| Description | Triglycerides (TG), also known as triacylglycerols, are fat molecules formed from three long-chain fatty acids and glycerol. They are the most abundant lipids in the human body. Most tissues can utilize the breakdown products of triglycerides for energy. Simultaneously, tissues like the liver and Triglycerides (TG), also known as triacylglycerols, are fat molecules formed from three long-chain fatty acids and glycerol. They are the most abundant lipids in the human body. Most tissues can utilize the breakdown products of triglycerides for energy. Simultaneously, tissues like the liver and adipose tissue can synthesize triglycerides. The enzymatic method for measuring TG is commonly used in biochemical assays due to its characteristics: 1. High sensitivity, accuracy, and precision; 2. Use of mild reaction conditions; 3. Simple operation; 4. Suitable for semi-automatic biochemical analyzers.Detection Principle: Triglycerides are hydrolyzed by lipoprotein lipase (LPL) into glycerol and free fatty acids. Glycerol is then phosphorylated by glycerol kinase (GK) and adenosine triphosphate (ATP) to form glycerol-3-phosphate (G-3-P). G-3-P is subsequently oxidized by glycerol-3-phosphate oxidase (GPO), producing hydrogen peroxide. The hydrogen peroxide, in the presence of peroxidase (POD), 4-aminoantipyrine (4-AAP), and phenol (collectively known as PAP), reacts to form a red-colored quinoneimine dye (Trinder reaction). The quinoneimine dye has a maximum absorption at 510 nm. The absorbance is directly proportional to the triglyceride concentration in the sample and can be measured using a microplate reader between 500-520 nm.This kit is used for the quantitative determination of triglyceride content in serum, cells, tissues, and other samples from humans or animals. This kit is intended for research use only and is not suitable for clinical diagnosis or other purposes.Component100TStorageBuffer Solution24 mL2-8℃. Store in the dark.Enzyme Reagent6 mL2-8℃. Store in the dark.Glycerol Standard (1.7 mmol/L)1 mL2-8℃User-Prepared Instruments and ReagentsddH₂O, Physiological Saline or PBSCentrifuge tubes or small test tubes, Water bath or incubatorMicroplate reader, 96-well plate, Semi-automatic biochemical analyzerExperimental Procedure1. Sample Preparation1.1 Serum, Plasma, Cerebrospinal Fluid SamplesSerum or plasma separated from the test sample should not be hemolyzed. Assay directly. If the concentration exceeds the linear range, dilute with physiological saline before assaying.1.2 Cell Samples(1) Take an appropriate amount of cells (generally recommended >10⁶), centrifuge at 1000 g for 10 min, discard the supernatant, keep the pellet.(2) Wash the pellet 1-2 times with PBS or physiological saline, centrifuge at 1000 g for 10 min, discard the supernatant, keep the pellet.(3) Add 200-300 µL of PBS or physiological saline to homogenize. Sonicate the cells on ice (power 300W, pulse 3-5s, interval 30s, repeat 3-5 times). Alternatively, homogenize manually. Do not centrifuge the prepared homogenate. Alternatively, lyse with 1-2% Triton X-100 on ice for 30-60 min. Do not centrifuge the prepared lysate.1.3 Tissue SamplesAccurately weigh an appropriate amount of tissue sample. Add physiological saline or PBS at a mass (g) to volume (mL) ratio of 1:9. Homogenize manually or mechanically on ice. Centrifuge at 2500-3000 g for 10 min. Collect the supernatant for assay.2. Preparation of GPO-PAP Working SolutionBefore use, mix the Buffer Solution and Enzyme Reagent at a 4:1 volume ratio. Mix well. Store at 4°C.3. TG Assay Steps using Microplate Reader3.1 Add reagents sequentially to the 96-well plate according to the table below. Mix thoroughly and incubate at 37°C in a water bath or incubator for 10 minutes.Reagent (µL)Blank WellStandard WellTest WellddH2O2.5//Glycerol Standard (1.7 mmol/L)/2.5/Test Sample//2.5GPO-PAP Working Solution2502502503.2 Measure the absorbance between 500-520 nm using the microplate reader. Zero the instrument with the blank well, then read the absorbance of the standard well and all test wells.4. TG Assay Steps using Semi-Automatic Biochemical Analyzer4.1 Instrument Parameter Settings:WavelengthTemperatureDelay TimeMeasurement TimeReagent BlankReaction TypeAspiration Volume510-550nm37℃2s2sYesEndpoint800µL4.2 Add reagents sequentially to tubes according to the table below. Mix thoroughly and incubate at 37°C in a water bath for 10 minutes.Reagent (µL)Blank TubeStandard TubeTest TubeddH2O10//Glycerol Standard (1.7 mmol/L)/10/Test Sample//10GPO-PAP Working Solution1000100010004.3 Zero the instrument with the blank tube, then read the absorbance of the standard tube and all test tubes.5. Calculation Formula5.1 For serum, plasma, and other liquid samples (Blank zeroed):TG (mmol/L) = (Absorbance of Test Well/Tube / Absorbance of Standard Well/Tube) × 1.7 mmol/L5.2 For cell, tissue, and other samples (Blank zeroed):TG (mmol/g prot) = (Absorbance of Test Well/Tube / Absorbance of Standard Well/Tube) × 1.7 mmol/L / Sample Protein Concentration (mg/mL)Reference Interval (Healthy Adults)Desirable range: < 1.7 mmol/L (< 150 mg/dL)Borderline high: 1.7 – 2.25 mmol/L (150 – 199 mg/dL)High: 2.26 – 5.64 mmol/L (200 – 499 mg/dL)Very high: ≥ 5.65 mmol/L (≥ 500 mg/dL)Precautions1. Avoid repeated freeze-thaw cycles for the low-temperature reagents mentioned above to prevent inactivation or decreased efficiency.2. The GPO-PAP Working Solution should be prepared immediately before use and is not suitable for long-term storage at 4°C.3. This method can be directly used to detect TG content in cerebrospinal fluid but cannot directly detect TG in urine, as untreated urine contains reducing substances that interfere with the peroxidase reaction.4. If test samples cannot be assayed immediately, they should be stored at 2-8°C and are stable for 3 days.5. The linear range of this method is up to 9.0 mmol/L. If the sample TG concentration is too high, results may be falsely low. Dilute the sample with physiological saline and re-assay, multiplying the result by the dilution factor.6. The working reagent should be protected from contamination by substances like glucose and cholesterol.7. The reagent is susceptible to oxidation by air, turning red. A blank measurement is necessary.8. For your safety and health, please wear a lab coat and disposable gloves during operation.9.Use the reagents as soon as possible after opening to prevent affecting subsequent experimental results... Read More | CFDASE cell proliferation and tracking detection kit is a kit for cell proliferation and tracking detection based on CFDA se. This kit is composed of CFDASE powder, solvent and staining buffer. CFDASE is a derivative of fluorescein diacetate (FDA), which has cell membrane permeability and CFDASE cell proliferation and tracking detection kit is a kit for cell proliferation and tracking detection based on CFDA se. This kit is composed of CFDASE powder, solvent and staining buffer. CFDASE is a derivative of fluorescein diacetate (FDA), which has cell membrane permeability and does not have fluorescence luminescence. When CFDASE penetrates the cell membrane into living cells, it can be catalysed by esterases in the cytosol to produce carboxyfluorescein succinimidyl ester (CFSE), which can emit strong green fluorescence, cannot penetrate the cell membrane, and can remain intact in the cell. CFSE can also spontaneously and irreversibly covalently bind to intracellular amino groups to couple to cellular proteins. Meanwhile, the excess and uncoupled CFDASE returned to the extracellular medium by passive diffusion and was cleared by subsequent washing steps. The fluorescence of non dividing cells labeled by CFDASE is very stable, and the stable labeling time can reach several months, so it is very suitable for cell community analysis. The fluorescence of CFDASE labeled cells is very homogeneous, which is superior to other cell tracking fluorescent probes used previously, such as PKH26, and the fluorescence distribution of the divided progeny cells is also very uniform. In the process of cell division and proliferation, CFSE labeled fluorescence can be evenly distributed to the two progeny cells, and the fluorescence intensity becomes half of the parental cells. According to the fluorescence intensity, flow cytometer (FL1 channel) can detect undivided cells, cells that divide once (1 / 2 of the fluorescence intensity), twice (1 / 4 of the fluorescence intensity), three times (1 / 8 of the fluorescence intensity), and cells that divide more times. CFDASE can detect up to eight or more cleavages. CFDASE labeled cells can be used for proliferation studies in vitro and in vivo, and have the function of not staining adjacent cells. CFDASE is most commonly used to detect the proliferation of lymphocytes, and can also be used to detect the proliferation of fibroblasts, NK cells and other cells. CFDASE labeled cells showed green fluorescence. In addition to flow cytometry to detect cell proliferation, fluorescence microscopy can also be used for homogeneous staining of cell tracking observation.Components:ComponentsC598182-20TC598182-500TA. CFDA SE1 tube1 tubex5B.CFDA SE solvent20 µL500 µLC.10x CFDA SE Buffer1 mL x250 mLMatters needing attention：1. please centrifuge the product to the bottom of the tube immediately before use, and then conduct subsequent experiments. 2. CFDA and Se are easily hydrolyzed and will deteriorate quickly in aqueous solution. Please avoid contact with water during use. Contact with water during the process of labeling cells is within the permitted range. 3. CFDA se solvent will solidify at lower temperatures such as 4 º C and ice bath and stick to the bottom, wall or cover of the centrifugal tube. It can be used after incubating in a 20-25 º C water bath for a while until it is completely dissolved. 4. this kit optimizes the CFDA se staining system, but users are advised to explore the optimal working concentration and staining time according to their own cell type, culture conditions and application direction. Different cells have different lactonase activities, so the staining effect is different. 5. fluorescent dyes have quenching problems. Please avoid light during operation to slow down fluorescence quenching. 6. for your safety and health, please wear experimental clothes and disposable gloves.Usage method:1. Preparation of reagents(1) Preparation of CFDA SE storage solution: Take one tube of CFDA SE provided in the reagent kit and restore it to room temperature. Instantly centrifuge to allow the powder to fully settle to the bottom of the tube. Add 100 µ L CFDA SE solvent (add 20 µ L CFDA SE solvent) to it and dissolve it thoroughly to prepare CFDA SE storage solution (1000 ×). Prepared CFDA SE storage solution, stored at -20 ℃ in the dark, with a shelf life of two months- Storing at 70 ℃ in the dark can extend the usage time appropriately.(2) Preparation of CFDA SE Buffer: Dilute 10 x CFDA SE Buffer to 1 x with sterile cell culture grade water as needed. The prepared 1 × CFDA SE Buffer can be stored at 4 ℃ and can be stored at -20 ℃ if not in use for a long time.2. Marking and detection(1) Centrifuge the collected cells, use 1 mL 1 × CFDA SE Buffer to re suspend the cells in a 15 mL centrifuge tube, and adjust the cell concentration to 1-5 × 106 cells/mL.(2) Preparation of CFDA SE working solution: Dilute the CFDA SE storage solution (1000 ×) with 1 × CFDA SE Buffer to 2 ×.(3) Staining: Add 1 mL of CFDA SE working solution (2 x) to 1 mL of cell suspension to be labeled, invert and mix well, and incubate at 37 ℃ for 10 minutes.(4) Immediately add 5 times the volume of preheated complete culture medium (including serum) to the centrifuge tube, invert and mix well to terminate the labeling reaction.(5) Centrifuge at 1000 rpm for 5 minutes at room temperature to remove the supernatant, then wash once with 5-10 mL of complete culture medium.(6) Add 5-10 mL of complete culture medium and incubate at 37 ℃ for 5 minutes to promote the residence of CFDA SE in the cells and the entry of unreacted CFDA SE into the complete cell culture medium. Centrifuge at 1000 rpm for 5 minutes at room temperature to remove the supernatant and complete the final wash.(7) Subsequently, the cells can be cultured using the normal cultivation method. The labeling effect can be directly observed under a fluorescence microscope, or cell proliferation can be detected by flow cytometry after appropriate cultivation time, showing green fluorescence. The labeled cells can also be used for transplantation in live animals and for fluorescence tracing.Note: a If cell fixation is required, use aldehyde fixatives such as 4% paraformaldehyde to fix at room temperature for 15 minutes; If additional labeling such as antibody labeling is required afterwards, please permeabilize the cells with ice acetone for 10 minutes. b. The optimal labeling concentration and incubation time for CFDA SE vary for different cells. The initial experiment can be conducted according to the experimental steps. If the effect is not satisfactory, it is recommended to adjust the staining concentration and incubation time to achieve the best labeling effect.Scope of application：Cell proliferation assay... 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 | Products content S666097Component200 TStorageS666097A5×SuperFast One Step RT-qPCR U+ Buffer1 mL-20℃. Avoid freeze/thaw cycle.S666097BSuperFast One Step U+ Enzyme200 µL-20℃. Avoid freeze/thaw cycle.S666097CRNase-Free Water2×1.5 mL-20℃. Avoid freeze/thaw cycle. Products content S666097Component200 TStorageS666097A5×SuperFast One Step RT-qPCR U+ Buffer1 mL-20℃. Avoid freeze/thaw cycle.S666097BSuperFast One Step U+ Enzyme200 µL-20℃. Avoid freeze/thaw cycle.S666097CRNase-Free Water2×1.5 mL-20℃. Avoid freeze/thaw cycle. Products IntroductionThe SuperFast Probe One Step RT-qPCR U+ Kit is designed for quantitative PCR assays using RNA as a template (e.g., RNA viruses). Using gene-specific primers (GSP), reverse transcription and qPCR reactions are completed in a single tube, eliminating the need for additional tube-opening/pipetting operations, greatly increasing throughput and reducing the risk of contamination. The dUTP/UNG anti-contamination system is introduced in this kit. The heat-sensitive UNG rapidly degrades U-containing contaminants at room temperature; it is rapidly inactivated by reverse transcription at 55°C, without affecting the efficiency and sensitivity of qRT-PCR. Combined with optimized buffer systems and antibody-modified Taq enzymes and mutated M-MLV, the SuperFast Probe One Step RT-qPCR U+ Kit provides sensitivity up to 0.1 pg of total RNA or <10 copies of RNA template and enhanced thermal stability. 5× SuperFast One Step RT-qPCR U+ Buffer contains the following components The 5× SuperFast One Step RT-qPCR U+ Buffer contains an optimized buffer system and dNTP/dUTP Mix, which is particularly suitable for high specificity, low template concentration and multiplexed rapid detection of fluorescently labeled probes such as TaqMan. caveatBefore use, please mix the product gently by turning it up and down after it is completely melted to avoid foaming, and use it after brief centrifugation. Avoid repeated freezing and thawing of the product.ROX dye is used to correct the fluorescence signal error between the quantitative PCR wells, this product does not contain ROX dye, if you need to match the ROX dye with the instrument you are using, please contact your local business or call CombiSense customer service at 4006-222-360. PCR reaction system Attention: (1) Usually, the final primer concentration of 0.2 µM can get better results, and 0.1-1.0 µM can be used as a reference for setting the range. If the amplification efficiency is not high, the concentration of primer can be increased; if non-specific reaction occurs, the concentration of primer can be decreased to optimize the reaction system.(2) The final concentration of the probe used is related to the fluorescence quantitative PCR instrument used, the type of probe, and the type of fluorescent labeling substance, please refer to the instrument manual or the specific requirements for the use of each fluorescent probe to adjust the concentration.3) Because templates from different species contain different numbers of copies of the target gene, the template can be diluted in a gradient to determine the optimal amount of template to usePCR reaction conditionsmovetemptimingcirculatereverse transcription55°C1 min1premutability95°C10s1)1denaturation95°C1 s40-45Annealing/Extension55-60°C2)10-15s3)40-45Attention: (1) The enzyme used in this product is activated under the condition of pre-denaturation at 95℃ for 30s. Under this condition, most of the templates can be well unchained. For templates with high GC content and complex secondary structure, the pre-denaturation time can be extended to 1min, so as to make the starting template fully unchained, and if the high temperature treatment time is too long, it will affect the activity of the enzyme; for simple templates, pre-denaturation time of 1-10s can also be used, and the optimal pre-denaturation time can be determined according to the template situation.(2) It is recommended to use two-step PCR reaction program, the annealing temperature should be 55-60℃ as the reference range, and the annealing temperature can be increased when non-specific reaction occurs. If you can't get good results due to the use of primers with low Tm values or long amplification products, you can try three-step PCR amplification.3) Whether the actual Real Time PCR instrument used supports rapid amplification cycles, please perform a pre-experiment to verify this for the first attempt... Read More | Products contentProducts IntroductionThis kit is suitable for simple, rapid and efficient isolation and purification of DNA/RNA from whole blood, tissue homogenates, swabs, serum, plasma and other cell-free body fluids, etc. The unique buffer system enables the viral nucleic acids in the lysate to Products contentProducts IntroductionThis kit is suitable for simple, rapid and efficient isolation and purification of DNA/RNA from whole blood, tissue homogenates, swabs, serum, plasma and other cell-free body fluids, etc. The unique buffer system enables the viral nucleic acids in the lysate to bind to the silica gel centrifugal adsorbent columns in a highly efficient manner, and the viral nucleic acids obtained are of high purity and stable quality, free of protein, nuclease and other impurities, and can be used in a variety of routine operations, including PCR, fluorescence quantitative PCR and other experiments. It can be used for a variety of routine operations, including PCR, fluorescence quantitative PCR and other experiments.Bring your own instrumentsThermostatic mixer.Pre-experiment Preparation and Important Notes1. Read these instructions carefully before experimenting.2. If Proteinase K is to be stored for a long period of time, please keep it at -20℃.3. Check Buffer RLC for crystallization or precipitation prior to use, and if crystallization or precipitation occurs, redissolve Buffer RLC in a 56°C water bath.4. Pre-treatment of tissue samples: Take 20 mg of tissue samples into 1.5 mL centrifuge tubes (self-provided), add 500 µL of Buffer RLC, and after the tissue homogenizer breaks up, centrifuge the samples for 1 minute at 12,000 rpm (~13,400×g), and take 200 µL of supernatant as samples. procedure1. Take a 1.5 mL centrifuge tube (provided), add 500 µL of Buffer RLC, 200 µL of sample, 20 µL of Proteinase K, vortex for 5 s, and then place it in a thermostatic mixer at 1200 rpm for 10 min at room temperature. Note: For wet swab samples, 200 µL of sample was taken after sufficiently shaking and mixing. Note: For wet swabs, 200 µL was taken from the sample after it was soaked in 400 µL of saline, shaken and mixed thoroughly for 5 minutes, and then centrifuged at 12,000 rpm for 1 minute, and 200 µL was taken for extraction.2. Instantly remove the centrifuge tube and add the solution from step 1 to the Spin Columns DM in the collection tube. centrifuge at 12,000 rpm (~13,400 x g) for 1 minute, pour off the waste liquid from the collection tube, and return the column to the collection tube.3. Add 500 µL of Buffer PGWT to the adsorbent column, centrifuge at 12,000 rpm for 1 minute, pour off the waste liquid from the collection tube, and return the column to the collection tube.4. Add 500 µL of Buffer GWT2 to the adsorbent column, centrifuge at 12,000 rpm for 1 minute, pour off the waste liquid from the collection tube, and return the column to the collection tube.5. Centrifuge at 12,000 rpm for 2 minutes and pour off the waste liquid in the collection tube. Place the adsorption column at room temperature for 2 minutes and allow to dry.6. Place the column in a new collection tube (RNase-Free Centrifuge Tube), add 40-100 µL of RNase-Free Water to the center of the column membrane, let it stand at room temperature for 2 minutes, and then centrifuge at 12,000 rpm for 1 minute to collect the nucleic acid solution. Store at -80℃ for a long time... Read More |