the tube before removing it so that as much of the bacterial fluid as possible can be squeezed out of the swab to minimize sample loss. 1b: For viscous samples, e.g. sputum, take ~500 µl of sample, add 1.5 times the volume (~750 µl) of Buffer GB1 and incubate at 37°C, 600 rpm for 15-30 min until the sample is completely liquefied.Note: The sample volume can be increased or decreased appropriately and the amount of Buffer GB1 added adjusted accordingly.1c: For alveolar lavage fluid containing a small amount of viscous sputum, centrifuge as much of the alveolar lavage fluid as possible, carefully remove the supernatant, and retain the lower viscous fraction (containing sputum, cells, and organisms), add 1.5 times the volume of Buffer GB1, and incubate for 15-30 min at 37°C, 600 rpm until the sample is completely liquefied.1d: For non-viscous body fluid samples such as blood and cerebrospinal fluid, liquefaction treatment is not required, and an appropriate amount of sample is taken directly, the operation of step 2 is carried out, and the cell precipitate is collected by centrifugation.2. Centrifuge at 10000 rpm for 5-10 min at room temperature and carefully discard the supernatant.Note: Do not disturb the lower cell sediment to avoid sample loss.3. Add 500 µl Buffer GB2, vortex to mix, and incubate at room temperature, 600 rpm for 10 min. 4. Centrifuge at 12000 rpm for 2 min and carefully remove the supernatant.Note: Do not disturb the bacterial precipitate when removing the supernatant to avoid sample loss.5. Add 200 µl of Buffer GB2 to the precipitate, add 2 µl of Benzonase and incubate for 30 min at 37°C, 600 rpm. 6. Centrifuge at 12000 rpm for 2 min, discard the supernatant, add 500 µl of Buffer GB2, vortex and wash the precipitate. Repeat the procedure once.7. Centrifuge at 12000 rpm for 2 min, discard the supernatant, and finally aspirate the residual Buffer GB2 with a small-volume tip. 8. Add 180 µl Lysozyme (10 mg/ml), resuspend the bacterial precipitate and transfer the bacterial resuspension to a Lysis Tube.9. The Lysis Tube is incubated at 37°C, 600 rpm for 20-30 min, then vortexed for 10 min or processed on a thermostatic homogenizer for 10 min at maximum vibration speed (2500-2900 rpm).10. Centrifuge briefly, add 20 µl proteinase K, vortex to mix, add 200 µl buffer GL, vortex to mix, and incubate for 30 min at 56°C, 600 rpm. Note: 1) Do not add Proteinase K directly to Buffer GL.2）For RNA removal, add 4 µl DNase-Free RNase A (100 mg/ml) before adding Buffer GL, shake to mix, and let stand at room temperature for 5-10 minutes.11. Centrifuge at 12000 rpm for 1 min and carefully aspirate the supernatant into a new centrifuge tube. Note: Do not aspirate the glass beads.12. Add 200 µl of anhydrous ethanol, vortex to mix, and centrifuge momentarily to collect the solution to the bottom of the tube. Note: The addition of anhydrous ethanol may produce a white precipitate that will not affect subsequent experiments.13. Add all of the solution from step 12, including the precipitate, to the Spin Columns DM in the collection tube, or transfer the solution several times if it cannot be added all at once. centrifuge at 12,000 rpm for 1 minute, pour off the waste from the collection tube, and return the column to the collection tube.14. Add 500 µl Buffer GW1 to the adsorbent column (check that anhydrous ethanol has been added before use), centrifuge at 12,000 rpm for 1 min, pour off the waste liquid from the collection tube, and put the adsorbent column back into the collection tube.15. Add 500 µl Buffer GW2 to the adsorbent column (check that anhydrous ethanol has been added before use), centrifuge at 12,000 rpm for 1 minute, pour off the waste liquid in the collection tube, and put the adsorbent column back into the collection tube. Note: Step 15 can be repeated once if further improvement of DNA purity is required.16. Centrifuge at 12,000 rpm for 2 minutes and pour off the waste liquid in the collection tube. Leave the column at room temperature for a few minutes and dry thoroughly. Note: The purpose of this step is to remove residual ethanol from the adsorbent column; ethanol residue can interfere with subsequent enzymatic reactions (digestion, PCR, etc.).17. Place the adsorbent column in a new centrifuge tube (supplied), add 50 µl of Buffer GE to the center of the adsorbent column overhang, let stand at room temperature for 5 minutes, centrifuge at 12,000 rpm for 1 minute, collect the DNA solution, and store the DNA at -20 °C. Attention:1）If the downstream experiments are sensitive to pH or EDTA, sterilized water can be used for elution. The pH value of the eluent has a great influence on the elution efficiency. If the eluent is made of water, the pH value should be 7.0-8.5 (the pH value of water can be adjusted to this range with NaOH), and the elution efficiency is not high when the pH value is lower than 7.0.2）Incubation at room temperature for 5 minutes prior to centrifugation increases yield.3）If the final concentration of DNA is to be increased, the DNA eluate obtained in step 17 can be re-spiked onto the adsorbent membrane and step 17 repeated. 4）DNA stored in water will be affected by acidic hydrolysis. For long-term storage, it is recommended to elute with Buffer GE and store at -20℃... Read More | Inquire |