| Description | Cell proliferation assays are widely used in the evaluation of cell viability, genotoxicity, and the efficacy of antitumor drugs. Direct detection of DNA synthesis in cells is considered the most accurate method for assessing cell proliferation. EdU (5-ethynyl-2′-deoxyuridine) is a Cell proliferation assays are widely used in the evaluation of cell viability, genotoxicity, and the efficacy of antitumor drugs. Direct detection of DNA synthesis in cells is considered the most accurate method for assessing cell proliferation. EdU (5-ethynyl-2′-deoxyuridine) is a novel thymidine (thymine deoxyribonucleoside) analogue. During DNA synthesis, EdU can be incorporated into newly synthesized DNA in place of thymidine. The ethynyl group on EdU can undergo a covalent reaction with fluorescently labeled small-molecule azide probes (such as Azide Alexa Fluor 488, Azide Alexa Fluor 555, Azide Alexa Fluor 594, Azide Alexa Fluor 647, etc.) via Cu(I)-catalyzed click chemistry, forming a stable triazole ring. This reaction is highly efficient and is referred to as the Click reaction. Through this process, newly synthesized DNA is labeled with the corresponding fluorescent probes, enabling the detection of proliferating cells using appropriate fluorescence detection equipment.Product Component TableE1456506Component50 T100 T200 TStorage conditionsQuantity Per TestE1456506AEdU(10 mM)100µL200µL400µL-20℃.Store in the dark.2 µL per 1.0-2.0 × 10⁶ cellsE1456506B6-FAM125µL250µL500µL-20℃.Store in the dark.2.5 µL per 1.0-2.0 × 10⁶ cellsE1456506CClick Reaction Buffer13 mL26 mL52 mL-20℃.Store in the dark.237.5 µL per 1.0-2.0 × 10⁶ cellsE1456506DCuSO40.5 mL1 mL2 mL-20℃.10 µL per 1.0-2.0 × 10⁶ cellsE1456506EClick Additive248 mg496 mg992 mg-20℃.Store in the dark.250 µL per 1.0-2.0 × 10⁶ cellsE1456506FDAPI Staining Solution（1000×）25 µL50 µL100µL-20℃.Store in the dark.0.5 µL per 1.0-2.0 × 10⁶ cells Usage Protocol1. Preparation1) Preparation of Click Additive Solution: For a 50-test kit: Add 12.5 mL of pre-chilled deionized water to the tube. Mix thoroughly until completely dissolved to obtain the Click Additive Solution. For a 100-test kit: Add 25 mL of pre-chilled deionized water to the tube. Mix thoroughly until completely dissolved to obtain the Click Additive Solution. For a 200-test kit: Add 50 mL of pre-chilled deionized water to the tube. Mix thoroughly until completely dissolved to obtain the Click Additive Solution. After preparation, aliquot the solution as needed and store at -20°C. If a white precipitate forms after dissolution, invert the tube repeatedly until it is fully dissolved before use. If the solution turns brown, it indicates degradation of the active component; discard it.2) Upon initial dissolution of the Click Reaction Buffer, aliquot it according to the number of samples per experiment and store at -20°C.2. EdU Labeling of CellsIt is recommended to use a final EdU concentration of 10 µM (1×). A 1:500 dilution of EdU (10 mM) in cell culture medium yields a 2× EdU working solution (20 µM). Mix an equal volume of pre-warmed (37°C) 2× EdU working solution (20 µM) with the cell suspension to achieve a final 1× EdU concentration. Incubate in a 37°C, 5% CO₂ incubator. Factors such as cell culture medium, cell density, cell type, and other experimental conditions may affect labeling efficiency. Therefore, the optimal EdU concentration and labeling duration must be empirically determined based on the cell type under investigation.3. Fixation and Permeabilization1) Harvest cells and centrifuge at 300 ×g for 5 min. Wash cells twice with PBS containing 2% FBS.2) Fix cells with 4% paraformaldehyde solution. Mix thoroughly and incubate for 15 min at room temperature protected from light.3) Collect cells and centrifuge at 300 × g for 5 min. Wash cells twice.4) Resuspend cells in PBS containing 0.1% Triton X-100. Mix well and incubate for 15 min at room temperature.5) Centrifuge at 300 × g for 5 min and wash cells twice.4. Fluorescent Labeling1) This protocol is based on a 500 µL reaction system per 2 × 10⁶ cells. The volume of the Click reaction mixture can be adjusted according to the experimental sample size.2) Centrifuge the cells at 300 ×g for 5 minutes. Add 500 µL of Click reaction mixture per sample, mix gently, and incubate for 30 minutes at room temperature protected from light.3) After the reaction, wash the cells twice with PBS containing 2% FBS.4) Dilute the DAPI Staining Solution (1000×) to 1× using PBS containing 2% FBS. Add 250 µL of the diluted DAPI solution to each sample and incubate for 5 minutes at room temperature.5) Add an additional 250 µL of PBS containing 2% FBS, mix gently, and proceed to detection using an appropriate flow cytometry instrument.Precautions1. Strictly adhere to the component order and volumes specified in the table above when preparing the Click reaction mixture, as deviations may affect subsequent experimental results.2. The Click reaction mixture must be used within 15 minutes of preparation.3. To avoid fluorescence quenching, perform detection as soon as possible after sample preparation... 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:D665967Component200 TStorageD665967ABuffer PB120 mLRTD665967BBuffer PS60 mLRTD665967CBuffer PW (concentrate)25 mLRTD665967DBuffer EB30 mLRTD665967ESpin Columns DM with Collection Tubes200 EART Product Introduction: This reagent kit adopts a new silicon-based membrane technology and Product content:D665967Component200 TStorageD665967ABuffer PB120 mLRTD665967BBuffer PS60 mLRTD665967CBuffer PW (concentrate)25 mLRTD665967DBuffer EB30 mLRTD665967ESpin Columns DM with Collection Tubes200 EART Product Introduction: This reagent kit adopts a new silicon-based membrane technology and reagent formula. Through a rapid and simple three-step process of binding, washing, and elution, 100 bp-10 kb DNA fragments can be purified and recovered from PCR products or enzyme reaction solutions (enzyme cutting, linking, probe labeling, etc.). Each adsorption column can adsorb up to 10 kb of DNA fragments µ G DNA, while minimizing impurities such as primers, oligonucleotides, enzymes, etc. The purified and recovered DNA has high purity and concentration, good integrity, and high recovery rate, and can be directly used for molecular biology experiments such as sequencing, linking and transformation, labeling, and in vitro transcription.Self prepared reagent: anhydrous ethanol.Preparation and important precautions before the experiment:1. All components can be stably stored in a dry, room temperature (15-30 ℃) environment for 1 year, and can be stored at 2-8 ℃ for longer periods of time. When the solution is stored at low temperature, it should be left at room temperature for a period of time before use, and then restored to room temperature before use.2. This reagent kit can selectively recover all DNA fragments from the solution. If you need to selectively recover specific fragments while removing other fragments of different sizes, please choose our company's gel recovery reagent kit.3.Before the first use, anhydrous ethanol should be added to the Buffer PW according to the instructions on the reagent bottle label.4. Before use, please check if there is any crystallization or precipitation in the Buffer PB. If there is any crystallization or precipitation, you can take a water bath at 37 ℃ for a few minutes to restore clarity.5. The recovery efficiency is related to the initial amount of DNA and the elution volume. The smaller the initial amount, the smaller the elution volume, and the lower the recovery rate.6. All centrifugation steps can be performed at room temperature.Operation steps:1. Estimate the volume of DNA reaction solution, add 5 times the volume of Buffer PB, and mix thoroughly (without removing paraffin or mineral oil).Note: 1) If the DNA reaction system is 50 µ l (excluding paraffin oil volume), add 250 µ l Buffer PB.2) After adding Buffer PB, check the pH value of the solution. If the pH value is greater than 7.5, add 10-30 to it µ 3 M sodium acetate (pH 5.0) was used to adjust the pH value to 5-7.2. Column balance: Add 200 to the spin columns DM that have been loaded into the collection tube µ Centrifuge at 13000 rpm (~16200 × g) for 1 minute, discard the waste liquid in the collection tube, and place the adsorption column back into the collection tube.3. Add the solution obtained in step 1 to the adsorption column that has been loaded into the collection tube, let it stand at room temperature for 1 minute, centrifuge at 13000 rpm for 30-60 seconds, discard the waste liquid in the collection tube, and place the adsorption column in the collection tube.Attention: The volume of the adsorption column is 750 µ l. If the sample volume is greater than 750 µ l, it can be added in batches.4. Add 500 µ l of Buffer PW to the adsorption column (please check if anhydrous ethanol has been added before use), centrifuge at 13000 rpm for 30-60 seconds, discard the waste liquid in the collection tube, and place the adsorption column in the recovery tube.Note: If purified DNA is used for salt sensitivity experiments (such as flat end ligation experiments or direct sequencing), it is recommended to add Buffer PW and let it stand for 2-5 minutes before centrifugation.5.13000 rpm for 1 minute and discard the waste liquid from the collection tube. Place the adsorption column at room temperature for a few minutes to thoroughly air dry.Note: The purpose of this step is to remove residual ethanol from the adsorption column, which can affect subsequent enzymatic reactions (such as enzyme digestion, PCR, etc.). To ensure that downstream experiments are not affected by residual ethanol, it is recommended to open the cover of the adsorption column and place it at room temperature for a few minutes to thoroughly dry the residual ethanol in the adsorbent material at the bottom.6. Place the adsorption column into a new centrifuge tube (provided by oneself), add 30-50 µ l Buffer EB to the middle position of the adsorption membrane by hanging droplets, and let it stand at room temperature for 1 minute. Centrifuge at 13000 rpm for 1 minute and collect DNA solution- Store DNA at 20 ℃.Attention:1) The pH value of the eluent has a significant impact on the elution efficiency. If water is used as the eluent, its pH value should be ensured to be between 7.0-8.5 (the pH value of water can be adjusted to this range using NaOH).2) To improve the recovery of DNA, the solution obtained by centrifugation can be added back to the adsorption column, left at room temperature for 2 minutes, and centrifuged at 13000 rpm for 1 minute.3) The elution volume should not be less than 30 µ l. A small volume will affect the recovery efficiency... Read More | This kit is used to extract and purify high-quality total RNA from various plants, and is also suitable for the extraction of fungal hyphal RNA. A unique Shredder separation column is used for homogenization and filtration of high viscosity plant or fungal lysates, while silica based membrane is This kit is used to extract and purify high-quality total RNA from various plants, and is also suitable for the extraction of fungal hyphal RNA. A unique Shredder separation column is used for homogenization and filtration of high viscosity plant or fungal lysates, while silica based membrane is used to adsorb RNA for purification, effectively removing various pollutants such as polysaccharides through washing. The washed RNA can be directly used in various downstream experiments. RNA with a molecular weight greater than 200 bases was extracted using this reagent kit, with high purity and almost no DNA residue. If it is an RNA experiment that is very sensitive to trace amounts of DNA, the remaining DNA can be digested and removed on a column using DNase I without RNase. The extracted RNA can be used for experiments such as Northern Blot, Dot Blot, RT-PCR, and in vitro translation. R665489Component50 TStorageR665489ABuffer RL35 mLRTR665489BBuffer RLC35 mLRTR665489CBuffer RW140 mLRTR665489DBuffer RW2 (concentrate)11 mLRTR665489ERNase-Free Water10 mLRTR665489FSpin Columns FL with Collection Tubes50 setsRTR665489GSpin Columns RM with Collection Tubes50 setsRTR665489HRNase-Free Centrifuge Tubes (1.5 mL)50 EART Self prepared reagents:β- Mercaptoethanol, anhydrous ethanol (newly opened or dedicated for RNA extraction).Preparation and important precautions before the experiment:To prevent RNase pollution, attention should be paid to the following aspects:1) Use RNase free plastic products and gun heads to avoid cross contamination.2) Glassware should be dry baked at a high temperature of 180 ℃ for 4 hours before use, while plastic containers can be soaked in 0.5 M NaOH for 10 minutes, thoroughly rinsed with water, and then sterilized under high pressure.3) Prepare the solution using water without RNase.4) Operators should wear disposable masks and gloves, and change gloves frequently during the experiment.2. The extracted samples should avoid repeated freeze-thaw cycles, otherwise it will affect the quantity and quality of RNA extraction.3. Please add Buffer RL before use β- Mercaptoethanol, with a final concentration of 1%. Add 10 to 1 ml Buffer RL µ L β Mercaptoethanol. join β- The buffer RL room temperature of mercaptoethanol can be stored for one month. No need to add buffer RLC when using it β- Mercaptoethanol.Before the first use, anhydrous ethanol should be added to Buffer RW2 according to the instructions on the reagent bottle label.5. If precipitation occurs in Buffer RL and Buffer RLC, please heat them to dissolve and place them at room temperature.6. All centrifugation steps should be carried out at room temperature unless otherwise specified, and all operation steps should be carried out quickly.7. If downstream experiments are highly sensitive to DNA, it is recommended to treat RNA with DNase I without RNase.Operation steps:1. Take 50-100 mg of fresh plant tissue, add liquid nitrogen and quickly grind it into powder.2. Collect the ground powder into a centrifuge tube (provided by oneself) and add 600 µ L Buffer RL (check if it is added before use) β- Sulfhydryl ethanol or Buffer RLC, vortex oscillation causes it to fully decompose.Attention:1) The main component of Buffer RL is guanidine isothiocyanate, which is suitable for the lysis of most plant tissues. However, in some plant tissues (such as corn endosperm), due to the unique secondary metabolites, guanidine isothiocyanate causes precipitation in the sample, resulting in poor RNA extraction efficiency. In this case, Buffer RLC can be added instead of Buffer RL.2) Incubating at 56 ℃ for 1-3 minutes helps with tissue lysis, but plants with high starch content should not be subjected to high-temperature incubation.3. Transfer all the liquid obtained in step 2 to the spin columns FL that have been loaded into the collection tube, centrifuge at 12000 rpm (~13400 × g) for 2 minutes, and transfer the supernatant from the collection tube to a new centrifuge tube (provided by oneself).Attention:1) When aspirating liquid, the tip of the gun can be cut off for easy sampling.2) Spin Columns FL can remove most of the fragments, but there will still be a small amount flowing out. After centrifugation, precipitation will form in the collection tube. When proceeding to the next step, be careful not to absorb the sediment.4. Add 0.5 times the volume of anhydrous ethanol to the clean cracking solution obtained in step 3 and quickly mix well. Attention: Adding ethanol may cause precipitation, but it does not affect subsequent experiments.5. Add all the solutions obtained in step 4 to the spin columns RM that have been loaded into the collection tube. If it is not possible to add all the solutions to the adsorption column at once, please transfer them in two separate steps. Centrifuge at 12000 rpm for 15 seconds, discard the waste liquid in the collection tube, and place the adsorption column back into the collection tube.6. Add 700 to the adsorption column µ Centrifuge at 12000 rpm for 1 minute, discard the waste liquid from the collection tube, and place the adsorption column back into the collection tube. Optional steps: If conducting RNA experiments that are highly sensitive to trace amounts of DNA, replace step 6 with the following steps.1) Add 350 to the adsorption column µ L Buffer RW1, centrifuge at 12000 rpm for 15 seconds, discard the waste liquid, and place the adsorption column back into the recovery manifold.2) Preparation of DNase I mixture: Take 52 µ Add 8 RNase Free Water to it µ 10 x Reaction Buffer and 20 µ DNase I (1 U/ µ l) Mix well and prepare to a final volume of 80 µ The reaction solution of L.Attention:The above system is configured according to our company's DNase I reaction system. Please refer to the corresponding instructions for other company products.3) Add 80 µ l of DNase I reaction solution directly to the adsorption column and incubate at 20-30 ℃ for 15 minutes.4) Add 350 to the adsorption column µ L Buffer RW1, centrifuge at 12000 rpm for 15 seconds, discard the waste liquid, and place the adsorption column back into the recovery manifold.7. Add 500 to the adsorption column µ Buffer RW2 (check if anhydrous ethanol is added before use), centrifuge at 12000 rpm for 15 seconds, discard the waste liquid in the collection tube, and place the adsorption column back into the collection tube.8. Repeat step 7.Centrifuge at 9.12000 rpm for 2 minutes and discard the waste liquid from the collection tube. Place the adsorption column at room temperature for a few minutes to thoroughly dry the anhydrous ethanol in the column.Attention:The purpose of this step is to remove residual ethanol from the adsorption column, which will affect subsequent enzymatic reactions (such as enzyme digestion, PCR, etc.).10. Place the adsorption column in a new RNase free centrifuge tube, and add 30-50 to the middle of the adsorption column in the air µ Place RNase Free Water at room temperature for 1 minute, centrifuge at 12000 rpm for 1 minute, collect RNA solution, and store RNA at -70 ℃ to prevent degradation.Attention:1) The volume of RNase Free Water should not be less than 30 µ l. Small volume affects the recovery rate.2) If you want to increase RNA production, you can use 30-50 µ Repeat step 10 for the new RNase Free Water.3) If you want to increase the RNA concentration, you can add the obtained solution back to the adsorption column and repeat step 10... Read More | DescriptionMetathesis: Ruthenium-Based Metathesis CatalystsRuthenium metathesis catalysts kit I consists of 9 samples of Grubbs 1st and 2nd generation catalysts. These catalysts have applications in ring-closing and ring-opening metathesis, cross-metathesis, ring-opening metathesis polymerization (DescriptionMetathesis: Ruthenium-Based Metathesis CatalystsRuthenium metathesis catalysts kit I consists of 9 samples of Grubbs 1st and 2nd generation catalysts. These catalysts have applications in ring-closing and ring-opening metathesis, cross-metathesis, ring-opening metathesis polymerization (ROMP) and enyne metathesis.Metathesis: Ruthenium-Based Metathesis Catalysts... Read More |