• Sucrose Solutions
• Cesium Chloride Gradients

Gradients used in ultracentrifugation are difficult to count. When mixed with most cocktails, they form two-phase systems or self-absorption quench may occur due to the presence of highly concentrated salt or sucrose solutions.

Using cocktails formulated with emulsifier (Ready-Solv HP™, Ready Protein+™, Ready Safe™), it is possible to solubilize up to 25% sucrose gradients into a single-phase, quench correctable system. More concentrated solutions require a special procedure.

Dilution of concentrated CsCl gradients is essential to achieve homogeneous counting conditions without self-absorption quench. With minimal effort, a special CsCl solubilizing solution can be prepared.

Sucrose Solutions

Condition A: 10% sucrose
Add up to 2 mL of 10% sucrose to either 8-10 mL of Ready-Solv HP or 10 mL of Ready Protein+ or Ready Safe. Shake well and count.

Expected tritium efficiency is 40-45%.

Condition B: 25% sucrose
Add up to 2 mL of 25% sucrose to 8-10 mL of Ready Protein+. Shake and count.

Expected tritium efficiency is 40-45%.

Condition C: 25% sucrose; large volumes¹
Add 1-4 mL of 25% sucrose to either 10 mL of Ready Gel™ or Ready Value™. Shake and count.

Expected tritium efficiency is 30-40%.

Note:
1. For ambient temperature counting only.

Condition D: Mini-vial counting
Add up to 2 mL of 25% sucrose to 4 mL of Ready Flow III™. Shake and count.

Expected tritium efficiency is 24-30%.

Condition E: 10 drop of 12-33% gradient fractions

1. Prepare a special stock solution by adding 50 mL of water per liter to Ready Flow III, Ready-Solv HP or Ready Protein+.
2. Add a 10 drop gradient fraction (approximately 0.5 mL) to a scintillation vial (the LS vial may be placed in the fraction collector).
3. Add to the vial any of the following:
   1. 2.5 mL of aqueous Ready Flow III
   2. 3.0 mL of aqueous Ready-Solv HP
   3. 5.5 mL of aqueous Ready Protein+
4. Shake well to form a clear solution.
   The volumes given above are necessary for the 12% fraction. Higher sucrose concentrations require less cocktail for solubilization.

Cesium Chloride Gradients

Add up to 0.8 mL of 6 M CsCl to 10 mL of CsCl solubilizing solution¹. Shake until clear and count.

Expected tritium efficiency is 35-40%.

Note:

1. CsCl solubilizing solution is prepared by mixing 100 mL of water with 960 mL Ready Gel.
    

Home | Products | Customer Support | Resource Center Employment | Our Company | Contact Us | Site Map  Privacy |  ©1998 - 2009 Beckman Coulter, Inc.