Nordic flow cytometry standards for CD34+ cell enumeration in blood and leukapheresis products: report from the second Nordic Workshop. Nordic Stem Cell Laboratory Group (NSCL-G).

Recently, Nordic protocols have been activated for the use of high-dose chemotherapy and radiotherapy to treat newly diagnosed young patients with multiple myeloma, breast cancer, and malignant lymphomas. The protocols include transplantation of autologous stem cells or progenitors as supportive therapy. This requires a safe number and optimal quality of the harvested cells. Consequently, the Nordic stem cell laboratories established a collaboration to optimize enumeration of CD34+ cells by flow cytometry. In the first workshop (WS-I) report, we addressed the issue of standardization of technical variables. Taking into consideration the variations in estimation of progenitors, a safe graft should contain > 2.0 x 10(6) CD34+ cells per kilogram patient weight, harvested by two or three leukaphereses. This report presents the results from the second workshop (WS-II). Twelve blood or leukapheresis products were shipped and analyzed in 24 stem cell laboratories, and all results were returned to a central database at Herlev Hospital, Copenhagen. The data documented a significant correlation between CD34+ cell enumerations of the 12 samples analyzed in each of the 24 different stem cell laboratories within the Nordic stem cell laboratory group (NSCL-G). At a practical workshop, the NSCL-G developed a standard for the performance of CD34 enumeration. This is based on the simple Milan Protocol, which uses the anti-HPCA-2PE marking of CD34+ (FL-2) nongranulated (lowSSC) cells. The NSCL-G has decided to hold a third workshop (WS-III) to establish working recommendations for double marking of CD34+ lineage-specific subsets and for handling cellular material for storage of RNA and DNA from blood and leukapheresis products.