| Description | Annexins are a family of calcium-dependent phospholipid-binding proteins that preferentially bind phosphatidylserine (PS). Under normal physiologic conditions, PS is predominantly located in the inner leaflet of the plasma membrane. Upon initiation of apoptosis, PS loses its asymmetric distribution Annexins are a family of calcium-dependent phospholipid-binding proteins that preferentially bind phosphatidylserine (PS). Under normal physiologic conditions, PS is predominantly located in the inner leaflet of the plasma membrane. Upon initiation of apoptosis, PS loses its asymmetric distribution across the phospholipid bilayer and is translocated to the extracellular membrane leaflet marking cells as targets of phagocytosis. Once on the outer surface of the membrane, PS can be detected by fluorescently labeled Annexin V in a calcium-dependent manner.In early-stage apoptosis, the plasma membrane excludes viability dyes such as propidium iodide (PI), 7-AAD. These cells will stain with Annexin V but not a viability dye, thus distinguishing cells in early apoptosis. However, in late stage apoptosis, the cell membrane loses integrity thereby allowing Annexin V to also access PS in the interior of the cell. A viability dye can be used to resolve these late-stage apoptotic and necrotic cells (Annexin V, viability dye-positive) from the early-stage apoptotic cells (Annexin V positive, viability dye-negative).We offer recombinant Annexin V conjugated to a numerous fluorophores, as well as an Annexin V biotin conjugate which can be detected with fluorophore-labeled streptavidin. By binding to PS, fluorophores labeled Annexin V can be used to detect and quantify apoptotic cells via flow cytometry or fluorescence microscopy. The excitation and emission maxima of the Annexin V conjugates are summarized in the following table. Cat.No.Ex/Em (nm)Formatrp226056NABiotinrp225999401/422AF405rp226057490/525AF488rp226060650/668AF647rp226002681/704AF680rp226003752/776AF750rp226053498/517FITCrp226004410/455Pacific Bluerp226006647/665Cy5rp226054650/660APCrp226055565/575PErp226058565/670PE-Cy5rp226059565/774PE-Cy7Precautions1. Please try to avoid light when using to slow down the quenching of fluorescence.2. Propidium Iodide Solution is toxigenic and mutagenic; handle with care.3. Due to the calcium dependence of the Annexin V:PS interaction, it is critical to avoid buffers containing EDTA or other calcium chelators during Annexin V experiments.Instruction for use1. Dilute 10X Binding Buffer (A1372288) to 1X using distilled water (1 mL 10X Binding Buffer + 9 mL ddH2O).2. Wash cells twice with cold PBS and then resuspend the desired amounts of cells in Annexin V Binding Buffer at a concentration of 1.0-5.0 x 106 cells/mL.3. Add 5 µL of Annexin V-AF488 to 100 µL of the cell suspension. Stain with a viability dye, such as PI (P1373641; P1372285), 7-AAD (A1372406), or DAPI (D1372407) dyes, if desired.4. Gently vortex the cells and incubate for 10 min at RT (25°C) in the dark.5. Add 100 µL of 1X Binding Buffer to each assay. Analyze by flow cytometry within 1 hr... Read More | Taq MasterMix is a premixed system composed of Taq DNA Polymerase, Mg2+, dNTPs, as well as PCR stabilizers and enhancers, with a concentration of 2 ×. The pre prepared PCR mixture makes the operation simpler and faster, and can minimize human error and contamination to the greatest extent Taq MasterMix is a premixed system composed of Taq DNA Polymerase, Mg2+, dNTPs, as well as PCR stabilizers and enhancers, with a concentration of 2 ×. The pre prepared PCR mixture makes the operation simpler and faster, and can minimize human error and contamination to the greatest extent possible. The original MasterMix formula results in high yield, strong repeatability, and good stability of amplified products. This product has been added with a dye (blue), and can be directly subjected to electrophoresis detection after the reaction is completed. The amplified PCR product has an "A" base attached to its 3 'end, making it suitable for direct use in T/A cloning. Mainly suitable for PCR amplification of DNA, DNA sequencing and other experiments. T665590Component5 mL25 mLStorageT665590A2×Taq MasterMix (Dye)5×1 mL5×5 mL-20℃. Avoid freeze/thaw cycle.T665590BddH₂O5×1 mL5×5 mL-20℃. Avoid freeze/thaw cycle.2×Taq MasterMix contains Taq DNA Polymerase, 3 mM Mg Cl₂ and 400 µM each dNTP. Quality control:After testing, there was no exogenous nuclease activity; PCR method for detecting residual DNA without host; Can effectively amplify single copy genes from multiple genomes.Usage:The following is an example of a PCR reaction system and reaction conditions for amplifying a 1 kb fragment using human genomic DNA as a template. In practical operation, corresponding improvements and optimizations should be made based on the template, primer structure, and target fragment size.1. PCR reaction system Reagent 50 µlReaction system Final concentration 2×Taq MasterMix（Dye） 25 µL 1× Forward Primer，10 µM 2 µL 0.4 µM Reverse Primer，10 µM 2 µl 0.4 µM Template DNA <0.5 µg <0.5 µg/50 µL ddH2O up to 50 µL /Attention: The primer concentration should be between 0.1 and 1.0 as the final concentration µ M serves as a reference for setting the range. In the case of low amplification efficiency, the concentration of primers can be increased; When non-specific reactions occur, the primer concentration can be reduced to optimize the reaction system.2. PCR reaction conditions Step Temperature Time / Pre denaturation 94℃ 2 min / Denaturation 94℃ 30 s 25-35 cycles Anneal 55-65℃ 30 s 25-35 cycles Extend 72℃ 30 s 25-35 cycles Finally extended 72℃ 2 min / Attention:1) In general experiments, if the annealing temperature is 5 ° C lower than the melting temperature Tm of the amplification primer, and the ideal amplification efficiency cannot be achieved, the annealing temperature should be appropriately reduced; When non-specific reactions occur, increase the annealing temperature to optimize the reaction conditions.2) The extension time should be set according to the size of the amplified fragment. The amplification efficiency of Taq DNA Polymerase in this product is 2 kb/min.3) The number of cycles can be set based on the downstream application of the amplification product. If the number of cycles is too small, the amplification amount is insufficient; If there are too many cycles, the probability of mismatches will increase, and non-specific backgrounds will be severe. So, while ensuring product yield, the number of cycles should be minimized as much as possible... Read More | Tyrosine decarboxylase catalyzes the removal of the carboxyl group from tyrosine to produce tyramine and carbon dioxide. Pyridoxal 5'-phosphate is a necessary cofactor. By using the apoenzyme prepared from cells grown on a vitamin B6 deficient medium pyridoxal phosphate may be determined. The Tyrosine decarboxylase catalyzes the removal of the carboxyl group from tyrosine to produce tyramine and carbon dioxide. Pyridoxal 5'-phosphate is a necessary cofactor. By using the apoenzyme prepared from cells grown on a vitamin B6 deficient medium pyridoxal phosphate may be determined. The HOLOenzyme may be used to determine tyrosine, phenylalanine and dihydroxyphenylalanine either manometrically or colorimetrically.L-Tyrosine decarboxylase apoenzyme from Streptococcus faecalis has been used in a study to purify and characterize tyrosine decarboxylase and aromatic-L-amino-acid decarboxylase.L-Tyrosine decarboxylase apoenzyme from Streptococcus faecalis has also been used in a study to investigate the stereospecificity of sodium borohydride reduction of tyrosine decarboxylase... Read More | Inquire | Product Characteristics UNI-StabilPLUS is a universal stabilizer for the dilution and stabilization of both Horseradish Peroxidase (HRP) and Alkaline Phosphatase (AP) labeled proteins and antibodies, in order to maintain the molecular conformation and prevent loss of activity over time. This enablesProduct Characteristics UNI-StabilPLUS is a universal stabilizer for the dilution and stabilization of both Horseradish Peroxidase (HRP) and Alkaline Phosphatase (AP) labeled proteins and antibodies, in order to maintain the molecular conformation and prevent loss of activity over time. This enables the making of pre-diluted, ready-to-use conjugates, minimizing assay errors in dilution. Superior stabilization of HRP and AP conjugated antibodies in low as well as high protein dilutions is seen, when using UNI-StabilPLUS. When tested with AP conjugated antibody stability is seen as follows: • at least 3 years at 2-8 °C • at least 2 years at room temperature • at least 4 weeks at 37 °C When tested with HRP conjugated antibody stability is seen as follows: • at least 2 years at 2-8 °C • at least 1 years at room temperature • at least 2 weeks at 37 °CUNI-StabilPLUS is recommended for the dilution of antibodies directed against rabbit immunoglobulins unlike HRP-StabilPLUS (cat. no. H494387) and Antibody Enhancer (cat. no. A494276).Composition & Properties UNI-StabilPLUS is a ready-to use buffer that appears as an opaque solution. The product is based on a mild acid Tris buffer containing proprietary stabilizing components. UNI-StabilPLUS contains neither BSA, nor other material from bovine serum, no azide, mercury or other toxic components.Working Procedure 1.Make a series of dilutions of the HRP- or AP conjugated protein in UNI-StabilPLUS in order to determine the optimal dilution. 2.Run the assay as usual or store the diluted conjugated protein preferably at 2-8 °C.Tips & Tricks • Avoid using phosphate buffers for AP-conjugated antibody assays. We recommend the use of Tris/HCl, Tween as the washing buffer, instead of a PBS buffer which will reduce signal significantly. • For extended stability of HRP conjugated antibodies, HRP-StabilPLUS (cat. no. H494387) is recommended. Handling & Storage • Store solution at 2-8 °C... Read More |