| Description | BCIP/NBT (5-bromo-4-chloro-3-indolyl phosphate/ Nitro Blue Tetrazolium) liquid ready-to-use substrate is a highly active and stable formulation utilized for colorimetric detection of Alkaline Phosphatase (AP) activity in membrane assays. Positive reactions form an intense blue/purple precipitate at BCIP/NBT (5-bromo-4-chloro-3-indolyl phosphate/ Nitro Blue Tetrazolium) liquid ready-to-use substrate is a highly active and stable formulation utilized for colorimetric detection of Alkaline Phosphatase (AP) activity in membrane assays. Positive reactions form an intense blue/purple precipitate at the site of the reaction. The color develops when AP catalyzes the dephosphorylation of BCIP and converts NBT to insoluble blue/purple NBT formazan. The intense blue/purple precipitate is very stable and resists fading when exposed to light.Product Characteristics BCIP/NBT (5-bromo-4-chloro-3-indolyl phosphate/ Nitro Blue Tetrazolium) liquid, ready-to-use substrate is a highly active and stable formulation utilized for colorimetric detection of Alkaline Phosphatase (AP) activity in membrane assays. Positive reactions form an intense blue/purple precipitate at the site of the reaction. The color develops when AP catalyzes the dephosphorylation of BCIP and converts NBT to insoluble blue/purple NBT formazan. The intense blue/purple precipitate is very stable and resists fading when exposed to light.Composition & Properties The BCIP/NBT is a liquid, ready-to-use substrate. The solution contains NBT, BCIP and non-toxic stabilizers in an aqueous buffer, pH 9.6. The formulation is free of organic solvents.Working Procedure 1.Allow the solution to reach room temperature prior to use. 2.After the final incubation with the AP-labelled probe, wash the membrane thoroughly in a Tris-Buffered Saline (TBS Buffer) containing 0.1% Tween 20. 3.After the final wash, completely cover the membrane with BCIP/NBT solution and allow the color reaction to develop until optimal signal intensity is reached (usually 5-30 minutes). Incubation time will depend on enzyme activity. A further adjustment of the dilution of the AP probe may be necessary. The formazan deposit may flake off the membrane or background staining of the membrane may occur if the color development is too fast or too slow, respectively. 4.Stop the enzyme reaction by thoroughly washing membrane with deionized water. 5.Dry the membrane and store at room temperature. It is recommended to protect the membrane from light for a permanent record of results.Tips & Tricks Normal appearance of the solution is clear, pale yellow. Discard if solution is turbid or purple. Crystalline precipitate can occur in the product. Do not use phosphate buffers, as inorganic phosphate is a powerful inhibitor of AP. The products are not recommended for immunohistochemical or in situ hybridization techniques.Handling & Storage Store solution at 2-8 °C in the dark. Avoid exposure to light and heat. Re-dispense only into bottles made of High Density Polyethylene (HDPE), amber color. Dispensing guidelines are available upon request... Read More | Inorganic pyrophosphates are inevitably produced in the process of mRNA transcription in vitro. These substances have a great inhibitory effect on transcription. Inorganic pyrophosphatase (PPase) can hydrolyze the inorganic pyrophosphates produced in nucleic acid amplification experiments, promote Inorganic pyrophosphates are inevitably produced in the process of mRNA transcription in vitro. These substances have a great inhibitory effect on transcription. Inorganic pyrophosphatase (PPase) can hydrolyze the inorganic pyrophosphates produced in nucleic acid amplification experiments, promote the shift of reaction equilibrium to the product generation end, and increase the amount of products.The molecular weight of PPase (pyrophosphatase, inorganic, inorganic pyrophosphatase) is about 63kd, which can catalyze the hydrolysis of inorganic pyrophosphate to produce orthophosphate: P2O74_+H2O+PPase→2HPO42_. In the nucleic acid amplification experiment, PPase can hydrolyze the inorganic pyrophosphate generated with the reaction to avoid its inhibition on the reaction system. The removal of pyrophosphate can shift the reaction equilibrium to the product generation end.This product is a GMP level recombinant inorganic pyrophosphatase (yeast source) expressed by large-scale fermentation of E. coli. It is produced with raw and auxiliary materials of medicinal specifications, and the host protein residue and nucleic acid residue are strictly controlled. The product production and quality management procedures in line with GMP specifications ensure that the production process and all raw and auxiliary materials can be traced.Quality requirements project standard appearance Clear liquid Visible foreign matter Compliance with regulations PH value 7.5±8.5 activity 98U/ml-102U/ml purity ≥95% Endonuclease residues Degradation of substrate shall not exceed 10% Exonuclease residues Degradation of substrate shall not exceed 10% RNase residue Degradation of substrate shall not exceed 10% Bacterial endotoxin content ≤10EU/ml Exogenous DNA residue ≤100pg/mg Host protein residue ≤50ppm Mycoplasma detection negative Heavy metal residues ≤10ppm Follow the following specifications1. ISO 9001:2015, certified facility。2. GMP appendix - cell therapy products State Drug Administration.3. general introduction to human gene therapy - Chinese Pharmacopoeia 2020, National Pharmacopoeia Committee.4. USP chapter <1043>, adjuvant materials for cell, gene, and tissue engineered products.5. USP chapter <92>, growth factors and cytokines used in cell therapy manufacturing.6. Ph. Eur. General chapter 5.2.12, raw materials of biological origin for the production of cell-based and gene therapy medical products.Product features1. hydrolyze inorganic pyrophosphate.2. DNA synthesis: significantly enhance DNA replication ability.3. RNA synthesis: increase RNA production in in vitro transcription reaction.4. The optimal reaction temperature is 25℃, and the enzyme can be inactivated at 65℃ for 10min.Product usage1. optimize RNA transcription: improve the RNA yield of in vitro transcription reaction.2. remove PPI contamination from reagents for SNP genotyping by pyrophosphate assay.3. promote the synthesis of protein, RNA and DNA.4. catalyze the reaction of PPI + H2O → 2pi.5. ssr-pcr optimization：Improve efficiency and increase DNA production.Activity definitionCatalytic inorganic pyrophosphate formation 1 per minute under standard reaction conditions µ The amount of enzyme required for mol phosphate was defined as 1 active unit.Preservation system20 mM Tris-HCl; 100 mM NaCl; 1 mM DTT; 0.1 mM EDTA; 50% (v/v) Glycerol; pH 8.0。 Storage temperature-20±5 ℃。Matters needing attention1. the enzyme has activity in various reaction buffers. Generally, the enzyme can be directly added in HDA, lamp and other experiments.2. the dosage of the enzyme needs to be optimized in different experiments, usually adjusted at the concentration of 0.05~1u/ml.3. the optimum reaction temperature of the enzyme was 25 ℃, and it was active at 16~37 ℃, and the enzyme could be inactivated at 65 ℃ for 10min.4. cofactor: mg2+ is necessary for enzyme activity... Read More | Purity≥95% SDS-PAGE. Recombinant human MIF, fused to His-tag at N-terminus, was cloned into an E. coli expression vector and was purified to apparent homogeneity by using conventional column chromatography techniques.FunctionPro-inflammatory cytokine. Involved in the innate immune response to Purity≥95% SDS-PAGE. Recombinant human MIF, fused to His-tag at N-terminus, was cloned into an E. coli expression vector and was purified to apparent homogeneity by using conventional column chromatography techniques.FunctionPro-inflammatory cytokine. Involved in the innate immune response to bacterial pathogens. The expression of MIF at sites of inflammation suggests a role as mediator in regulating the function of macrophages in host defense. Counteracts the anti-inflammatory activity of glucocorticoids. Has phenylpyruvate tautomerase and dopachrome tautomerase activity (in vitro), but the physiological substrate is not known. It is not clear whether the tautomerase activity has any physiological relevance, and whether it is important for cytokine activity... Read More | Purity>97% by SDS-PAGE and HPLC analyses.Additional sequence informationN-terminal Glycine.FunctionChemotactic for monocytes and T-lymphocytes. Binds to CXCR3.Post-translationalCXCL10(1-73) is produced by proteolytic cleavage after secretion from keratinocytes | Inquire |