Clinical cytometry and immunophenotyping. I. Flow cytometric analysis of normal peripheral blood and bone marrow.
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Publications and source records attributed to C W Patrick.
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A monoclonal antibody recognizing a natural killer (NK) cell-associated antigen was used to sequentially quantify numbers of peripheral blood NK cells in a small group of patients with large cell lymphoma. Patients with active disease had low numbers (less than 100/mm3) of NK cells. Patients in complete remission after therapy had normal numbers of NK cells, but those patients who relapsed had a fall of NK cell number to less than 100/mm3 preceding or concurrent with clinical relapse. A role for NK cells in the surveillance and control of abnormal lymphoproliferation is suggested, as is the possible prognostic utility of sequential peripheral blood NK cell quantification in patients with large cell lymphoma.
Peripheral blood samples from 20 normal adults and 12 patients with diagnosed B cell lymphoproliferative disease were phenotyped with a panel of 20 monoclonal antibodies (Coulter) using a whole blood lysis technique at zero time and after storage in M199 containing insulin, transferrin and L-glutamine at 24, 48 or 72 h. The samples were analyzed using flow cytometry (EPICS V) to determine the percentage of positive events for each sample. A 2-way analysis of variance showed that there were no significant differences between the initial sample and those phenotyped after 24, 48 or 72 h. These data suggest that this medium can be utilized for the storage and transport of blood samples from patients and ensure satisfactory cell surface phenotypic analysis.
Immunoregulation was assessed in a group of patients with myelodysplasia (MDS) by flow cytometric analysis of peripheral blood lymphocyte subsets and in vitro studies of mitogen-stimulated T-lymphocyte blastogenesis. Mitogenesis was significantly depressed in MDS patients compared to controls (p less than .001) and a similar defect was found in a small group of patients with untreated acute nonlymphocytic leukemia (ANLL) (p less than .005). The impaired mitogenic response ability of T-cells in these patients did not appear to be the result of alteration in lymphocyte subpopulation ratios. The observed defect might result from defective cooperation between T-lymphocytes and abnormal myeloid elements. Alternatively, the lymphocytes themselves could be derived from the abnormal clone and thus be functionally abnormal.
A 40-year-old woman presented with splenomegaly, macrocytic anemia, and red cell aplasia. Although lymphocytosis was absent in the peripheral blood, large atypical lymphoid aggregates were present in the bone marrow. Splenectomy resulted in partial remission of red cell aplasia, but a gradual increase in the number of peripheral blood lymphocytes followed during the next 36 months. Flow cytometric analysis demonstrated that the majority of these peripheral blood lymphocytes had suppressor, natural killer T-cell phenotype. No other treatment was given until red cell hypoplasia worsened 42 months after initial presentation. Repeat bone marrow evaluation again demonstrated severe erythroid hypoplasia and large abnormal lymphocytic infiltrates. Cyclophosphamide given for 8 months resulted in complete resolution of the red cell aplasia and complete clinical remission of CLL. However, flow cytometric analysis revealed persistent increase in bone marrow T-cells, and bone marrow co-culture studies demonstrated residual ability of peripheral blood mononuclear cells to inhibit erythropoiesis in vitro, suggesting that residual, clinically undetectable leukemia persists in spite of complete clinical remission.
A panel of monoclonal antibodies recognizing myeloid cell surface differentiation-associated antigens was used to study the peripheral blood myeloid population of 23 patients with myelodysplastic syndromes (MDS) and 23 controls. A marker for immaturity, defined by the presence of the antigen recognized by the My9 antibody, was found to persist on the surface of mature neutrophils in a subgroup of MDS patients. Abnormal My9 positivity was concentrated primarily, but not exclusively, in previously described morphologically defined subgroups of MDS patients considered to be at highest risk for leukemic conversion. Longitudinal study of a larger number of patients will be required to test the hypothesis that abnormal persistence of My9 may have prognostic significance.
The development of a large cell lymphoma in a patient with chronic lymphocytic leukemia (Richter syndrome) is associated with a poor prognosis. There is dispute regarding the clonal origin of the large cell component. We described a patient in whom prolonged remission of the large cell component was achieved with combination chemotherapy, followed by subsequent reemergence of the less aggressive CLL clone. Serial cell surface phenotypic data are presented suggesting origin of both histologies from a common B-cell clone.
The technicon H-6000 has the unique ability among automated hematology systems to discriminate basophils from other hematopoietic cells on the basis of astra blue positivity of heparin-containing granules. We therefore examined the ability of this flow cytochemistry system to predict allergic diathesis in vitro by examining the number of basophils detected before and after incubation with a variety of allergens. One hundred subjects with documented atopic reactions to known allergens were examined and compared to 13 patients who had undergone successful desensitization and 12 normal subjects who had no history of atopy. One subject who had a previous anaphylactic response to penicillin but was skin-test negative to the major antigenic determinant demonstrated 55% degranulation after incubation with penicillin G. Finally, the basophil degranulation as assessed by the Technicon H-6000 was compared to a histamine release assay to determine the sensitivity of the automated system. The results suggest that flow cytochemistry may prove a valuable tool in predicting the success of desensitization therapy for common allergens and may have the potential to screen atopic reaction in the routine hematology laboratory.