Advanced Flow Workshop Shumway Murphey

More documents

Recommendations

Info

Workshop 2: Advanced Flow Workshop- Wayne Shumway, CHT, CHS and Cathi Murphey, MT(ASCP), CHS, BLM 1 “RELATIVE” LINEAR VALUES RLV-BD 0.1 RLV-BC 10 100 1000 10,000 1 10 100 1,000 64 128 192 50 100 150 200 250 End of 1 st decade End of 2 nd decade End of 3 rd decade 256 512 768 256 End of 4 th decade 1024 “RELATIVE” LINEAR VALUES Current XM: Last XM PMT settings X 4 = Last XM: Last XM PMT settings MCF = 256 MCF = 410 MFI Ratio = 4 MCF = 77 MCF = 231 MFI Ratio = 4 154 MCF shift 154 MCF shift 8 – 2 = 6 MFI Shift 40 – 10 = 30 MFI Shift 400% OF BACKGROUND 400% OF BACKGROUND 200 400 600 800 1000 1 2 3 4 5 6 7 8910 200 400 20 30 40 50 60 70 100 “RELATIVE” LINEAR VALUES X 4 = Current XM: NEW XM PMT settings MCF = 256 MCF = 410 MFI Ratio = 4 154 MCF shift 40 – 10 = 30 MFI Shift 400% OF BACKGROUND “RELATIVE” LINEAR VALUES X 4 = Current XM: CONSTANT PMT settings Last XM: CONSTANT PMT settings MCF = 256 MCF = 410 MFI Ratio = 4 MCF = 77 MCF = 231 MFI Ratio = 4 154 MCF shift 154 MCF shift 8 – 2 = 6 MFI Shift 40 – 10 = 30 MFI Shift 400% OF BACKGROUND 400% OF BACKGROUND 1 2 3 4 5 6 7 8910 200 400 20 30 40 50 60 70 100 1 2 3 4 5 6 7 8910 200 400 20 30 40 50 60 70 100 MESF Values ** **** MESF Values are Independent of Instrument and Settings **** 200,000 50,000 12,500 MESF line at initial PMT settings MESF line at increased PMT settings ** Relative positions are not to scale. For illustration only. MESF Acquisition 2 8 10 40 Cytometer Scale (RLV) ** Reference Bead MESF 2546 6393 19019 47474 6
Workshop 2: Advanced Flow Workshop- Wayne Shumway, CHT, CHS and Cathi Murphey, MT(ASCP), CHS, BLM Universal Standardization of the FCXM Will Require More Than MESF Conversion due to the Variability of: 1) FCXM protocols 2) Secondary Ab characteristics 3) Negative control sera reactivity 4) HLA expression FCXM Interpretation Using MESF 1) The key to obtaining the clearest perspective on FCXM data is the differential comparison of fluorescence intensities that are expressed as linear values 2) MESF are simply standardized linear values 3) If a lab FCXMs on multiple instruments, MESF conversion provides a convenient way to standardize data between machines - BUT 4) MESF conversion alone is not sufficient to universally standardize the FCXM between laboratories 5) MESF reference lines can be used to verify the proper functioning of fluorescence detection systems Auto XM Negative Positive Donor XM IS NOT Due to auto-ab ab Patient Cells + Patient Serum Positive Positive Donor XM MAY BE Due to auto-ab ab Negative Positive Donor XM IS NOT Due to auto-ab ab Patient Cells + Patient Serum Positive Positive Donor XM MAY BE Due to auto-ab ab Cell Washer IF Positive Donor XM Donor Cells + Donor Adsorbed patient serum Negative HLA-ab likely Positive HLA-ab NOT likely 7
Page 1 and 2: Workshop 2: Advanced Flow Workshop-
Page 3 and 4: Workshop 2: Advanced Flow Workshop-
Page 5: Workshop 2: Advanced Flow Workshop-

Advanced Flow Workshop Shumway Murphey

You also want an ePaper? Increase the reach of your titles

Delete template?

Save as template?