(minimal residue disease, MRD), ,,, ALL %, 8

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1 Chin J Hematol June 2003 Vol 24 No B (minimal residue disease MRD) 58 B (B2ALL) (89. 7 %) B2 ALL MRD CD 10 ΠCD 34 ΠCD 19 CD 38 CD 65 CD 66c CD % % % % % % % % % MRD B ; ; ; Detection of minimal residual disease in childhood B2ALL by flow cytometry XU Chong ZHAO Hui2jun WU Zheng2hong XUE Hui2liang TANG Jing2yan CHEN Jing PAN Ci CHEN Jing LI Li GU Long2jun SHEN Li2song. Laboratory Diagnostic Center Xin Hua Hospital Shanghai Children s Medical Center Shanghai Second Medical University Shanghai China Abstract Objective To establish a flow cytometric method for detecting minimal residual disease (MRD) in children with B2ALL and evaluate its clinical application. Methods Fifty2eight childhood B2ALL cases entered this study and 30 MRD analyses were performed after remission induction therapy. Four2color fluorochrome labeled monoclonal antibodies were used to analyze the cell immunophenotypes. Cells from normal bone marrow were used as controls. The leukemic cell populations located in flow cytometry dot plots different from those of normal were con2 sidered to be the markers of interest in the first step screening and then used to monitor MRD step after therapy. Results Fifty2eight cases of childhood B2ALL were screened for antibodies combinations of interest and were identi2 fied in %(52Π58) of these cases. The four2color antibody combinations consisted of CD 10 ΠCD 34 ΠCD 19 plus an2 other effective marker such as CD 38 CD 65 CD 66c CD 21. The sensitivity of this method was % much higher than microscopic inspection. In 8 cases whose bone marrow microscopically showed no residual leukemic cells the percentage of leukemic cells were identified with this method of % % % % % % % and % respectively. Conclusion The application of flow cytometry in MRD monitoring can significantly improve the detection sensitivity in childhood B2ALL thus facilitate the further treatment decision and follow2up. Key words Leukemia B2cell acute ; Antibody monoclonal ; Minimal residual disease ; Flow cy2 tometry (MRD) B (B2ALL) MRD : B2ALL

2 Chin J Hematol June 2003 Vol 24 No. 6 L 1 15 L 2 39 L 3 4 FAB % EDTA 0. 2 % PBS) 10 min PVDL Wash 1 ( ) 10 min Wash 2 1 MRD 3. 3 : 2 CD 38 2FITC CD 21 2FITC CD 65 2FITC CellQuest (version 3. 2) CD 56 2FITC KOR2SA35442FITC CD 33 2FITC Immunotech ;CD 10 2PE anti2tdt2fitc 3. 4 :CellQuest (version 3. 2) Dako ; CD 34 2PerCP CD 19 2APC CD 15 2FITC CD 45 2FITC BD ; IgM2PE Southern Biotechnology Associates MRD : 3. 1 :B2ALL Ficoll Ficoll PBSA 2 PBSA (2 3) 10 5 CD 38 CD 45 CD 56 CD 65 CD 66c ( KOR2SA3544 ) CD 15 CD 21 CD 22 CD 33 CD l MRD : CD 38 CD 45 CD 58 CD 65 CD 66c CD 21 CD 22 CD 34 c 1 B2ALL 10-4 ( CD 56 ) CD 15 CD 13 CD 33 CD 66c 2 B2ALL ( ) 1 1 MRD 1 CD 38 Π FITC PE PerCP APC CD 38 CD 10 CD 34 CD 19 CD 45 CD 10 CD 34 CD 19 CD 65 CD 10 CD 34 CD 19 CD 56 CD 10 CD 34 CD 19 CD 66c CD 10 CD 34 CD 19 CD 15 CD 10 CD 34 CD 19 CD 21 CD 10 CD 34 CD 19 CD 22 CD 10 CD 34 CD 19 CD 33 CD 10 CD 34 CD 19 CD 13 CD 10 CD 34 CD 19 TdT 3 CD 10 CD 34 CD 19 TdT 3 c 3 CD 34 CD 19 : : min PBSA(0. 2 % ( 0. 2 % PBS) 2 8E ) ( St. Jude Children s Research Hospital Dr Dario Campana ) 40 min 5 MRD Wash (5 % 1. 5 % 30 CD 10 ΠCD 34 ΠCD 19 CD 22 ΠCD 10 ΠCD 34 ΠCD 19 CD 45 ΠCD 10 Π CD 34 ΠCD 19 MRD B2ALL 2 B2ALL ( ) CD 22 CD 13 CD 21 CD (89. 7 %) MRD

3 Chin J Hematol June 2003 Vol 24 No MRD : % % 2 58 B2ALL % % % % % [1 ] %( CD 38 ΠCD 10 ΠCD 34 ΠCD 19 ( %) ( %) CD 45 ΠCD 10 ΠCD 34 ΠCD TdTΠCD 10 ΠCD 34 ΠCD CD 66c ΠCD 10 ΠCD 34 ΠCD CD 22 ΠCD 10 ΠCD 34 ΠCD CD 33 ΠCD 10 ΠCD 34 ΠCD CD 13 ΠCD 10 ΠCD 34 ΠCD CD 21 ΠCD 10 / CD 34 ΠCD TdTΠcΠCD 34 ΠCD CD 65 ΠCD 10 ΠCD 34 ΠCD CD 58 ΠCD 10 ΠCD 34 ΠCD ) 3 CD 38 ΠCD 10 ΠCD 34 ΠCD 19 2CD 38 CD 45 B2ALL B CD 10 B2 ALL 2 CD 10 (CD 38 ) a b c CD 38 ΠCD 10 ΠCD 34 ΠCD 19 CD 22 ΠCD 10 ΠCD 34 ΠCD 19 CD 45 ΠCD 10 ΠCD 34 ΠCD 19 CD + 19 d e f 1 B2ALL g h i d CD 38 CD 10 g CD 38 ΠCD 10 ΠCD 34 ΠCD 19 e CD 22 (R5 ) h R % f CD 10 CD 45 (R4 ) i R % 1 MRD

4 Chin J Hematol June 2003 Vol 24 No. 6 a b c a R1 ;b R2 CD 19 ;c G2 = R1 and R2 CD 10 ΠCD 38 CD + 19 dπg eπh fπi 3 B2ALL d e f 3 d f CD 38 R3 e CD 38 CD 10 R4 g h i 3 g h < % MRD i R % MRD 2 CD 38 ΠCD 10 ΠCD 34 ΠCD 19 MRD MRD MRD B2ALL CD 65 CD 21 CD 22 CD 13 Π : CD 19 CD 10 CD 34 B / 10 [2 ] CD 38 CD 45 ( ) [3 ] ; ( (CD 66c ) (CD 21 CD 34 c CD 34 ) [1 ) MRD ] B ( CD 13 CD 33 CD 15 CD 65 CD 56 ) B ALL

5 Chin J Hematol June 2003 Vol 24 No MRD 1 Campana D Coustan2Smith E. Detection of minimal residual disease in acute leukemia by flow cytometry. Cytometry : Lavabre2Bertrand T Janossy G Ivory K et al. Leukemia2associated changes identified by quantitative flow cytometry :. CD10 expression. Cytometry : Behm FG Raimondi SC Schell MJ et al. Lack of CD45 antigen on blast cells in childhood acute lymphoblastic leukemia is associated with chromosomal hyperdiploidy and other favorable prognostic features. Blood : ( : ) ( : ) T T (Subcutaneous panniculitis2like T2cell lymphoma SPTCL) T 40 mgπd 15 ) :SPTCL T Gonzalez WHO T ( NK ) SPTCL LCA CD 8 CD 3 CD 45RO CD 4 SPTCL 50 % (HS) 0. 2 gπd 10 d T : 3 cm 3 cm 5 cm 5 cm CD 43 CD 45RO CD 3 CD 4 ( + ) CD 8 ( - ) 1 cm 1 cm :Hb(133 CD 4 ( - ) CD 8 ( + ) CD 4 ( - ) CD 8 ( - ) TCR 144) gπl RBC( ) ΠL WBC ( ) 10 9 ΠL BPC(138174) 10 9 ΠL : 59 gπl Perniciaro SPTCL : 23 gπl : ; CTOP( 600 mg 1 ; 20 mg 1 40 mg 2 ; 1 mg 1 ; CHOP HS ( : ) : ( : )