CTNs
Calf Thymocyte Nuclei
These natural cycling cells provide daily cell-cycle statistics, and also evaluate the doublet discrimination module or electronic pulse-processing system.
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| 2.0 mL Catalog No. 1008$118.00 |
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BioSure® Calf Thymocyte Nuclei (CTNs) are an internal biological reference material that can function as a performance standard when calibrating and troubleshooting flow cytometers for DNA analyses. The published genome size is 3.7 picograms1. CTNs are isolated from fresh calf thymus tissue and fixed in formaldehyde-PBS. The procedure yields single nuclei, doublets, and a few larger aggregates. The control has been tested on FACScan flow cytometers equipped with a Doublet Discrimination Module (DDM) to assure that nuclei for each phase of the cell cycle are present.
FORM
BioSure® CTNs are supplied as an unstained suspension in 2.0 milliliter volumes. The nuclei count per milliliter is at least 2.0 X 107.
STABILITY
CTNs are stable at 2° - 8° C until the expiration date denoted on each vial label.
PREPARATION OF PROPIDIUM IODIDE (50 microgram/mL) STAINING SOLUTION
To 48 milliliters of calcium and magnesium free Dulbecco's PBS, add 2.5 mg of propidium iodide (PI) and 0.3 mL of Nonidet P40 (NP40). Mix the solution and Q.S. to 50 mL. Staining solution is stable at 2° - 8° C for at least 30 days when stored protected from light in an amber bottle.
PREPARATION OF WORKING SOLUTION
The recommended working solution is a 1:10 dilution of the stock CTNs in the PI stain prepared previously. This provides an adequate density of nuclei and sufficient working solution for several hours.
Typical CTN graph
1. Gently mix the CTN stock for 5 - 10 seconds to suspend the nuclei, and add one drop to a test tube (13 X 75 mm).
2. Add 1.0 mL of PI stain, mix for 5 - 10 seconds, and incubate at room temperature (22° - 25° C) for ten minutes protected from light.
3. Mix sample 5 - 10 seconds and evaluate. Samples are typically stable at 2° - 8° C for four hours when protected from light.
NOTE: The flow cell of some cytometers is susceptible to clogging. Filtering the stained preparation through a 30 - 50 mesh nylon screen will eliminate large nuclear aggregate.
PRECAUTIONS
Formaldehyde has been listed as a carcinogen by the Environmental Protection Agency (EPA). Wear gloves when handling this material and avoid contact with skin and eyes.
