Procedure for

Alignment and Linearity

Check Your Cytometer Performance with BioSure CEN's

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Here's everything you need to know about using BioSure Chicken Erythrocyte Nuclei (CEN's) for cytometer alignment and linearity checks. It's so easy and effective, you'll want to make it a routine process in your lab! It's so economical, you can do this check daily, or whenever your instrument appears not to be performing. Get ready for optimized performance.

Alignment and Linearity Procedure

All you do is prepare the PI solution, prepare a working solution with BioSure® CEN's, incubate, analyze, and evaluate.

Prepare the BioSure PI Solution

1)
Remove 4.0 mL of PI Staining Buffer from Bottle B and add it to the PI Stain in Bottle A.

CAUTION: Wear gloves and protective eye wear when handling the PI Stain and solution. Always handle and dispose of bottles and contents in accordance with good laboratory practices.

2) Replace the cap on Bottle A and gently invert the bottle several times.

3) Pour the contents of Bottle A into Bottle B and mix.

4) Rinse Bottle A with 4.0 mL of the mixture in Bottle B to remove any remaining dye and add the mixture back into Bottle B.

5) Place the enclosed PI Staining Solution label over the label on Bottle B and store at 2 - 8 degrees C. The staining solution is ready for use.

Prepare the BioSure PI Solution
1)	Remove 4.0 mL of PI Staining Buffer from Bottle B and add it to the PI Stain in Bottle A. CAUTION: Wear gloves and protective eye wear when handling the PI Stain and solution. Always handle and dispose of bottles and contents in accordance with good laboratory practices.
2)	Replace the cap on Bottle A and gently invert the bottle several times.
3)	Pour the contents of Bottle A into Bottle B and mix.
4)	Rinse Bottle A with 4.0 mL of the mixture in Bottle B to remove any remaining dye and add the mixture back into Bottle B.
5)	Place the enclosed PI Staining Solution label over the label on Bottle B and store at 2 - 8 degrees C. The staining solution is ready for use.

Prepare the Working Solution

The recommended working solution is a 1:10 dilution of BioSure® CEN's to PI Staining solution. This dilution provides an adequate density of nuclei for analysis, and supplies sufficient working solution for several hours.

1) Gently mix BioSure CEN's for 5 - 10 seconds to suspend the nuclei.

2) Add one or two drops of CEN's to a 13 x 75-mm test tube.

3) Add 1.0 mL of PI stain to the test tube, mix for 5 - 10 seconds, and incubate at room temperature (22 - 25 degrees C) for 10 minutes. Protect from light.

4) Mix the sample for 5 - 10 seconds before analyzing.

Prepare the Working Solution

The recommended working solution is a 1:10 dilution of BioSure® CEN's to PI Staining solution. This dilution provides an adequate density of nuclei for analysis, and supplies sufficient working solution for several hours.
1)	Gently mix BioSure CEN's for 5 - 10 seconds to suspend the nuclei.
2)	Add one or two drops of CEN's to a 13 x 75-mm test tube.
3)	Add 1.0 mL of PI stain to the test tube, mix for 5 - 10 seconds, and incubate at room temperature (22 - 25 degrees C) for 10 minutes. Protect from light.
4)	Mix the sample for 5 - 10 seconds before analyzing.

Evaluate the Linearity and Alignment

Look at the single-nuclei peak near the ordinate, and the other peaks with mean channel values about two, three, and four times greater. The flow cytometer is properly aligned when CV's for the single, double, and triple peaks are less than or equal to 3%.

Procedural Notes

The mixed BioSure® PI staining solution is stable for at least 90 days when refrigerated at 2 - 8 degrees C in the amber bottle.
BioSure® CEN's are stable when refrigerated at 2 - 8 degrees C, until the expiration date on each vial label.
Mixed samples (PI solution and CEN's) remain stable for four hours when refrigerated at 2 - 8 degrees C and protected from light.
The flow cells of some cytometers are susceptible to clogging. If clogging occurs, filter the stained preparation through a 30 to 50 micron mesh nylon screen to eliminate large nuclear aggregates.

Click here for the BioSure® CEN's data sheet

Click here to find out more about BioSure® CEN's and other controls

Click here to find out more about our convenient PI Staining Solution

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BioSure®
1050 Whispering Pines Lane, Suite F
Grass Valley, CA 95945

Phone (800) 345-2267 or
(530) 273-5095
Fax (530) 273-5097

Alignment and Linearity

Check Your Cytometer Performance with BioSure CEN's

Alignment and Linearity Procedure

Prepare the BioSure PI Solution

Prepare the Working Solution

Evaluate the Linearity and Alignment Look at the single-nuclei peak near the ordinate, and the other peaks with mean channel values about two, three, and four times greater. The flow cytometer is properly aligned when CV's for the single, double, and triple peaks are less than or equal to 3%.

Evaluate the Linearity and Alignment

Procedural Notes

Evaluate the Linearity and Alignment

Look at the single-nuclei peak near the ordinate, and the other peaks with mean channel values about two, three, and four times greater. The flow cytometer is properly aligned when CV's for the single, double, and triple peaks are less than or equal to 3%.