1. What experiments are suitable for D-fluorescein?
D-fluorescein, particularly in the form of D-luciferin potassium salt and 5-fluoro-luciferin, is widely used as a substrate for luciferase in various experimental applications. These include: - Reporter gene assays to monitor gene expression - In vivo bioluminescence imaging for tracking cellular activity in living organisms - In vitro imaging for studying cell behavior in controlled environments - ATP quantification, as the luciferase reaction is directly dependent on ATP levels - Research involving luciferase enzymes and their corresponding genes These substrates are essential tools in molecular biology, offering high sensitivity and specificity for detecting biological processes in real-time.2. The basic principle of D-fluorescein in live imaging
The use of D-fluorescein in in vivo and in vitro imaging relies on the enzymatic reaction catalyzed by luciferase. When D-luciferin is present in excess, the amount of light emitted (photons) correlates directly with the concentration of the enzyme and the number of cells. This allows researchers to quantify cellular activity or track the progression of biological events within living systems.3. Stability of D-fluorescein potassium salt solution
D-luciferin potassium salt dissolves readily in water and buffer solutions, with a solubility up to 25 mg/mL. It is typically used at concentrations between 3–15 mg/mL. However, its stability is influenced by several factors. The pH of the solution plays a critical role: below 6.5, hydrolysis can occur, while above 7.5, racemization may convert the D-form into the L-form, which is less effective in luciferase reactions. Oxygen in the solution also accelerates degradation. Under optimal storage conditions—such as -80°C, neutral pH, and an oxygen-free environment—the degradation rate is approximately 0.2% per day. Therefore, it is recommended to use the solution within one year after preparation. Additionally, moisture exposure can lead to racemization, where D-luciferin converts to L-luciferin, potentially inhibiting the luciferase reaction. Proper handling and storage are crucial for maintaining the integrity and performance of the reagent.4. Differences between D-luciferin, sodium D-luciferin, and potassium D-fluorescein
While all these forms are similar in function, the main difference lies in their solubility and dissolution properties. D-luciferin potassium salt exhibits excellent water solubility and fast dissolution, making it ideal for most biological assays. Sodium D-luciferin may have slightly different solubility characteristics, depending on the buffer system used.5. Storage and transportation conditions for D-luciferin potassium salt
For optimal preservation, D-luciferin potassium salt should be stored at ≤ -20°C, away from light, and kept dry. An unopened bottle can remain stable for up to two years under these conditions. During transportation, high-purity D-luciferin potassium salt can be shipped at room temperature for short periods. Each original container is flushed with argon gas to prevent oxidation and maintain stability. Once opened, it must be handled carefully and stored under the same strict conditions. Due to its hygroscopic nature, it is important to allow the material to reach room temperature before weighing and to work quickly to avoid moisture absorption. Proper storage and handling ensure consistent performance and reliable results in all experiments that rely on D-fluorescein as a substrate.Dog Rubber Toys,Latex Dog Toy,Dog Chew Toys,Interactive Dog Toys
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