Procedure for

Daily Monitoring

Establish Daily Values to Provide a Laboratory Record of Instrument Stability with BioSure CRBC's

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Once target reference values for the CRBC's have been established, monitoring of the parameters is performed on a daily basis. Instrument settings for the target values are recalled and the new daily values compared to the target reference values. Recording of the new daily values onto Levey-Jennings plots provides a laboratory record of instrument stability over time.

Daily Monitoring Procedure

1) Prepare a fresh dilution of the CRBC's to achieve a flow rate of about 500 - 1000 events/second. Place the tube on the cytometer. Set the instrument parameters as follows: FSC-linear, SSC-linear, FL1-log, FL2-log.

2) Recall the instrument settings for the FSC parameter. The CRBC high scatter peak and the low scatter peak should both be on scale in the forward scatter histogram display. Calculate the difference in mean channels between the CRBC high scatter peak and the low scatter peak. Both mean channels should be within plus or minus ten channels of the target values. A significant loss (approximately 10%) of the high peak's target mean channel value or delta probably indicates optical system problems. Record the values.

3) Proceed to the SSC histogram display. Recall the instrument settings for the SSC parameter. Compare the side scatter mean channel number with the mean channel number established for the SSC during the initial target values run. A variation of more than 10% may indicate the need for realignment or instrument servicing

4) Proceed to the FL1 histogram display. Recall the averaged FL1 PMT setting established by the control donor lymphocytes. Adjust the PMT voltage in the FL1 parameter (if necessary) to place the CRBC's in the same FL1 channel as determined by the initial target values. Record the adjusted PMT voltage and C.V. A variation of more than 10% may indicate the need for realignment or instrument servicing.

5) Proceed to the FL2 histrogram display. Recall the averaged FL2 PMT setting established by the control donor lymphocytes. Adjust the PMT voltage in the FL2 parameter (if necessary) to place the CRBC's in the same FL2 channel as determined by the initial target values. Record the adjusted PMT voltage and C.V. A variation of more than 10% may indicate the need for realignment or instrument servicing.

6) Repeat for FL3 or other parameters, if desired.

Before any realignment or service of the instrument, prepare a fresh dilution of the CRBC standards and repeat the test protocol. If variations persist with the freshly prepared sample of CRBC's, then servicing or realignment may be necessary.

Daily Monitoring Procedure
1)	Prepare a fresh dilution of the CRBC's to achieve a flow rate of about 500 - 1000 events/second. Place the tube on the cytometer. Set the instrument parameters as follows: FSC-linear, SSC-linear, FL1-log, FL2-log.
2)	Recall the instrument settings for the FSC parameter. The CRBC high scatter peak and the low scatter peak should both be on scale in the forward scatter histogram display. Calculate the difference in mean channels between the CRBC high scatter peak and the low scatter peak. Both mean channels should be within plus or minus ten channels of the target values. A significant loss (approximately 10%) of the high peak's target mean channel value or delta probably indicates optical system problems. Record the values.
3)	Proceed to the SSC histogram display. Recall the instrument settings for the SSC parameter. Compare the side scatter mean channel number with the mean channel number established for the SSC during the initial target values run. A variation of more than 10% may indicate the need for realignment or instrument servicing
4)	Proceed to the FL1 histogram display. Recall the averaged FL1 PMT setting established by the control donor lymphocytes. Adjust the PMT voltage in the FL1 parameter (if necessary) to place the CRBC's in the same FL1 channel as determined by the initial target values. Record the adjusted PMT voltage and C.V. A variation of more than 10% may indicate the need for realignment or instrument servicing.
5)	Proceed to the FL2 histrogram display. Recall the averaged FL2 PMT setting established by the control donor lymphocytes. Adjust the PMT voltage in the FL2 parameter (if necessary) to place the CRBC's in the same FL2 channel as determined by the initial target values. Record the adjusted PMT voltage and C.V. A variation of more than 10% may indicate the need for realignment or instrument servicing.
6)	Repeat for FL3 or other parameters, if desired.

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