When validating a new Planer Integra 750 plus we observed very slow engraftment for the first 5 patients compared to other prior patients receiving stem cell grafts frozen with our old Planer as well as patients receiving stem cell grafts frozen with dump freezing (-80°C over night). Validation test on cryo vials after freezing with Intergra 750 plus (CD34+ count/viability) and CFU assays where acceptable. At our institution stem cell grafts are frozen at a volume of 200 ml in 1000 ml Cryomacs Freezing bags (Miltenyi), with 10% DMSO.
Single dose 4 x 106 CD34+ per Kg
Below is the freezing protocol
Seeding Type: No seeding
Step list:
Ramp step: Ramp 0.0°C @ -10.0°C/min
Hold step: 15:00
Ramp step: Ramp -3.0°C @ -1.0°C/min
Ramp step: Ramp -20.0°C @ -20.0°C/min
Hold step: 01:00
Ramp step: Ramp -14.0°C @ 10.0°C/min
Ramp step: Ramp -50.0°C @ -1.0°C/min
Ramp step: Ramp -150.0°C @ -20.0°C/min
Hold step: 02:00
The engraftment day of ANC and Plt varied considerably between these first 5 patients but in all cases was above the average in our operation of day 15 for ANC and Plt (20) and day 16 for Plt (50)
If someone has encountered similar problems or has any suggestions for us on how to proceed with our validation please respond.
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