Objective:To prepare anti-human c-kit monoclonal antibody(McAb) by genetic immunization in spleen,and to determine practicability of these means to produce McAbs based on the biological activity of anti-human c-kit antibody.Methods:Recombinant plasmid pcDNA3.1/c-kit extracellular domain was constructed by molecular cloning techniques,and was used to immunize BALB/c mice in spleen directly to prepare mAb against human c-kit by routine hybridoma technique.FASC、fluorescence microscope and Western blot were utilized to identify the prepared antibody.Results:c-kit extracellular region was cloned and insert pcDNA3.1 plasmid successfully.Three hybridoma cell lines 6C4、2C5 and 5D5 that secrete anti-human c-kit McAbs were obtained after using intra-spleen immunization with a DNA vaccine.The isotypes of these three antibodies were all IgM,and the epitopes were different with each other.Conclusion:The method of genetic immunization into spleen can be used to prepare anti-human c-kit monoclonal antibodies.

Objective:To analyze the clinical manifestations and laboratory features in patients with myeloid neoplasms complicated with clonal T large granular lymphocyte (T-LGL) proliferation.Methods:The clinical data of 5 patients with myeloid neoplasms complicated with clonal T-LGL proliferation from November 2017 to November 2018 in Institute of Hematology & Blood Diseases Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College were analyzed retrospectively.Results:The median age was 60 years old. All patients had a history of abnormal peripheral blood cell counts for over 6 months. The absolute lymphocyte count in peripheral blood was less than 1.0×10 9/L. In addition to the typical T-LGL phenotype, the immunophenotype was heterogenous including CD4 +CD8 - in 2 patients, the other 3 CD4 -CD8 +. Four patients were αβ type T cells, the other one was γδ type. STAT3 mutation was detected in 1 patient by next-generation sequencing, the other 4 cases were negative. Conclusions:Clonal T-LGL proliferation with myeloid neoplasm develops in an indolent manner, mainly in elderly patients. Hemocytopenia is the most common manifestation. The diagnosis of T-LGL proliferation does not have specific criteria, that it should be differentiated from other T cell proliferative disorders, such as T-cell clones of undetermined significance. STAT3 or STAT5b mutation may help distinguish.

OBJECTIVETo investigate the myeloperoxidase (cMPO) expression pattern by flow cytometry (FCM) in patients with acute myeloid leukemia (AML) and its role in classifying AML.

METHODSEight- color multiparametric FCM with CD45/SSC gating was used to determine the cMPO expression in 502 AML patients.

RESULTSThe positive rate of cMPO in all patients was 58.0%, in which the proportion of normal positivity, dim positivity and partial positivity was 21.5%, 34.1% and 2.4%, respectively. The remaining case (42.0%) were all negative. In AML with t (15;17）(q22;q12)/PMLRARα, the positive rate was the highest (100%) and the intensity was similar to that of the normal granular leukocytes, followed by AML with t (8;21（q22;q22/RUNX1-RUNX1T1, the positive rate was 91.4% and the intensity was mostly dim. AML with minimal differentiation and acute megakaryoblastic leukemia were all cMPO negative. The positive rates of cMPO in the remaining subtypes were between 22.7% and 76.2%.

CONCLUSIONThe positive rate and intensity of cMPO were significantly different among different subtypes of AML.

Cell Differentiation ; Flow Cytometry ; Granulocytes ; Humans ; Leukemia, Myeloid, Acute ; classification ; genetics ; Peroxidase ; genetics