| Description | Product Introduction:Blood samples are an important source of biological information in clinical research. They contain circulating proteins from multiple tissues and organs, which are involved in a wide range of biological processes and can be used as biomarkers or drug targets. However, blood Product Introduction:Blood samples are an important source of biological information in clinical research. They contain circulating proteins from multiple tissues and organs, which are involved in a wide range of biological processes and can be used as biomarkers or drug targets. However, blood samples have problems such as complex types of proteins, a large dynamic range, and a high proportion of high-abundance proteins, which bring great difficulties to proteome detection. This kit enriches circulating proteins in plasma samples based on the immunomagnetic bead capture method. In downstream proteomics detection, it can reduce the coverage of low-abundance protein signals by high-abundance proteins, thereby increasing the signal intensity of low-abundance proteins and improving the number of identified proteins and the repeatability of quantification.Product Components and Storage Conditions:M1456358Component12T24T48TStorageM1456358AMagnetic beads Max225 µL450 µL900 µL4℃M1456358BIncubation Buffer I6.25 mL12.5 mL25 mLRTM1456358CIncubation Buffer II7.5 mL15 mL30 mLRTM1456358DWashing Buffer7.5 mL15 mL30 mLRTOperating Procedure:1.Centrifuge the plasma sample (3000g, 10min), and take the supernatant for later use (if storage is needed, store it at -80°C for long-term preservation to avoid repeated freezing and thawing).2.Take 50-100µL of centrifuged plasma, add 400µL of Incubation buffer I, then add 18µL of Magnetic beads, vortex to mix, and incubate at room temperature on a shaking mixer for 1h.3.After incubation, use a magnetic rack to magnetically attract for 3min, and discard the supernatant.4.Remove the EP tube, add 500µL of Incubation buffer II, gently invert up and down to mix several times, magnetically attract for 3min, and discard the supernatant.5.Remove the EP tube, add 500µL of Washing Buffer, gently invert up and down to mix several times, magnetically attract for 3min, and discard the supernatant.6.Add 50µL of Protein Lysis Buffer (P1408622) to the beads obtained in step 5.7.It can be processed according to the conventional proteomics pretreatment steps in the laboratory, or our Proteomics Pretreatment Kit (P1456469) (to be purchased separately) can be used for processing.Precautions:1.Magnetic beads will precipitate after standing. Please shake gently and thoroughly before each use to keep the magnetic beads in a uniform suspension state.2.During the storage and use of magnetic beads, operations such as freezing, drying and high-speed centrifugation should be avoided, as they may damage the structure of magnetic beads and affect their protein-binding ability.3.This product is limited to scientific research use by professionals, and must not be used for clinical diagnosis or treatment, nor for food or drugs... Read More | H665581 Component 100 T Storage H665581A gDNA Eraser 50 µL -20℃. Avoid freeze/thaw cycle. H665581B 10×gDNA Eraser Buffer 120 µL -20℃. Avoid freeze/thaw cycle. H665581C HiFiScript, 200 U/µL 100 µL -20℃. Avoid freeze/thaw cycle. H665581D 5×ScriptRT H665581 Component 100 T Storage H665581A gDNA Eraser 50 µL -20℃. Avoid freeze/thaw cycle. H665581B 10×gDNA Eraser Buffer 120 µL -20℃. Avoid freeze/thaw cycle. H665581C HiFiScript, 200 U/µL 100 µL -20℃. Avoid freeze/thaw cycle. H665581D 5×ScriptRT Buffer 500 µL -20℃. Avoid freeze/thaw cycle. H665581E Primer Mix 120 µL -20℃. Avoid freeze/thaw cycle. H665581F RNase-Free Water 2×1 mL -20℃. Avoid freeze/thaw cycle.Product IntroductionThis product is a kit for removing genomic DNA for reverse transcription. The kit removes genomic DNA in 2 minutes at 42°C. Since the reverse transcription reagent contains a component that inhibits gDNA Eraser, cDNA can be synthesized directly by reverse transcription of gDNA Eraser-treated samples.The kit is equipped with a new high-efficiency reverse transcription enzyme, HiFiScript, with novel mutation sites that dramatically increase the transcriptional activity of the enzyme, resulting in higher efficiency and yield of cDNA first-strand synthesis. The first strand of cDNA can be synthesized with higher efficiency and yield, and the first strand of cDNA can be synthesized from pg total RNA or mRNA. If the reverse transcription product cDNA is used for downstream fluorescence quantitative detection, the reverse transcription reaction can be completed at 42℃ in 15 minutes. This kit is suitable for the synthesis of first-strand cDNA and subsequent RT-PCR, RT-qPCR, and the construction of full-length cDNA libraries.Product Features1. Rapid genome removal: contains gDNA Eraser for genomic DNA removal, which removes genomic DNA in just 2 minutes.2. Rapid reverse transcription: 15 minutes to obtain fluorescent quantitative PCR template cDNA first strand synthesis.3. High sensitivity: cDNA first strand can be synthesized using pg-level total RNA or mRNA templates.4. Highly efficient reverse transcription: Novel mutation sites dramatically increase enzyme activity, resulting in higher yields of cDNA.matters needing attention1. During operation, RNase contamination should be avoided to prevent RNA degradation or cross-contamination in the experiment. It is recommended that operators wear masks and disposable gloves and change the gloves frequently, and use specialized instruments and consumables.2. The reverse transcription system is prepared and operated on ice to prevent degradation of RNA. Store the kit enzymes at -20ºC as soon as possible after use and try to avoid repeated freezing and thawing.3. The reaction system can be scaled up to a maximum of 1 µg of total RNA in 10 µl of reaction system.4. Primer Mix is prepared by Oligo(dT) and Random primer, and Oligo-dT Primer or Gene Specific Primer can also be used according to the experimental needs.5. If the amount of starting RNA is less than 50ng, it is recommended to add RNAase inhibitor (RNasin).6. For RNA templates with complex secondary structures, it is recommended to incubate the template RNA at 65°C for 5 minutes immediately on ice prior to the manipulation step and centrifuge briefly before proceeding to the next step.UsageThaw template RNA on ice; place kit components on ice immediately after thawing at room temperature. Each solution was mixed by vortexing and shaking before use and briefly centrifuged.I. Genomic DNA removal reactions1. Prepare the reaction system according to the following table on ice in a total volume of 10 µl. To ensure the accuracy of the reaction solution preparation, prepare the premixed system in the amount of reaction number + 2 before dispensing it into each reaction tube and finally adding the RNA sample.Note: 1) If the amount of total RNA is greater than 1µg, scale up the reaction system proportionally. If the amount of starting RNA is less than 50ng, it is recommended to add RNAase inhibitor (RNasin).2. Mix by vortex shaking and centrifuge briefly so that the solution on the walls of the tube collects at the bottom.3. Incubate at 42°C for 2 minutes (this can be extended to 30 minutes for room temperature reactions).4．At the end of the reaction, centrifuge briefly and place on ice to cool.II. Reverse transcription reaction1. Prepare the reaction system on ice according to the following table. In order to ensure the accuracy of the reaction solution configuration, first prepare a premixed solution in the amount of number + 2, and then dispense 10 µl into each reaction tube, take 10 µl of the prepared premixed solution and add it to the reaction tube of step 1 where the de-etching of the genome has been completed.Note: 1) Oligo-dT Primer or Gene Specific Primer can be used according to the needs of the experiment, it is recommended to use 50 pmol of Oligo-dT Primer or 2 pmol of Gene Specific Primer for 20 µl reaction system.2. Mix well and centrifuge briefly so that the solution on the walls of the tube collects at the bottom.3. cDNA synthesis reaction conditions:1) If fluorescent quantitative PCR assay is performed downstream, incubate at 42°C for 15 minutes and 85°C for 5 minutes.2) If downstream for normal PCR assay, incubate at 42°C for 30-50 minutes and 85°C for 5 minutes. Note: For templates with complex secondary structure or high GC content, the reverse transcription temperature can be increased to 50°C to enhance reverse transcription efficiency.4. At the end of the reaction, centrifuge briefly and place on ice before proceeding with subsequent PCR or fluorescence quantitative PCR, or place at -20°C if prolonged storage is required.Note: When performing Real-time PCR reactions, the amount of reverse transcription product added should not exceed 1/10 of the total volume of the PCR reaction... Read More | Product introduction：Used to isolate lymphocytes from human organsMatters needing attention：1. samples, reagents and experimental environment in the whole process shall be carried out at 20 ± 2 ℃. In order to obtain the best experimental results, it is best to carry out the Product introduction：Used to isolate lymphocytes from human organsMatters needing attention：1. samples, reagents and experimental environment in the whole process shall be carried out at 20 ± 2 ℃. In order to obtain the best experimental results, it is best to carry out the experiment within 2 h of sampling. The longer the sample is stored, the worse the cell separation effect is. The separation effect is even worse after the sample is placed for more than 6 h, or even cannot achieve the purpose of separation. 2. in this experiment, it is better not to use plastic products with high polymerization materials (such as polystyrene), but use non-static, low static ionization heart tubes and glass products without alkali treatment, because the electrostatic effect will lead to cell adhesion, and the surface of alkali treated glass will become rough, which will affect the effect of cell separation. 3. aspirating too many lymphocyte layers and separation liquid layers will cause the granulocytes at the junction of separation liquid to be aspirated, thus increasing the number of mixed granulocytes. 4. when the amount of separating solution is greater than that of tissue single cell suspension sample, the separation effect is better.Scope of application：Lymphocyte isolation... Read More | Apoptosis refers to the cell autonomous and orderly death controlled by genes to maintain the stability of the internal environment. Apoptosis is different from cell necrosis. Apoptosis generally refers to a programmed cell death process that occurs during the development of body cells or under the Apoptosis refers to the cell autonomous and orderly death controlled by genes to maintain the stability of the internal environment. Apoptosis is different from cell necrosis. Apoptosis generally refers to a programmed cell death process that occurs during the development of body cells or under the action of some factors through the regulation of intracellular genes and their products. Cell necrosis is a cell death process that is caused by strong physical and chemical or biological factors to cause disordered changes in cells. The difference between apoptosis and necrosis lies in the characteristic morphological and biochemical changes, including the changes of cell membrane permeability and nuclear chromatin, the contraction of cytoplasm and the loss of membrane asymmetry. The oxazole yellow/pi membrane permeability apoptosis detection kit produced by our company is a dual fluorescence detection kit based on oxazole yellow and PI dyes. This kit is suitable for fluorescence microscopy, flow cytometry, fluorescence microplate reader and other fluorescence detection systems. Oxazole yellow is a non cell membrane penetrating cyanine monomer green fluorescent dye with high affinity for DNA. It basically has no fluorescence when it is not bound to DNA, but can emit bright green fluorescence after binding to DNA. When apoptosis occurs, the permeability of cell membrane changes. At this time, oxazole yellow can enter the cell and bind to DNA, emitting bright green fluorescence. Therefore, it is often used for the detection of apoptosis. It should be noted that oxazole yellow can also stain dead cells, so it needs to be double stained with PI that specifically fluorescently stains dead cells to effectively determine apoptosis. PI (propidium iodide) is a red fluorescent dye that can stain DNA. It is an analog of pyridine bromide that releases red fluorescence after embedding double stranded DNA. Although PI cannot pass through the membrane of living cells, it can cross the damaged cell membrane of dead cells to stain nuclei. Therefore, oxazole yellow combined with PI can be directly used for the detection of apoptosis. Apoptotic cells show green fluorescence, dead cells show both red and green fluorescence positive, and living cells have little or no fluorescence.Components: Components O598364-50T A. Oxazole yellow dye 50 µL B. Propidium Iodide (PI) 50 µLUsage (using flow cytometry as an example):1. Cell preparation(1) For adherent cells, after trypsin digestion, resuspend in culture medium and wash once with pre cooled PBS; The digestion time of trypsin should not be too long to prevent false positives. Note: Digest with trypsin and allow the cells to recover in the optimal cell culture conditions and medium for about 30 minutes, then stain.(2) For suspended cells, centrifuge at 1000 rpm for 5 minutes, discard the supernatant, and wash once with pre cooled PBS.2. Cell stainingSuspend cells in pre cooled PBS, with a recommended cell count of 106 cells/mL per sample. Add 1 µ L Oxazole Yellow and 1 µ L to 1 mL of the samplePI， Gently blow and mix well. Incubate on ice in the dark for 30 minutes. Note: We suggest adding the following two experimental controls:Blank tube: negative control group cells, without dye, used to regulate voltage.Single staining tube: Positive control group cells were treated with only two tubes, Oxazole yellow and PI, for regulating compensation.3. Flow detectionAfter incubation, the sample can be directly detected by flow cytometry, or centrifuged at 1000 rpm for 5 minutes, the supernatant can be aspirated, and the sample can be resuspended in 1 mL of pre cooled PBS for flow cytometry detection. Oxazole yellow can be excited by a 488 nm laser, and the detected fluorescence emission spectrum is around 530 ± 30 nm (FITC channel), while the PI channel emission spectrum is around 617 nm (PI or PE channel).Product parameters：Oxazole yellow dye:ex/em = 491 / 509 nm (bound DNA); Propidium iodine:ex/em = 535 / 617 nm (combined with DMatters needing attention：1. please centrifuge the product to the bottom of the tube immediately before use, and then conduct subsequent experiments. 2. fluorescent dyes have quenching problems. Please try to avoid light to slow down fluorescence quenching. 3. for your safety and health, please wear experimental clothes and disposable gloves.Scope of application：Membrane permeability apoptosis assay... Read More | Product DescriptionAcetyl esterase (sialate-O-acetylesterase) is a recombinant protein from Tannerella forsythia, ATCC 43037 strain, expressed in Escherichia coli. The enzyme removes acetyl groups attached via an O- group, mainly 9-, 8- and 7-. It can be used for monitoring of diacetylation of Product DescriptionAcetyl esterase (sialate-O-acetylesterase) is a recombinant protein from Tannerella forsythia, ATCC 43037 strain, expressed in Escherichia coli. The enzyme removes acetyl groups attached via an O- group, mainly 9-, 8- and 7-. It can be used for monitoring of diacetylation of sialic acids on products such as erythropoietin (EPO).The Zyme Acetyl Esterase Kit removes 9-, 8- and 7-O-acetyl groups from released sialic acids, released glycans or glycoproteins. It is commonly used for the characterization of highly-sialylated biotherapeutics such as EPO, FSH and blood clotting factors.Molecular Weight76.3 kDContentsAcetyl esterase – PBS pH7.5 buffer containing 10 mM Tris-HClReaction Buffer – 500 mM sodium acetate pH5.5Number of SamplesSufficient for up to 50 samples.Amount of SampleUp to 10 µg glycoprotein, up to 2.5 µg released glycans and up to 1 µg free sialic acid per digestion.Suitable SamplesAcetyl esterase (sialate-O-acetylesterase) can act upon complex glycoprotein samples, such as erythropoietin (EPO), bovine submaxillary mucin and oral epithelial cell-bound glycans, and on N- and O-glycans released from a glycoprotein. Either fluorescently labelled or unlabelled glycans are suitable. It can also be used on released sialic acids.Unit DefinitionOne unit (U) of acetyl esterase is defined as the amount of enzyme required to produce 300 µmole of 4-nitrophenol and acetate in 1 minute at 30°C in a buffer containing 50 mM Tris-HCl, 140 mM NaCl, pH 8.5, from 4-nitrophenyl acetate, a chromogenic esterase substrateStorageProtect from sources of heat and light. When stored correctly, the enzyme should be stable for 24 months from date of purchase. Exposure to ambient temperatures (20 – 26°C) over 3 days does not result in a reduction of enzymatic activity.ShippingThe product should be shipped at 4°C.HandlingEnsure that any glass, plastic ware or solvents used with this item are free of environmental carbohydrates. Use powder-free gloves for all sample handling procedures and avoid contamination with environmental carbohydrate.SafetyPlease read the Safety Data Sheets (SDSs) for all chemicals used. All processes involving labelling reagents should be performed using appropriate personal safety protection – safety glasses, chemically resistant gloves (e.g. nitrile), lab coat, and when appropriate, in a laboratory fume cupboard.For research use only. Not for human or drug use ApplicationAcetyl esterase (sialate-O-acetylesterase) can be used to remove 9-, 8- and 7-O-acetyl groups from released sialic acids, released glycans or glycoproteins... Read More |