Is there any one who knows how a passive reference dye gives flourescence during real time PCR run?
472148
I have recently wondered the answer to this question myself.  I think that passive dyes, such as ROX, constantly emit a flourescent signal.  This signal is not supposed to be influenced by the PCR reaction cycle (ie "passive").  I believe the ROX dye is a rhodamine labeled amino acid, or maybe a short peptide. Other passive dyes are probably similar.
737939
Yes. It produces a bright yellow precipate:\nAgNO3 (aq) + KI (aq) → KNO3 (aq) + AgI (ppt)
1131203
G.W. Jr. Shipps, U.P. Spitz, J. Jr. Rebek, Solution-Phase Generation of Tetraurea Libraries, Bioorganic & Medicinal Chemistry \nfound at the site... \nhttp://www1.elsevier.com/cdweb/journals/09680896/viewer.htt?viewtype=keywords&rangeselected=32\n\nlooks like fun stuff :) don't inhale
827602
You're doing the PCR reaction to look for an amplification of a DNA segment.  When you run it out on a gel and look for the presence of bands of DNA, water will be your negative control because there is no DNA.  You want to do this to have something to compare your amplified sequence with; you also want to do this to make sure that bands in your amplified lanes are due to PCR and not to something DNA that was present in the water.
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AgNO3 + NaCl => AgCl(white ppt) + NaNO3\nAgNO3 + KI => AgI( yellow ppt) + KNO3\n\nthere is a change of color because AgI is produced
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temperature changes de amount of energy in molecules, which has a direct efect on the amount of light transmited through a  solution and, therefore, afects absorbance.  Also, in chemical analysis, the formation of colored compunds is necesary and if the correct temperature is not set, the compound doesn't form correctly. (just some reaction mechanisms from organic chemistry)\nThe AOAC method for testing nitrate content in water is a good example. If you don't give the reaction the correct temperature, the colored compound will never form and the test is invalid.
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Experimentally you can do primer extension, which will show the +1 site, from there you can identify the RBS and -10 and -35 sequences.\n\nYou can also do mobility shift assays with RNA polymerase to show that a certain piece of DNA contains the promoter, then do DNAse protection assays to show where the polymerase is actually binding.
210520
When the ink is sprayed onto a porous material the water in the ink reacts with carbon dioxide in the air to form carbonic acid. The carbonic acid then reacts with the sodium hydroxide in a neutralization reaction to form sodium carbonate. Neutralizaton of the base causes a color change of the indicator and the stain disappears:\n\nCarbon dioxide in the air reacts with water to form carbonic acid:\n\nCO2 + H2O -> H2CO3\n\nThe neutralization reaction is sodium hydroxide + carbonic acid -> sodium carbonate + water:\n\n2 Na(OH) + H2CO3 -> Na2CO3 + 2 H2O \n\nHere's how to make your own disappearing ink:\n\n   1. Dissolve the thymolphthalein (or phenolphthalein) in the ethyl alcohol.\n   2. Stir in 90 ml of water (will produce a milky solution).\n   3. Add sodium hydroxide solution dropwise until the solution turns a dark blue or red (might take slightly more or less than the number of drops stated in the Materials section).\n   4. Test the ink by applying it to fabric (cotton tee-shirt material or a table cloth works well). Paper allows less interaction with air, so the color change reaction takes more time.\n   5. In a few seconds, the 'stain' will disappear. The pH of the ink solution is 10-11, but after exposure to air will drop to 5-6. The damp spot will eventually dry    5. A white residue may be visible on dark fabrics.\n      Sponsored Links\n\n      AP Chemistry TutoringAccess to 30,000+ Discovery Videos For K-12. Sign up for 30-Day Trial!www.Cosmeo.com\n\n      Selenium ChemicalsSelenium dioxide and other Selenium Chemicals producer in the worldwww.vitalchem.com\n\n      Acids and Bases Homework?Chemistry Wizards for Your Homework Easy Step-by-Step Solutions Nowwww.molecularsoft.com\n      The residue will rinse out in the wash.\n   6. If you brush over the spot with a cotton ball that has been dampened in ammonia the color will return. Similarly, the color will vanish more quickly if you apply a cotton ball dampened with vinegar or if you blow on the spot to improve air circulation.\n   7. Leftover ink may be stored in a sealed container. All of the materials may be safely poured down the drain. \n\nDisappearing Ink Safety\n\n    * Never spray disappearing ink into a person's face. Particularly avoid getting the solution in the eyes.\n    * Preparing/handling the sodium hydroxide (lye) solution requires adult supervision, as the base is caustic. In case of skin contact, immediately rinse well with water. \n\nHere's what you need in order to make your own blue or red disappearing ink:\n\n    * 0.10 g thymolphthalein for blue ink or phenolphthalein for red ink (1/3 of 1/8 tsp)\n    * 10 ml (2 tsp) ethyl alcohol (ethanol) [can substitute 14 ml or 3 tsp of ethyl rubbing alcohol]\n    * 90 ml water\n    * 20 drops of 3M sodium hydroxide solution or 10 drops 6M sodium hydroxide solution [make a 3 M sodium hydroxide solution by dissolving 12 g of sodium hydroxide NaOH (1 level tablespoon of lye) in 100 ml (1/2 cup) of water.] \n\nLet's try it out...
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as far as i know its used for DNA figerprinting in electophoresis (i didnt spell that right).  basically you use a gel like substance, put DNA fragments in it (mixed w. dye) and put this in a solution and hook the one end of the whole device to a negative battery side or something and the other end to a positive side and the DNA fragments will seperate and u can use it to match up DNA samples w. people etc.
42811
http://www.uni-regensburg.de/Fakultaeten/nat_Fak_IV/Organische_Chemie/Didaktik/Keusch/D-Dyeing_vat-e.htm\n\nFound by simple web search
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I haven’t done any serious histology for over 10 years but…\nFirst off what do you mean by “without altering the mycelial growth”? Are you attempting to produce a live culture slide so that you can exmaine growth? If so then for God's sake don't use ethanol. Alcohol will certainly alter the mycelial growth. It will kill any growing mycelia. Alcohol also has a string tendency to cause shrinkage in soft tissues.\n\nDo you perhaps mean “without altering the mycelial structure”? That's a whole different story.\n\nLactophenol cotton blue is a combined stain and mounting fluid. The stain component (cotton blue) is designed to make the various cellular components contrast more so you can see them more easily. The lactophenol is just there as a mounting medium, not as a fixative. It serves approximately the same purpose as water on a wet slide but t lasts a little longer. Alcohol is a fixative. You can’t use it to replace the stain and it evaporates too fast to serve as a mounting medium. \n\nWhat do you want to use this slide for? How long do you need to store it?  I seem to recall, and a quick websearch seems to confirm, that lactic acid and acid fuschin does pretty much the same job as LCB. I’ll include one of the websites I turned up that suggest this. It also has a few other suggestions for mounting media/dye combinations that might suit you.\n\nSorry I can’t be more help. It’s probably because it’s been so long since I’ve done this, but I really don’t quite understand what you are trying to achieve.
1094870
We have used the pH-dependent fluorochrome fluorescein-dextran (FD) to study the acidification of prelysosomal vacuoles (endosomes) and lysosomes isolated from cultured macrophages and fibroblasts. FD was internalized by pinocytosis under conditions that allowed its selective localization in endosomes (1- to 5-min pulse) or in lysosomes (5-min pulse, 30-min chase). Fibroblasts were also exposed to FD at 20 degrees C, at which temperature endosome-lysosome fusion is inhibited. Cells were homogenized and labeled organelles were separated by centrifugation in Percoll density gradients. The addition of ATP rapidly decreased the internal pH of both endosomes and lysosomes, as indicated by a decrease in fluorescence intensity. The pH gradient was dissipated by H+ ionophores and ammonium chloride. Acidification was not affected by inhibitors of the mitochondrial F1,F0-ATPase or the Na+, K, K+-ATPase and did not require permeant anions, Na+, or K+. Of the inhibitors tested, only N-ethylmaleimide prevented the ATP-dependent acidification of both compartments. These findings provide direct support for the existence of an acidic prelysosomal compartment that may be acidified via the same type of H+ pump believed to operate in lysosomes and secretory granules.
222016
I am in a Microbiology college course. The closest I can come to your question from my text is about 50 NM (nanometer).
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I believe the difference is that in real-time PCR, when the first series runs, a second is started, where in Nested PCR, the primers, proteins etc are added a little at a time, making the process continuous, as opposed to in starts and stops.
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Fluorometry is the measurement of light emitted, for example, by a labeled sample that has been excited by light at a shorter wavelength. The greater the difference in wavelength between the excitation and emission light (this difference is known as the Stokes’ shift) the greater the potential for sensitive measurement of the fluorescence. \n\nTime-resolved fluorometry is possible when the fluorescent label has a long decay time. Light is collected at a time when non-specific fluorescence from the sample matrix and the microplate, etc. has died down. The elimination of interfering background in this way makes a further big contribution towards sensitivity. The DELFIA method exploits the unique fluorescence properties of lanthanide chelate labels. These labels have a long decay time as well as a wide Stokes’ shift.
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Methylene Blue is a homologue of Toluidine Blue O. This stain is used to visualize intracellular metachromatic granules. \n\nProcedure:\n\nSmears are fixed with heat and allowed to cool.\n\n\nStain with the methylene blue (homologue of toluidine blue O) solution two to seven minutes.\n\n\nWash slide and blot dry. \n \n\nResults: By this method, the intracellular metachromatic granules stain a ruby-red to black color; with the remainder of the cell staining a pale blue color. \n\n  \nhttp://www.life.umd.edu/classroom/bsci424/LabMaterialsMethods/MethyleneBlueStain.htm
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Paper chromatography is a modern method used separate mixtures. Paper chromatography uses paper as the stationary phase and a liquid solvent as the mobile phase.\n\nYou will use paper chromatography to test food colorings to see if the color results from a single dye or mixture of dyes. If you cannot get food coloring you can use colored felt pens or other water soluble coloring pens. Your exercise is simple but uses very essential principles. An optional part of the activity is to check a household product for the presence of the same dyes that are in food coloring.\n\nThe technique relies on the idea that the solvent and the paper both have an attraction for the components in a mixture. The solvent creeps along the surface of the paper. If a material is placed on one spot on the paper and is soluble in the liquid solvent, the material will be dissolved when the solvent moves over it. The material will move along with the solvent. Each compound in a mixture will have its own characteristic balance of attractions to solvent and to paper, so all will not move at the same speed. Eventually this difference in speed will separate the compounds.\n\nIn paper chromatography when the conditions are kept constant, a particular compound always travels a fixed percentage of the distance traveled by the solvent front. The ratio of the distance the compound travels to the distance the solvent travels is called the Rf value. The symbol Rf stands for "retardation factor" or "ratio-to-front". It is expressed as a decimal fraction. When the conditions are duplicated, the same average relative positions will turn up for the solvent and solute; thus the Rf value is a constant for a given compound. The Rf value is a physical property for that compound. The Rf value is useful in identifying compounds, but other properties should be used in combination with the Rf value to confirm compound identification. Since it is difficult for different laboratories to exactly duplicate conditions for a chromatography experiment, Rf values are more useful for comparisons within one lab than for comparisons of data from different labs.\n\nSubstances that are not colored can also be separated by chromatography. They are detected using ultraviolet or black light. These substances appear to glow in the dark.