Does DTT affect pH?
DTT is an unusually strong reducing agent, with a redox potential of -0.33 V at pH 7. The pKa of thiol groups is typically ~8.3. Since protonated sulfurs have lowered nucleophilicities, DTT becomes less potent as the pH lowers.
What does DTT do in a buffer?
DTT is a reducing agent and usage will ensure that the protein is unfolded and soluble, easy to purify. Cytoplasmic proteins usually lack disulfide bonds. To keep the cysteine side chains in their normal reduced state, a reducing agent such as DTT is included in the purification.
What concentration is DTT?
DTT quantitatively reduces disulfide bonds and maintains monothiols in a reduced state (see Reference 1). At a final 0.1 M concentration, DTT is also widely used for disruption of protein disulfide bonds in SDS-polyacrylamide gel electrophoresis.
Does DTT dissolve in water?
This product is soluble in water (50 mg/ml), yielding a clear, colorless solution. DTT is also soluble in ethanol, acetone, ethylate, chloroform, and ether. DTT solutions should be prepared fresh daily.
Is DTT temperature sensitive?
Hi Mahesh, The nature of DTT is keep on reduction of the SH group in your protein about only few days(recommendable amount up to 5mM). During dialysis it shouldn’t affect your protein. That transfer time only my protein in room temperature.
How toxic is DTT?
The thiol-containing compound Dithiothreitol (DTT) has been shown to be toxic to cultured cells by inducing the generation of reactive oxygen species that ultimately cause cell death. However, its effects on multicellular organisms and the environment have not been investigated yet in detail.
Is DTT volatile?
Non-volatile. Effective at room temperature. When refrigerated 2-8ºC, it is stable up to 3 years.
What is DTT used for?
Dithiothreitol (DTT) is a redox reagent also known as Cleland’s reagent. It is used to break down protein disulfide bonds and stabilize enzymes and other proteins.
Can DTT go bad?
If DTT is kept at room temperature it will degrade/oxidize. The main role of DTT is to keep proteins in a reduced state. If the DTT was left out for more than 72 hours it is definitely too late. Also it is better to store DTT at -20C for long term storage.
How long does DTT last at 4c?
Before bottle opening , the reagent remains stable, protect from light, for 12 months at 2-8 °C, while, after opening it, the stability not is beyond 30 days at 2-8°C, always from light protect.
How do I get rid of DTT?
Vacuum dust with equipment fitted with a HEPA filter and place in a closed, labeled waste container. Dispose of via a licensed waste disposal contractor. Note: see Section 1 for emergency contact information and Section 13 for waste disposal.
What’s the best pH range for DTT reagent?
The optimal pH range for DTT is between 7.1 and 8.0, but the reagent can be used effectively at pH 6.5-9.0. DTT is well stable (longer shelf life as a powder than 2-
What is the redox potential of DTT at pH 7?
DTT is an unusually strong reducing agent, with a redox potential of -0.33 V at pH 7. The pKa of thiol groups is typically ~8.3.The reduction of a typical disulfide bond proceeds by two sequential thiol-disulfide exchange reactions and is illustrated:
What’s the difference between HCl and DTT in water?
Yes, when stressed at 30ºC, DTT demonstrates degradation starting after 3 days and increasing rapidly after 5 days. Sensitive to nickel. Effective at lower/wider pH. Effective pH range is 1.5 to 8.5. When dissolved in water resulting pH is ~2.5. Yes, the reducing power is limited to pH values >7.
Do you need to minimize exposure to air with DTT?
Exposure to air should be minimized, even though DTT has a lower tendency to be oxidized directly by air than other reducing agents. Note: DTT becomes less a less potent reducing agent with decreasing pH levels. TCEP HCl is an alternative reducing reagent which is more stable and works even at low pH.