| Description | Protein Labeling Kit (Aggregation-induced emission, AIE) is developed based on the principle of aggregation-induced emission (AIE) and is used for labeling general proteins. Its labeling principle is that the AIE dye in the kit carries active groups, which can react with the primary amino groups (Protein Labeling Kit (Aggregation-induced emission, AIE) is developed based on the principle of aggregation-induced emission (AIE) and is used for labeling general proteins. Its labeling principle is that the AIE dye in the kit carries active groups, which can react with the primary amino groups (lysine residues) on the protein surface to form stable amide bonds. The AIE dye has high reactivity and can quickly label general proteins. The optimal excitation wavelength of the AIE dye is 460 nm, and the emission wavelength range is 570–700 nm, which is suitable for a series of instruments equipped with a blue light excitation wavelength detector. Marking renderings:Notes:After receiving the product, customers should store it according to the temperature indicated on the product label.Customers need to prepare dimethyl sulfoxide (DMSO) reagents and disposable syringe consumables by themselves. Experimental Methods 1.Replacement of protein sample buffer solvent: Before starting to label the protein, ensure that the protein sample buffer is compatible with the kit. If your protein sample buffer contains one or more of primary amino groups (such as Tris and glycine), imidazole, DTT, and β-mercaptoethanol, please replace it with Buffer 1× buffer solution before labeling.2.Preparation of AIE dye stock solution: Add 20 µL of DMSO to the small tube containing AIE dye lyophilized powder, mix well to dissolve, and prepare a 10 mM AIE dye stock solution.Notes: 1) Before dissolving with DMSO, let the AIE dye lyophilized powder return to room temperature and centrifuge for 10 seconds. 2) After dissolution, it can be aliquoted for storage as needed.3.Protein labeling method (recommended) 1) Dilute the protein sample with Buffer 1× to a concentration of approximately 150 µg/mL. 2) Add 10 mM AIE dye stock solution to the above protein sample solution to make its final concentration in the protein sample solution 10 µM (for example, when the volume of the protein sample solution is 1 mL, add 1 µL of 10 mM AIE dye stock solution; the dosage can be adjusted according to the actual situation of the protein sample), and mix well. 3) Incubate at room temperature in the dark for 5–10 minutes. 4) After labeling, filter the labeled solution with a 0.1 µm needle filter to remove excess AIE dye.PrecautionsThe AIE dye is sensitive to humidity, so ensure that it is stored at -20°C in a dry and dark place. Before preparing the stock solution, let the dye return to room temperature and centrifuge for 10 seconds. After preparing the stock solution, aliquot it for storage and use it as soon as possible.The buffer solution (pH = 8.5) provided in the kit is the first choice for the AIE dye to work. You can choose any buffer solution that does not contain primary amino groups, imidazole, DTT, or β-mercaptoethanol as a substitute, but it may affect the labeling efficiency.It is recommended that the minimum concentration of the protein during labeling is not less than 15 µg/mL.Due to the differences in the number and reactivity of amino groups on the protein surface, the labeling effects of different proteins may vary. If the labeling effect is not good, the dye labeling concentration can be appropriately increased or the reaction time can be prolonged.For your safety and health, please wear a lab coat and disposable gloves during operation.This product is only for scientific research purposes... 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 | Glycogen and starch generate glucose-1-phosphate (1PG/G1P) during the process of phosphohydrolysis. This reagent kit provides a simple, sensitive, and rapid determination method: Glucose-1-phosphate (1PG/G1P) is reduced from NADP+to NADPH by the sequential action of phosphoglucose mutase and Glycogen and starch generate glucose-1-phosphate (1PG/G1P) during the process of phosphohydrolysis. This reagent kit provides a simple, sensitive, and rapid determination method: Glucose-1-phosphate (1PG/G1P) is reduced from NADP+to NADPH by the sequential action of phosphoglucose mutase and phosphoglucose dehydrogenase. The content of glucose-1-phosphate (1PG/G1P) in the sample can be calculated by detecting the increase in NADPH at 340nm.Composition and preparation of reagent kit: Reagent name Specifications Save requirements Remarks Extraction solution Liquid 100mL x 1 bottle 4 ℃ storage / Reagent 1 Powder mg x 1 tube 4 ℃ storage Shake or centrifuge the reagent a few times before use to make it fall to the bottom, then dissolve it in 1.2mL of distilled water for later use. Reagent 2 Powder mg x 1 tube Store at -20 ℃ Shake or centrifuge the reagent a few times before use to make it fall to the bottom, then dissolve it in 1.2mL of distilled water for later use. Reagent 3 Liquid 16mL x 1 bottle 4 ℃ storage / Reagent 4 Powder mg x 1 tube Store at -20 ℃ Shake or centrifuge the reagent a few times before use to make it fall to the bottom, then add 1 Dissolve 1mL of distilled water for later use. TRC 1 powder 4 ℃ storage Only used to identify whether the reagents in the kit are normal (not involved in result calculation). Usage: Use a pre standard tube (GIP) to shake the powder a few times until it falls to the bottom, then add 0.5mL of distilled water and mix well to dissolveDilute GIP with a concentration of 4mg/mL and then dilute it four times to 1mg/mL for later use: follow the instructions in the sample addition table for the measuring tube operationRequired instruments and supplies:ELISA reader, 96 well plate, desktop centrifuge, adjustable pipette, mortar, ice and distilled water.Determination of glucose-1-phosphate (1PG/G1P) content:1. Sample preparation① Organizational sample:Suggest weighing around 0 1g of tissue, add 1mL of extraction solution, and homogenize in an ice bath. Centrifuge at 12000rpm, 4 ℃ for 10 minutes, take the supernatant, and place it on ice for testing.[Note]: If the sample size is increased, it can be extracted in a ratio of tissue mass (g) to extraction solution volume (mL) of 1:5-10.② Bacterial/cellular samples:Collect bacteria or cells into a centrifuge tube first, centrifuge and discard the supernatant; Take about 5 million bacteria or cells and add them to 1mLExtract solution, sonicate bacteria or cells (ice bath, power 200W, sonication for 3s, interval 10s, repeated 30 times); Centrifuge at 12000rpm at 4 ℃ for 10 minutes, take the supernatant, and place it on ice for testing.[Note]: If the sample size is increased, extraction can be carried out in a ratio of 500-1000:1 of bacteria/cell quantity (104) to extraction solution (mL).③ Liquid sample: direct detection.2. Machine testing:① Preheat the enzyme-linked immunosorbent assay (ELISA) reader for at least 30 minutes and adjust the wavelength to 340nm.② Thaw the reagent to room temperature (25 ℃);③ Add reagents to the 96 well plate in the following order according to the table:② Thaw the reagent to room temperature (25 ℃);③ Add reagents to the 96 well plate in the following order according to the table: Reagent name (µL） Measurement tube Blank tube (only done once) Reagent 1 10 10 Reagent 2 10 10 Reagent 3 150 170 Sample 20 / Mix well, incubate at room temperature (25 ℃) for 20 minutes, and then read A1 at 340nm (if the A value continues to increase, the incubation time needs to be extended until the absorbance value remains unchanged within 2 minutes). Reagent 4 10 10 Mix well, incubate at room temperature (25 ℃) for 20 minutes, and then read A2 at 340nm (if the A value continues to increase, the incubation time needs to be extended until the absorbance value remains unchanged within 2 minutes). Δ A=(A2-A1) measurement - (A2-A1) blank.[Note] 1 If the difference in Δ A is hovering around zero, the sample size V1 can be increased (such as increasing to 50 µ L, the three phases of the reagent should be reduced while keeping the total volume unchanged), or the sample sampling mass W can be increased. The changed V1 and W need to be substituted into the formula for recalculation.If the A2 value exceeds 1.2, the amount of sample added V1 can be reduced (such as to 10 µ L, the three-phase reagent should be increased while keeping the total volume unchanged), or the sample can be diluted with distilled water (keeping the sample addition system unchanged), and the changed V1 and D need to be substituted into the formula for recalculation.Result calculation:1. Calculated by sample weight:1PG/G1P content (µ g/g fresh weight)=[(Δ A ÷ (ε× d) × V2 × 106 × MR] ÷ (W × V1 ÷ V) × D=836 × Δ A ÷ W × D2. Calculated by the number of cells:1PG/G1P content (µ g/104 cell)=[(Δ A ÷ (ε× d) × V2 × 106 × MR] ÷ (500 × V1 ÷ V) × D=1.7 × Δ A × D. 3. Calculated by liquid volume:1PG/G1P content (µ g/mL)=[(Δ A ÷ (ε× d) × V2 × 106 × Mr] ÷ V1=836 × Δ A ε---NADPH Molar extinction coefficient，6.22×103 L/mol/cm； d---96 Orifice plate optical diameter，0.5cm； V---Add volume of extraction solution，1 mL； V1---Add sample volume，0.02mL V2---Total reaction volume；0.2mL=2×10-4L； W---Sample quality，g； Mr---Glucose-1-phosphate（1PG/G1P）Molecular weight；260； 500---Number of cells, in millions； D---Dilution ratio，Undiluted is 1。 /... Read More | This kit is used to extract miRNAs from various animal tissues, plant tissues and cells. The extracted miRNA molecule is complete and high purity, which is suitable for various molecular biology experiments such as Northern blot, real timepcr, miRNA microarray chip, in situ hybridization, This kit is used to extract miRNAs from various animal tissues, plant tissues and cells. The extracted miRNA molecule is complete and high purity, which is suitable for various molecular biology experiments such as Northern blot, real timepcr, miRNA microarray chip, in situ hybridization, RNase protection assay, etcComposition:Scope of application：Nucleic acid extraction and purificationInstruction：1.Experimental preparation：1.1.All reagents were prepared with DEPC-treated solvents. Please use RNase-free tip and centrifuge tube to avoid RNA degradation by RNase during extraction.1.2.70 % ethanol, -20C pre-cooling.2.Operational procedure：There is a slight difference in the operation of miRNA extraction from different samples. The specific steps are as follows :【 Extraction of miRNA from animal tissues】1.Take 20-40 mg tissue, transfer to a pre-cooled mortar, and add liquid nitrogen to grind into powder.Please click below to describe the amount of organization used :①RNA-rich tissue ( e.g. liver ) : no more than 30 mg②Tissues with low RNA content ( e.g., muscle ) : no more than 100 mg③When the amount of tissue used was less than 20 mg : the amount of R-I, R-II and isopropanol used was halved.④When the amount of tissue used was more than 40 mg : the use of R-I, R-II and isopropanol increased proportionally.2.Add 400 ul Buffer R-I, repeatedly aspirate 8-10 times with a syringe equipped with a 21-25 needle, and transfer to a 1.5 m : centrifuge tube ( provided in the kit ). 3.Add 150 µl BufferR-1l, swirl for 15-30 s, centrifuge at 12,000 X g for 5 min. [ Centrifugation at 4 °C is recommended ] 4.Take the supernatant to 1.5ml centrifuge tube, add 180 u anhydrous ethanol, mix evenly.5.The preparation tube was placed in a 2 m : centrifuge tube ( provided in the kit ), the mixture in step 4 was transferred to the preparation tube, and 12,000 X g was centrifuged for 1 min. [ 1 Centrifugation at 4 °C is recommended ; 2 miRNA in the filtrate, pay attention to preserve the filtrate. ]6.Abandon the preparation tube, add 500µl isopropanol to the filtrate, and mix evenly.7.12,000Xg centrifuged for 10 min, discard the supernatant.8.Add 700µl 70 % ethanol ( pre-cooled at -20 °C ), centrifuged at 12,000Xg for 5min.9.The supernatant was discarded and dried at room temperature for 5-10 min.10.70 ul Buffer TE ( nucdease-free ) or RNase-free water was added to the centrifuge tube to elute miRNA.【 Extraction of miRNA from plant tissue 】1.Take 30-150 mg tissue, transfer to a pre-cooled mortar, and add liquid nitrogen to grind into powder.Please click below to describe the amount of organization used :①Plant leaves : usually 10-80 mg② Plant fiber tissue : usually 100-150 mg③When the amount of plant leaf tissue was less than 30 mg : the amount of R-I, R-II and isopropyl alcohol used was halved.④When the amount of plant leaf tissue was more than 80 mg : the use of R-I, R-II and isopropanol increased proportionally.⑤When the amount of plant fiber tissue was more than 150 mg : the use of R-I, R-II and isopropanol increased proportionally.2.Add 400 ul BufferR-I, use a syringe with a 21-25 needle to repeatedly suck 8-10 times, and transfer to a 1.5mI centrifuge tube ( provided in the kit ). 3.Add 150 ul Buffer R-1I, vortex oscillation 15-30 s, 12.000 x g centrifugation 5 min. [ Centrifugation at 4 °C is recommended ]4.Take the supernatant to 1.5ml centrifuge tube, add 180 mountain anhydrous ethanol, mix evenly.The preparation tube was placed in a 2 mI centrifuge tube ( provided in the kit ), the mixture in step 4 was transferred to the preparation tube, and 12.000 xg was centrifuged for 1 min. It is recommended to centrifuge at 4 °C ; 2 miRNA in the filtrate, pay attention to preserve the filtrate. ]Abandon the preparation tube, add 500µl isopropanol to the filtrate, and mix evenly.7.12,000xg high heart for 10 min, discard the supernatant.8.Add 700 ul 70 % ethanol ( -20 °C precooling ), 12,000 xg centrifuge for 5 min.9.The supernatant was discarded and dried at room temperature for 5-10 min.10.70 ul Buffer TE ( nucdease-free ) or RNase-free water was added to the centrifuge tube to elute miRNA.【miRNA extraction from cells】Steps 1-3 According to the different ways of cell culture, two experimental methods, a or b, can be selected.a. Suspension cultured animal cells or cell suspension obtained from petri dishes or culture flasks or freshly isolated animal tissue single cell suspension :1a.Collect 2X 10 * -1X 10 ' cells, centrifuge 2,000Xg for 5 min, discard the supernatant ;2a. Add 400 µl Buffer R-I, repeatedly draw 8-10 times with a syringe containing 21-25 needles, and transfer to a 1.5 mI centrifuge tube ( provided in the kit ) ;3a. Add 150µl Buffer R1I, vortex oscillation 15-30s, 12.000Xg centrifugal 5min. [ build at 4 °C centrifugal ].b. Cells cultured on 96-well L, 24-well, 12-well or 6-well plates :Cells were collected from 96-well, 24-well, 12-well or 6-well culture plates, and the medium was discarded as much as possible, and 400 u / well Buffer R-I was added to each well, and the pipette gun was used to blow up and down 8-10 times ;2b.Transfer the above cell suspension to a 1.5ml centrifuge tube ( provided in the kit ), and repeatedly draw 8-10 times with a syringe containing 21-25 needles ;3b. Add 150 µl Bufflr R-II, swirl for 15-30 s, centrifuge for 5 min at 12,000 × g. [ Recommended at 4 °C ]4.Take the supernatant to 1.5ml centrifuge tube, add 180 mountain anhydrous ethanol, mixing evenly.5.The preparation tube was placed in a 2 ml centrifuge tube ( provided in the kit ), the mixture in step 4 was transferred to the preparation tube, and centrifuged at 12.000 Xg for 1 min. [ 1 Centrifugation at 4 °C is recommended ; 2 miRNA in the filtrate, pay attention to preserve the filtrate. ]6.Abandon the preparation tube, add 500 u of isopropanol to the filtrate, and mix evenly.7.12,000Xg high heart for 10 min, discard the supernatant.8.Add 700µ70 % ethanol ( pre-cooled at − 20 °C ), centrifuged at 12,000 × g for 5 min.9.Abandon the supernatant, dry at room temperature for 5 - 10 min.10.70 ul Bufer TE ( nucdease-free ) or RNase-free water was added to the centrifuge tube to elute mRNA.3.Flow chartMatters needing attention：Buffer R-I contains irritating compounds, when operating to wear latex gloves and glasses, to avoid contamination of the skin, eyes and clothes, be careful not to inhale the nose and mouth. If the skin, eyes, to immediately rinse with a lot of water or saline, if necessary, seek medical advice... 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 |