In this article, I will briefly discuss some important factors to take into consideration when choosing your buffer and then discuss how to prepare the most commonly used buffers in life science. First and foremost, the pH range of the agents you plan to use in your experiment needs to coincide with the pH range of the buffer you choose. You can determine if they coincide by looking at the pH range of the buffer or by actually comparing the pKa of your buffer to the pH you intend to keep your experiment at.
The pKa value should be within one unit of your desired pH. Additionally, you must take into consideration concentration, buffer toxicity, temperature, and reactivity. The concentration of your buffer must be sufficient to account for the amount of acid or base you plan to use in your experiment. The more of the pH-altering component that you plan to use, the higher the buffer concentration you will need. Buffer toxicity is important because some buffers can be toxic to the cells you are working with, so if you are unsure about toxicity you can test the buffer out on your cells before you use it in your experiment.
You should also ensure that the buffer you decide to use is not capable of producing unwanted reactions in your experiment. Last, but not least, check that the temperature you plan to use in your experiment works with your buffer of choice because temperature can alter the buffering capacity of your buffer. For more information on choosing the right buffer for your experiment, refer to our user guide or our short article to help determine what is most appropriate.
Bis-Tris is a member of the Bis 2-hydroxyethyl amine family of buffers. You can use this buffer to resist pH changes in experiments where pressure is being varied, in gel electrophoresis experiments, and during anion exchange chromatography and NMR. The pH range of this buffer is 5. To prepare 1 liter of 1M Bis-Tris buffer stock solution, dissolve Adjust to desired pH using concentrated hydrochloric acid. Fill to a final volume of 1L with dH 2 O and sterilize by filter or autoclave. Store buffer at 4 degrees Celsius.
When working with a Bis-Tris buffer solution, remember that Bis-Tris forms complexes with lead and copper and various other metals. In addition to the uses mentioned above, Bis-Tris can be substituted as a safer alternative for cacodylate, which is a toxic buffering agent.
This buffer can be used in cell culture media for a variety of organisms. It can also be used as a binding buffer in protein studies, used in cation exchange elution experiments, and in gel electrophoresis as a running buffer. Adjust to desired pH using 10N sodium hydroxide. HEPES is soluble in water, inexpensive, and is typically biologically inert. However, it does participate in oxidation-reduction reactions yielding free radicals and interferes with reactions between DNA and restriction enzymes, but to a lesser extent than Tris due to steric hindrance.
It is also important to note that HEPES should not be used when working with heteromeric connexin channels as it can inhibit their function. MES is a buffer of the morpholinic family. This buffer is used in solutions that contain metal ions because it does not form complexes with most metals.
MES is typically used in buffered culture media for yeast, bacteria and mammalian cells, but it is toxic to plants at concentrations higher than 10 mM. MES is also a great choice of buffer for a variety of chromatography and electrophoresis experiments because it displays low ionic mobility and conductivity. It is an effective buffer in pH ranges from 5. X-gal 5-bromochloroindolyl-b-D-galactoside same recipe for X-phosphate. Use a glass or polypropylene tube.
It is not necessary to sterilize X-gal solutions. This Web page is maintained by Julie B. Useful links: Looking for career advice? Prepare a 5x stock solution in 1 liter of H2O: 54 g of Tris base Cover with parafilm and mix thoroughly. Remove 50 ml and set aside for the last two washes. To the remainder, add I-Block to 0. Heat to 68 C, and stir with a magnetic stirrer to assist dissolution.
If necessary, adjust the pH to 7. Store at room temperature. Sterilization is not necessary. Do not autoclave. Sodium Acetate 3 M, pH 5. Adjust the volume to ml with H2O. Minneapolis, MN Skip to main content. Antibiotics 6X Gel Loading Buffer 0.
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