| Description | Inquire | Ketone bodies, 3-hydroxybutyric acid (BOH) and acetoacetic acid (AcAc), are produced in the liver primarily from oxidation of fatty acids, and are normally present at low concentrations in urine and blood. Increased ketone concentrations in the blood may lead to metabolic acidosis, which has been Ketone bodies, 3-hydroxybutyric acid (BOH) and acetoacetic acid (AcAc), are produced in the liver primarily from oxidation of fatty acids, and are normally present at low concentrations in urine and blood. Increased ketone concentrations in the blood may lead to metabolic acidosis, which has been associated with diabetes, childhood hypoglycemia, growth hormone deficiency, alcohol or salicylate intoxication, and inborn errors of metabolism.Ketone Body Assay has been used to measure the release of ketone bodies in the human liver cancer cell line HepG2 culture medium... Read More | Product content N665859Component50 TStorageN665859ABuffer DS30 mLRTN665859BBuffer GTL15 mLRTN665859CBuffer GL15 mLRTN665859DBuffer GW1 (concentrate)13 mLRTN665859EBuffer GW2 (concentrate)15 mLRTN665859FBuffer TE10 mLRTN665859GProteinase K2×1.25 mLRTN665859HRNase A (100 mg/mL)0.4 Product content N665859Component50 TStorageN665859ABuffer DS30 mLRTN665859BBuffer GTL15 mLRTN665859CBuffer GL15 mLRTN665859DBuffer GW1 (concentrate)13 mLRTN665859EBuffer GW2 (concentrate)15 mLRTN665859FBuffer TE10 mLRTN665859GProteinase K2×1.25 mLRTN665859HRNase A (100 mg/mL)0.4 mLRTN665859ISpin Columns DF With Collection Tubes50 EA2-8℃N665859JCentrifuge Tubes (L-1.5 mL)50 EART Product IntroductionThis kit is suitable for the effective purification of genomic DNA from formalin-fixed, paraffin-embedded tissues.The product uses specially optimized dewaxing agent and lysis solution to release DNA from formalin-fixed or tissue sectioned samples, which does not involve the organic reagent xylene and does not need to be operated overnight; the digested samples are incubated at higher temperatures to remove formalin cross-linking of the free DNA, which can effectively improve the yield and purity of DNA; the optimized buffer system allows the inhibitors in the lysis solution to be specifically bound to the adsorbent membrane, which can be effectively removed by a two-step rinsing step. The optimized buffer system enables the DNA in the lysate to specifically bind to the adsorbent membrane, and the inhibitor is effectively removed by a two-step rinsing step, and finally eluted with low-salt buffer or water to obtain high-purity DNA.Meanwhile, configured with a high-efficiency microsorbent column, the elution volume can be as low as 20 µL.The purified DNA can be directly used for PCR, Real-time PCR, SNP Genotyping, STR genotyping, second-generation sequencing and pharmacogenomics research.The molecular weight of DNA isolated from formalin-fixed, paraffin-embedded samples is usually lower than that of DNA from fresh or frozen samples.The degree of DNA fragmentation depends on the type of sample, the duration of storage, and the conditions of fixation.Self-contained reagent: anhydrous ethanolPre-experiment Preparation and Important Notes1. After obtaining the sample, fix the sample in 4%-10% formalin as soon as possible, the fixation time should be 14-24 hours, too long a period of time will easily lead to genome breakage, affecting the downstream experiments. If the formaldehyde fixation time is too long or the sample has been stored for too long (> 1 year), it will easily lead to DNA integrity damage and unable to amplify long fragments.2. Ensure that the sample is thoroughly dehydrated before embedding; residual formalin will inhibit Proteinase K.3. Anhydrous ethanol should be added to Buffer GW1 and Buffer GW2 according to the instructions on the label of the reagent bottle before first use.4. Before use, please check Buffer GTL, Buffer GL and Buffer DS for any crystallization or precipitation. If there is any crystallization or precipitation, please re-dissolve Buffer GTL, Buffer GL and Buffer DS at 56℃ in a water bath.5. Preheat the water bath or thermostatic mixer to 56°C and keep the centrifuge at 25°C before starting the experiment.6. If downstream experiments are needed to reduce the low frequency of C>T:G>A transitions (artificial mutations) that occur to minimize the risk of false positives, 7 µL of UNG (1 U/uL) can be added after 1 hour of incubation at 90°C.Operation steps1. Sample processing:1a. Paraffin-embedded samples: Trim off excess paraffin from the tissue block with a scalpel to expose the tissue and then cut into 5-10µm slices. Take about 1×1cm2 slices (about 4-5 slices in total) and place them in a centrifuge tube (provided), add 160µL Buffer DS, vortex and shake for 10 seconds, then add 180µL Buffer GTL and 20µL Proteinase K, vortex and shake for 10 seconds. centrifuge the samples at 12,000rpm for 1 minute at 25℃.Note: 1) If the surface of the sample has been exposed to air, discard the 2-3 pieces that have been exposed to air and do not use them.2) DS will solidify below 18°C, and if it does it does not affect the following experiments.1b. Sample in formalin and other fixative: take about 20mg of sample, cut it into small pieces, place it in a centrifuge tube, add 500µL of 10mM PBS (PH7.4), vortex shaking, centrifuge at 12,000rpm for 1minute, discard the supernatant, and repeat 3 times. Add 180 µL Buffer GTL, 20 µL Proteinase K, vortex shaking to mix.2.56°C for 1 hour until the sample is completely dissolved. incubate at 90°C for 1 hour. centrifuge at 12,000 rpm, 25°C for 1 minute, and carefully pipette the lower aqueous phase (~180 µL) along the wall of the tube into a new centrifuge tube, trying to avoid aspirating the bottom precipitate and the upper layer of the wax solution.Note: 1) Samples can be left at room temperature after incubation at 56°C until the temperature of the water or dry bath reaches 90°C before placing the samples at 90°CIncubation.2) Optional step: add 7µL UNG (1U/µL), 50°C, 5min, no shaking. The purpose of this step is to minimize the risk of false positives by reducing the low-frequency occurrence of C>T:G>A transitions (artificial mutations) while effectively retaining the true occurrence of mutations.3. Optional step: If you need to remove RNA, you can lower the temperature of the sample to room temperature, then add 2µL of RNase A solution at a concentration of 100mg/mL, shake and mix well, and leave it at room temperature for 2 minutes.4. Add 20µL Proteinase K and incubate at 65℃, 450rpm for 15min.5. Add 200 µL of Buffer GL, mix well by vortexing and shaking, then add 200 µL of anhydrous ethanol and mix thoroughly by vortexing and shaking. Centrifuge briefly so that the solution on the wall of the tube collects at the bottom of the tube.Note: 1) Mix well immediately after adding Buffer GL and anhydrous ethanol.2) The addition of Buffer GL and anhydrous ethanol may produce a white precipitate that will not affect subsequent experiments.3) If more than one sample needs to be manipulated, the Buffer GL and anhydrous ethanol can be pre-mixed and spiked.6. Add all the solution obtained in step 5 to the adsorption columns (Spin Columns DF) that have been loaded into the collection tube, centrifuge at 25℃, 12000rpm for 2 minutes, pour out the waste liquid in the collection tube, and put the adsorption columns back into the collection tube.7. Add 500µL of Buffer GW1 to the adsorption column (check that anhydrous ethanol has been added before use), centrifuge at 12,000rpm for 1 minute, pour off the waste liquid in the collection tube, and put the adsorption column back into the collection tube.8. Add 500µL of Buffer GW2 to the adsorption column (check that anhydrous ethanol has been added before use), centrifuge at 12000rpm for 1 minute, pour off the waste liquid in the collection tube, and put the adsorption column back into the collection tube.Note: Step 8 can be repeated if further DNA purity is required.9.12 Centrifuge at 2000 rpm for 2 minutes and pour off the waste liquid in the collection tube. Leave the adsorption column at room temperature for several minutes to dry thoroughly.Note: The purpose of this step is to remove residual ethanol from the adsorption column; ethanol residue can interfere with subsequent enzymatic reactions.10. Place the adsorption column in a new 1.5 mL collection tube, add 20-100 µL of Buffer TE or sterilized water to the middle of the adsorption column overhanging the column, let it stand at room temperature for 2-5 minutes, centrifuge it at 12,000 rpm for 1 minute, and collect the DNA solution.-20°C to preserve DNA.Note: 1) The pH value of the eluent has a great influence on the elution efficiency, if water is used as the eluent should ensure that its pH value is 7.0-8.5, the pH value is lower than 7.0 when the elution efficiency is not high.2) If the final concentration of DNA is to be increased, the DNA eluate obtained in step 10 can be re-spiked onto the adsorbent membrane and left at room temperature for 2 minutes and centrifuged at 12,000 rpm for 1 minute... Read More | This plant protein extraction kit can extract soluble plant proteins from fresh, frozen, or dried plant tissues. Suitable for protein extraction from various plants and different parts of plants (such as roots, stems, leaves, flowers, seeds, etc.), with high extraction efficiency, high protein yieldThis plant protein extraction kit can extract soluble plant proteins from fresh, frozen, or dried plant tissues. Suitable for protein extraction from various plants and different parts of plants (such as roots, stems, leaves, flowers, seeds, etc.), with high extraction efficiency, high protein yield, high activity, and fast speed. The extracted protein can be directly subjected to protein electrophoresis analysis, immunoprecipitation, Western Blot, protein activity determination, and protein purification experiments. The concentration of the extracted protein can be determined using the BCA protein quantification kit. P665757Component100 TStorageP665757APlant Protein Extraction Reagent100 mLRTP665757BProtease Inhibitor Cocktail (100×)1 mL-20℃. Avoid freeze/thaw cycle. Precautions：1. This product contains 1mM EDTA.2. To prevent protein degradation, all operations should be carried out on ice as much as possible.3. After extracting protein using this product, the BCA method can be used for protein quantification.4. To achieve the best experimental results, please adjust the optimal usage amount according to the experiment.Operation steps:1. Please remove the required Plant Protein Extraction Agent for pre cooling before protein extraction.2. Weigh the weight of the experimental plant tissue. Add 5 ml of Plant Protein Extraction Agent to 1 g of tissue (add Protein Inhibitor Cocktail in a 1:99 ratio before protein extraction).Attention:1) Before homogenization, cut large pieces of plant tissue into small pieces and homogenize them with a mechanical homogenizer for 10 seconds, with an interval of 10 seconds. Repeat the process three times and select the appropriate homogenization method according to the different tissue samples.2) The amount of lysate used is adjusted according to different parts of the plant. If concentrated protein extracts are needed, the amount of Plant Protein Extraction Agent used can be appropriately reduced.3. After homogenization, incubate on ice for 20-30 minutes.4.4 ℃ 13400 × g, centrifuge for 20 minutes.5. Collect soluble proteins from the supernatant for further purification or downstream analysis... Read More | The commonly used method of eukarYOtic gene expression regulation research is the detection of reporter genes, and bioluminescence is the most commonly used and effective means of reporter gene detection. Luciferase can catalyze the conversion of the substrate luciferin and emit photons. This The commonly used method of eukarYOtic gene expression regulation research is the detection of reporter genes, and bioluminescence is the most commonly used and effective means of reporter gene detection. Luciferase can catalyze the conversion of the substrate luciferin and emit photons. This product provides a rapid, sensitive and stable detection method for the expression of Renilla luciferase reporter gene in mammalian cells. Product characteristic：1.Rapid : Cell lysis was completed within 10-15 min ；2.Convenience : The reagent is easy to prepare, and the sample detection steps are simple；Instruction：1. Cell lysis ( 1 ) Remove the culture medium and gently wash with PBS ( adherent cells can be directly performed this operation, suspension cells should be centrifuged to collect cells ). Add 1 × Lysis Buffer ( diluted component A with sterile water at 4 : 1 ) according to the following scheme, and then place the culture plate on a micro-oscillator at room temperature for 15 min to fully lyse the cells. Note : The pyrolysis products can be stored at room temperature for 6 h, and can be stored at − 70 °C for a long time ( the pyrolysis products cannot be repeatedly frozen and thawed ). ( 2 ) The pyrolysis products after full pyrolysis were centrifuged at 10000-15000 rpm for 3-5 min. After centrifugation, the supernatant was moved into a new EP tube for subsequent detection. 2. Preparation of working fluid ( 1 ) Restore all components to room temperature. ( 2 ) Dilute component C into renilla luciferase working solution with component B, and the dilution method is to add 1 µL C component to 49 µL B component. 3.chemiluminescence value detection ( 1 ) According to the operation instructions of the instrument, the instrument with chemiluminescence detection function was opened, such as multifunctional microplate reader. The parameters were set, the determination time was 10 s, and the determination interval was 2 s. ( 2 ) The cell lysis products were added to the measuring tube according to the volume of 20 ~ 100 µL ( keep the same amount of samples each time ). 1 × Lysis Buffer was blank control. ( 3 ) 100 µL renilla luciferase working solution was added to determine the RLU ( Relative light unit ) value ( Shaking mixing function is recommended for microplate reader ). Note : The renilla luciferase working solution cannot be stored for a long time. It is now ready for use and is used once. Component：RenillaLuciferase Lysis Buffer；RenillaLuciferase Assay Buffer；CoelenterazineMatters needing attention：Scope of application： Matters needing attention：1.Please instantaneously centrifuge the product to the bottom of the tube before use, and then carry out subsequent experiments ; 2.Due to the influence of temperature on the enzyme reaction, the sample and reagent should be measured after reaching room temperature. 3.The strongest wavelength of bioluminescence catalyzed by renilla luciferase is 480 nm, in order to prevent interference between holes, it is recommended to use white opaque orifice plate ;4. B component is recommended to carry out small batch packing according to the experimental requirements ; 5.It is recommended to use it now to avoid repeated freezing and thawing ; 6.For your safety and health, please wear experimental clothes and wear disposable gloves. Scope of application：Study on gene expression regulation and promoter... Read More |