| Description | Glutathione S-Transferase (GST) is a protein family with multiple physiological functions, primarily located within the cytoplasm. GST is a crucial component of the body's detoxification enzyme system. It mainly catalyzes the covalent binding of the sulfhydryl group of glutathione (GSH) to Glutathione S-Transferase (GST) is a protein family with multiple physiological functions, primarily located within the cytoplasm. GST is a crucial component of the body's detoxification enzyme system. It mainly catalyzes the covalent binding of the sulfhydryl group of glutathione (GSH) to various chemicals and their metabolites, converting electrophilic compounds into hydrophilic compounds that are more easily excreted in bile or urine. This process facilitates the degradation and elimination of potentially toxic substances from the body. Thus, GST plays a vital biological role in protecting cells from damage caused by electrophilic compounds. GST possesses GSH peroxidase activity (also known as non-Se-GSH-Px) and functions in repairing oxidatively damaged macromolecules such as DNA and proteins. The GST-catalyzed reaction consumes GSH but does not increase GSSG levels. Detection Principle: GST catalyzes the conjugation of GSH with CDNB (1-chloro-2,4-dinitrobenzene). The conjugation product has an absorption peak at 340 nm. The GST activity is calculated by measuring the rate of increase in absorbance at 340 nm. Detection Range: 2 - 76 U/L Sensitivity: 2 U/L Applicable Samples: Serum (plasma), animal/plant tissues, cells, bacteriaG1501773Component48T96TStorageG1501773AAssay Buffer60 mL120 mL2-8℃G1501773BChromogen11 mL22 mL2-8℃. Store in the dark.G1501773CSubstrate1EA1EA2-8℃. Store in the dark.Note: Before formal testing, it is recommended to perform a preliminary test with 2-3 samples expected to have significant differences.User-Prepared Instruments and ReagentsMicroplate reader or UV spectrophotometer (capable of measuring absorbance at 340 nm)96-well UV plate or micro quartz cuvettes, adjustable micropipettes and tipsIce maker, refrigerated centrifuge, water bathDeionized waterHomogenizer (for tissue samples)Experimental Procedure1. Reagent PreparationReagent NameReagent PreparationNotesAssay BufferReady-to-use; Equilibrate to room temperature before use.Store at 4℃.ChromogenReady-to-use; Equilibrate to room temperature before use.Store at 4°C protected from light. Skin irritant. Use appropriate protective equipment.Substrate Working ReagentBefore use, dissolve in 2.4 mL of deionized water.Unused reagent can be stored at 4°C protected from light for one month.2. Sample Preparation2.1 Animal/Plant TissuesWeigh 0.1 g of tissue sample. Add 1 mL of pre-cooled Assay Buffer and homogenize quickly on ice. Centrifuge the homogenate at 8,000 g, 4°C for 10 minutes. Collect the supernatant and keep it on ice for assay.2.2 Cells or BacteriaCollect 5×10⁶ cells or bacteria into a centrifuge tube. Wash with cold PBS, centrifuge, and discard the supernatant. Add 1 mL of Assay Buffer. Sonicate in an ice bath for 5 minutes (power 20% or 200W, pulse 3s on, 7s off, repeat 30 times). Centrifuge at 8,000 g, 4°C for 10 minutes. Collect the supernatant and keep it on ice for assay.2.3 Serum (Plasma)Assay directly.Note:Sample processing should be performed on ice. If not used immediately, samples can be stored at -80°C for one month.For GST activity measurement in cells, the cell count should be between 3-5×10⁶. Use Assay Buffer with sonication for cell extraction; do not use cell lysis buffers.If protein concentration measurement is required, Aladdin's BCA Protein Quantification Kit (B665595) or Ready-to-Use BCA Protein Quantification Kit (R1491648) is recommended.3. Assay Steps3.1 Preheat the microplate reader or UV spectrophotometer for at least 30 minutes. Set the wavelength to 340 nm. For UV spectrophotometers, zero the instrument with deionized water.3.2 Incubate the Substrate Working Reagent at 25°C (for general species) or 37°C (for mammals) for 15 minutes.3.3 Add reagents to a 96-well UV plate or micro quartz cuvette as follows:ReagentBlank (µL)Test (µL)Sample020Assay Buffer200Chromogen180180Substrate Working Reagent20203.4 Mix rapidly and immediately measure the change in absorbance at 340 nm. Record the absorbance for the Blank at 10 seconds (A1) and 310 seconds (A2). Record the absorbance for the Test at 10 seconds (A3) and 310 seconds (A4). Calculate ΔA blank = A2 - A1, ΔA test = A4 - A3. Note: Only one Blank is needed. A preliminary test with 2-3 samples showing expected significant differences is recommended. If the sample absorbance is greater than 1, dilute the sample with deionized water and multiply the result by the dilution factor. The reaction temperature significantly affects the results; maintain it at 25°C (general species) or 37°C (mammals). 4. Calculation of Results 4. Calculation of Results Note: We provide both derived and simplified calculation formulas. They are equivalent. The simplified formulas in bold are recommended for final calculation. 4.1 Calculation Formulas for 96-well UV Plate (1) Based on Protein Concentration Unit Definition: One unit of enzyme activity is defined as the amount that catalyzes the conjugation of 1 µmol of CDNB with GSH per minute per milligram of protein at 25°C or 37°C. Calculation Formula: GST Activity (U/mg prot) = (ΔAtest - ΔAblank) ÷ (ε × d) × 10⁶ × Vtotal reaction ÷ (Cpr × Vsample) ÷ T = 0.46 × (ΔAtest - ΔAblank) ÷ Cpr (2) Based on Sample Fresh Weight Unit Definition: One unit of enzyme activity is defined as the amount that catalyzes the conjugation of 1 µmol of CDNB with GSH per minute per gram of sample at 25°C or 37°C. Calculation Formula: GST Activity (U/g fresh weight) = (ΔAtest - ΔAblank) ÷ (ε × d) × 10⁶ × Vtotal reaction ÷ (Vsample ÷ Vtotal sample × W) ÷ T = 0.46 × (ΔAtest - ΔAblank) ÷ W (3) Based on Cell or Bacterial Count Unit Definition: One unit of enzyme activity is defined as the amount that catalyzes the conjugation of 1 µmol of CDNB with GSH per minute per 10⁴ cells or bacteria at 25°C or 37°C. Calculation Formula: GST Activity (U/10⁴) = (ΔAtest - ΔAblank) ÷ (ε × d) × 10⁶ × Vtotal reaction ÷ (500 × Vsample ÷ Vtotal sample) ÷ T = 0.46 × (ΔAtest - ΔAblank) ÷ 500 (4) Based on Liquid Volume Unit Definition: One unit of enzyme activity is defined as the amount that catalyzes the conjugation of 1 µmol of CDNB with GSH per minute per milliliter of liquid at 25°C or 37°C. Calculation Formula: GST Activity (U/mL) = (ΔAtest - ΔAblank) ÷ (ε × d) × 10⁶ × Vtotal reaction ÷ Vsample ÷ T = 0.46 × (ΔAtest - ΔAblank) 4.2 Calculation Formulas for Micro Quartz Cuvette Adjust the pathlength (d) in the formulas above from 0.5 cm to 1 cm for calculations. Parameter Definitions: ε: Molar extinction coefficient of the product, 9.6 × 10³ L/mol/cm d: Light path for 96-well plate (0.5 cm) 10⁶: Conversion factor (1 mol = 1 × 10⁶ µmol) V total reaction : Total reaction volume (220 µL = 2.2 × 10⁻⁴ L) Cpr: Protein concentration of the supernatant (mg/mL) W: Sample weight (g) V sample : Volume of supernatant added to the reaction system (20 µL = 0.02 mL) V total sample : Volume of extraction buffer added (1 mL) T: Reaction time (5 min) 500: Cell or bacterial count factor (5 × 10⁶ total cells / 10⁴ per unit = 500)Precautions1. Biochemical reagents are generally irritating and potentially biologically toxic. For your safety and health, implement appropriate biosafety precautions throughout the experiment, including wearing lab coats, masks, gloves, and head covers. Perform experiments in a fume hood or biosafety cabinet.2. This product is for research use only. Not for use in clinical diagnosis... Read More | Calcein AM /PI Double Staining Kitis utilized for simultaneous fluorescence staining of viable and dead cells. This kit contains Calcein-AM and Propidium Iodide (PI) solutions, which stains viable and dead cells, respectively(Fig. 1). Calcein-AM, an acetoxymethyl ester of calcein, is highly Calcein AM /PI Double Staining Kitis utilized for simultaneous fluorescence staining of viable and dead cells. This kit contains Calcein-AM and Propidium Iodide (PI) solutions, which stains viable and dead cells, respectively(Fig. 1). Calcein-AM, an acetoxymethyl ester of calcein, is highly lipophilic and cell membrane permeable. Though Calcein-AM itself is not a fluorescent molecule, the calcein generated from Calcein-AM by esterase in a viable cell emits a strong green fluorescence (excitationat 490 nm, emission at515 nm). Therefore, Calcein-AM only stains viable cells. On the other hand, PI, a nuclei staining dye, cannot pass through a viable cell membrane. It reaches the nucleus by passing through disordered areas of dead cell membrane, and intercalates with the DNA double helix of the cell to emit red fluorescence (excitation: 535 nm,emmision: 617 nm). Since both calcein and PI-DNA can be excited with 490 nm, simultaneous monitoring of viable and dead cells is possible with a fluorescence microscope. With 545 nm excitation, only dead cells can be observed (Fig. 1). Since optimal staining conditions differ from cell line to cell line, we recommend that a suitable concentration of PI and Calcein-AM be individually determined. Please note that PI is suspected to be highly carcinogenic;careful handling is required.Required Equipment and Materials:Microscope with 490 nm excitation filter and 530 nm emission filter；CO2incubator；10 µl and 200 µl adjustable pipettes, PBSSolution A (Calcein-AM)；Solution B (PI) Storage Condition: -20oC ；Shipping Condition: blue ice.Application:Assay Procedure1)Add 2.5 µl Solution A and 12.5 µl Solution B to 5 ml PBS to prepare assay solution.*2)Wash the cell with PBS several times to remove residual esterase activity.3)Add 100uLof assay solution to200uL105~106CELLSsolution and incubate the mixture at 37oC for 15 min.4)Detect fluorescence using a fluorescence mircoscope with 490 nm excitationfor simultaneous monitoring of viable and dead cells.With 545 nm excitation, only dead cells can be observed.*The following steps may be necessary tooptimizethe suitable concentration of each reagent:1)Prepare dead cells by 10 min incubation in 0.1% saponin or 0.1-0.5% digitonin or by 30 min incubation in 70% ethanol.2)Stain dead cells with 0.1-10 µM PI solution to find a PI concentration that stains the nucleus only, not the cytosol.3)Stain dead cells with 0.1-10 µM Calcein-AM solution to find a Calcein-AM concentration that does not stain the cytosol. Then stainviable cells with that Calcein-AM solution to check whether the viable cell can be stained... Read More | D665729 Component 50 T Storage D665729A Conversion Buffer CR 5×1 mL RT D665729B Buffer CL 30 mL RT D665729C Buffer MD 0.4 mL RT D665729D Buffer DB 10 mL RT D665729E Buffer WB (concentrate) 10 mL RT D665729F Buffer GW1 (concentrate) 13 mL RT D665729G Buffer GW2 (concentrate) 15 mL RT D665729H D665729 Component 50 T Storage D665729A Conversion Buffer CR 5×1 mL RT D665729B Buffer CL 30 mL RT D665729C Buffer MD 0.4 mL RT D665729D Buffer DB 10 mL RT D665729E Buffer WB (concentrate) 10 mL RT D665729F Buffer GW1 (concentrate) 13 mL RT D665729G Buffer GW2 (concentrate) 15 mL RT D665729H Buffer EB 4 mL RT D665729I Buffer PS 10 mL RT D665729J Spin Columns DF 50 Pcs 2-8 ℃ D665729K Collection Tubes 50 Pcs RTProduct Introduction:The basic principle of this reagent kit is that after DNA is treated with sodium bisulfite, unmethylated cytosine can be transformed into uracil, while methylated cytosine remains unchanged. And adopting an innovative high-temperature treatment method, the transformation time is greatly shortened, the transformation efficiency is improved, and the transformation efficiency can reach over 99%. At the same time, using a silicon-based membrane purification column, DNA can be recovered and purified from the methylated solution through a simple binding washing elution step. The recovered DNA has high purity and good integrity, and can be directly used for sequencing, methylated PCR detection, chip analysis, connection and transformation, enzyme digestion, labeling, microinjection, PCR and in vitro transcription and other molecular biology experiments.Self prepared reagents: anhydrous ethanol, 75% ethanol.Preparation and important precautions before the experiment1. Product usage method:(1) 10 times packaging preparation method: CT Conversion Agent is a solid mixture that must be prepared before first use. Add 2 ml sterile water and 100 µ M-Dissolving Buffer and 300 µ Add M-Diffusion Buffer to the CT Conversion Agent tube. Dissolve at 55 ° C and shake until completely dissolved. Store the CT Conversion Agent solution at room temperature (20 ° C-30 ° C) in the dark before use. The CT Conversion Agent for each tube is designed for 10 DNA treatments. In order to achieve better results, the prepared CT Conversion Agent should be used immediately. If not used immediately, the CT Conversion Agent solution can be stored at -20 ° C for 1 week. Before use, be sure to thaw the stored CT Conversion Agent solution at room temperature and mix thoroughly by shaking or inverting for 2 minutes, CT Conversion Reagent is sensitive to light, so it is important to minimize exposure to light as much as possible.(2) 50 times packaging preparation method: CT Conversion Agent and M-Dissolving Buffer are solid mixtures that must be prepared before first use. Add 5 ml of sterile water to the M-Dissolving Buffer and shake to dissolve. After all the solids have dissolved, transfer all the solution from the M-Dissolving Buffer tube to the CT Conversion Agent tube and add 5.5 ml of sterile water. Add 1.5 ml of M-Dilution Buffer to the CT Conversion Agent tube. Dissolve at 55 ° C and shake until completely dissolved. Store the CT Conversion Agent solution at room temperature (20 ° C-30 ° C) in the dark before use. The CT Conversion Agent for each tube is designed for 50 DNA treatments. In order to achieve better results, the CT Conversion Agent should be used immediately after preparation. If not immediately used, the CT Conversion Agent solution can be stored at -20 ° C for 1 week. Before use, be sure to thaw the stored CT Conversion Agent solution at room temperature and mix thoroughly by shaking or inverting for 2 minutes, CT Conversion Reagent is sensitive to light, so it is important to minimize exposure to light as much as possible.2. Before the first use, anhydrous ethanol should be added to the M-Wash Buffer according to the instructions on the reagent bottle label.Operation stepsThe range of DNA prepared each time is 1 ng-4 µ Between g, the optimal amount is 500 ng-2 µ G.1. Take 20 µ Add DNA sample into centrifuge tube (self provided), and if the sample amount is insufficient, replenish with water up to 20 µ L.2. Add 2.2 to the DNA sample µ Mix the sample well with the M-Dilution Buffer of l.3.42 ℃ water bath for 30 minutes.4. Add 220 to the sample obtained from the previous step µ Prepare the CT Conversion Agent solution, mix well, and incubate in an 80 ℃ constant temperature water bath in a dark place for 60 minutes.5. Add 480 to the solution in the previous step µ M - Buffer PA, gently mix upside down.6. Column balance: Add 200 to the spin columns DS that have been loaded into the collection tube µ Centrifuge at 12000 rpm (~13400 × g) for 2 minutes, discard the waste liquid in the collection tube, and place the adsorption column back into the collection tube.7.Add all the solution obtained from step 5 to the adsorption column (already loaded into the collection tube), let it stand at room temperature for 2 minutes, centrifuge at 12000 rpm for 1 minute, discard the waste liquid in the collection tube, and place the adsorption column back into the collection tube.Attention: The maximum capacity of the adsorption column is 750 µ l. If the sample volume is greater than 750 µ L can be added in batches.8. Add 500 to the adsorption column µ Centrifuge at 12000 rpm for 1 minute using M-Buffer PA, discard the waste liquid from the collection tube, and place the adsorption column in the recovery tube.9. Add 650 to the adsorption column µ M-Wash Buffer (please check if anhydrous ethanol has been added before use), centrifuge at 12000 rpm for 1 minute, discard the waste liquid in the collection tube, and place the adsorption column in the collection tube.10.12000 rpm for 2 minutes, discard the waste liquid, and 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 will affect subsequent enzymatic reactions (such as enzyme digestion, PCR, etc.).11. Place the adsorption column into a new centrifuge tube (provided by oneself), and add 20 drops to the middle position of the adsorption membrane in the air µ M-Elution Buffer (pH 8.5), leave at room temperature for 2 minutes. Collect DNA solution by centrifugation at 12000 rpm for 1 minute.12. Collect 20 µ Add 2.2 to DNA µ M-Diffusion Buffer, let it stand at room temperature for 30 minutes.13. Add 500 to the solution µ After pre cooling anhydrous ethanol, invert and mix well, and place the solution at -20 ℃ to precipitate for 30 minutes (overnight precipitation is more effective).14.12000 rpm for 15 minutes and gently discard the supernatant.15. Add 75% ethanol, centrifuge at 12000 rpm for 1 minute, pour out the supernatant, wait for ethanol to evaporate at room temperature, then add 20 µ Dissolve the M-Elution buffer and store the DNA at -20 ℃. The DNA collected in this step can be used for subsequent related experiments... Read More | Products contentN665989Component240 TStorageN665989AIndex N501 Primers for Illumina240 µL-20℃. Avoid freeze/ Thaw cycle.N665989BlIndex N901-N924 Primers for Illumina24×10 µL-20℃. Avoid freeze/ Thaw cycle.Note: The amount of individual primers used is 1 µl, each N7-endProducts contentN665989Component240 TStorageN665989AIndex N501 Primers for Illumina240 µL-20℃. Avoid freeze/ Thaw cycle.N665989BlIndex N901-N924 Primers for Illumina24×10 µL-20℃. Avoid freeze/ Thaw cycle.Note: The amount of individual primers used is 1 µl, each N7-end primer can perform 10 DNA library constructs, and each kit can perform 240 DNA library constructs. Products IntroductionThis kit is a companion kit to the transposase-based Rapid DNA Library Construction Kit for Illumina platform library construction. Each kit contains one N5 primer and 24 N7 primers, which can be used to prepare 24 different single-ended Index libraries. All reagents provided in the kits have been subjected to stringent quality control and functional validation to maximize the stability and reproducibility of library construction. The libraries can be used for sequencing on Illumina platforms such as HiSeq X-10/4000/2500/2000 and MiSeq. Provide your own instruments, reagents and consumables1. Magnetic frame: DynaMagTM-2 is recommended.2. DNA purification and recovery kit: It is recommended to use Kangwei DNA purification and recovery kit by magnetic bead method.3. DNA building kit: It is recommended to use the Kangwei Century transposase method second-generation sequencing rapid DNA building kit.4. Anhydrous ethanol.5. Reaction tubes: It is recommended to use low adsorption PCR tubes with 1.5 ml centrifuge tubes;Tip: It is recommended to use a high quality filter tip to prevent contamination of kits and library samples. Pre-experiment Preparation and Important NotesPlease centrifuge briefly before opening the cap so that the liquid collects at the bottom of the tube to avoid cross-contamination between different primers. ProcedureFor the use of the CombiVision Second Generation Sequencing Multisample Primer Kit, please follow the CombiVision Second Generation Sequencing Rapid DNA Library Kit protocol.Index N501 Primer for IlluminaIndex N901-N996 Primer for Illumina... Read More | The Succinic Acid (Succinate) assay kit is suitable for the specific assay of succinic acid in wine, cheese, eggs, sauce and other food products. Succinic acid (or succinate) is found in all plant and animal materials as a result of the central metabolic role played by this dicarboxylic acid in the The Succinic Acid (Succinate) assay kit is suitable for the specific assay of succinic acid in wine, cheese, eggs, sauce and other food products. Succinic acid (or succinate) is found in all plant and animal materials as a result of the central metabolic role played by this dicarboxylic acid in the Citric Acid Cycle. Succinic acid concentrations are monitored in the manufacture of numerous foodstuffs and beverages, including wine, soy sauce, soy bean flour, fruit juice and dairy products (e.g. cheese).Product Description： Succinic acid is found in all plant and animal materials as a result of the central metabolic role played by this dicarboxylic acid in the Citric Acid Cycle. Succinic acid concentrations are monitored in the manufacture of numerous foodstuffs and beverages, including wine, soy sauce, soy bean flour, fruit juice and dairy products (e.g. cheese). The ripening process of apples can be followed by monitoring the falling levels of succinic acid. The occurrence of > 5 mg/kg of this acid in egg and egg products is indicative of microbial contamination. Apart from use as a flavouring agent in the food and beverage industries, succinic acid finds many other non-food applications, such as in the production of dyes, drugs, perfumes, lacquers, photographic chemicals and coolants. Preparation Instructions：Suitable for succinate determination in food, beverage, agricultural products, and other biological samples.Note for Content：The number of manual tests per kit can be doubled if all volumes are halved. This can be readily accommodated using the MegaQuantTM Wave Spectrophotometer (D-MQWAVE).Browse all of our organic acid assay kits.Principle：The Succinate Assay Kit provides a simple, one step assay for measuring succinate. In this assay succinate is converted to pyruvate which reacts with specific reagents and dye to form a colored product. The color intensity at 570 nm or fluorescencAdvantages：Extended cofactors stability. Dissolved cofactors stable for > 1 year at 4oC.Very competitive price (cost per test)All reagents stable for > 2 years as suppliedVery rapid reaction (even at room temperature)Mega-Calc™ software tool is available from our website for hassle-free raw data processingStandard includedSuitable for manual, microplate and auto-analyser formats... Read More |