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L S Kaplow

Publications and source records attributed to L S Kaplow.

At least 19 recordsLinked to original sources

Assessment of leukocyte alkaline phosphatase by image analysis.

We have shown that it is possible to automate the assessment of leukocyte alkaline phosphatase by using an azo dye cytochemical staining procedure and a commercial, highly sophisticated image analysis instrument originally designed specifically as a differential white cell counter. The data to date indicate that values obtained by this approach are at least as precise and accurate as current manual techniques. Instrumental analysis avoids the subjectivity associated with manual interpretation of staining intensity and should permit meaningful interlaboratory comparisons. The stability of the stained smears upon exposure to immersion oil or Polymount mounting medium proved to be an unexpected bonus. In addition to such functional data on leukocytes as illustrated by this report, these instruments, with appropriate staining methods and software, can also provide clinically useful quantitative data on red cells, as have been described for reticulocytes. We hope to see more clinical applications in the future for these expensive and target-oriented image analysis instruments. They are capable of automatically providing objective quantitative information on a cell by cell basis--providing feature data that cannot be obtained by other means.

Alkaline Phosphatase

The relationship of myeloperoxidase activity to neutrophil maturity and other hematologic indicators of infection.

Myeloperoxidase activity (MPO) in 100 male hospitalized patients was quantified by enzyme cytochemical flow cytometric analysis. These data were compared with commonly used laboratory indicators of infection. The correlation coefficients were statistically significant for increased MPO activity with band count, toxic granulation, and the Schilling Index; however, the orders of magnitude were too low to suggest the use of MPO as a clinical parameter. Patients were given an infection rank based on their numbers of abnormal laboratory indicators of infection. There was a highly significant difference in mean neutrophil peroxidase (MNP) activity between those patients with three or more indicators of infection and those with two or less: t = 3.68, p less than 0.001. It is concluded that whereas MNP may not be a useful indicator of a left shift or of leukocytosis, it might be helpful as an indicator of infection or inflammation. Further studies will be required to support this hypothesis.

Humans

Relationship of reticulocyte age to polychromasia, shift cells, and shift reticulocytes.

Polychromatophilic erythrocytes on Wright-stained blood smears represent young reticulocytes. Ratios of polychromatophilic cells to total reticulocyte counts have been used to estimate marrow response to erythropoietin stimulation. These ratios, however, require both accurate counts of polychromatophilic cells on Wright-stained blood smears and reticulocytes on supravitally stained blood smears. Data from this study indicated that reticulocytes of Heilmeyer groups I, II, and III best represent polychromatophilic cells. Group III reticulocytes, however, were found in normal circulation and were difficult to distinguish from group IV reticulocytes. Groups I and II were not found in normal circulation and were easily identified on routine reticulocyte preparations. The term "shift reticulocyte" is proposed for reticulocytes of groups I and II only. The present study suggests that the "shift reticulocyte count," expressed as percent of 100 reticulocytes, is a more useful indicator of marrow response to anemia than total reticulocyte counts.

Erythrocyte Aging

Myeloperoxidase deficiency. Increased sensitivity for immunocytochemical compared to cytochemical detection of enzyme.

The discovery of hereditary deficiency of myeloperoxidase (MPO) in neutrophils and monocytes of affected individuals has been based on the absence of cytochemical staining in these peripheral blood cells. We report that an immunocytochemical method shows more sensitivity than either the benzidine or 4-chloro-1-naphthol cytochemical methods. In MPO-deficient subjects, immunocytochemistry detects a marked decrease, but not absence, of MPO.

Benzidines

Myeloperoxidase deficiency: prevalence and clinical significance.

Leukocyte differential counting by flow cytochemistry has shown 28 subjects with partial or complete neutrophil myeloperoxidase (MPO) deficiency in a population of about 60 000 patients screened at a general hospital. Partial (13 patients) or complete (13 patients) MPO deficiency was confirmed by examination of cytochemical stains in 26, biochemical measurement of total enzymatic activity in eight, and flow cytometry in six patients. None had apparent hematologic disorders. Only four patients had infections; of these, two had major systemic infections (one, candidiasis; one, bacteremia). In assays of leukocyte function only minor defects in killing of Staphylococcus aureus by MPO-deficient cells were noted whereas killing of Candida albicans was much more impaired. Family studies in eight patients have shown various degrees of partial or complete MPO deficiency in first-degree relatives of six. The findings indicate that the incidence of MPO deficiency is much higher than previously suspected. Although MPO appears to be necessary for killing of Candida species by neutrophils, the importance of its role in normal antibacterial defense must be re-evaluated.

Adolescent

Monocyte contamination in Ficoll-Hypaque mononuclear cell concentrates.

Monocyte contamination in mononuclear concentrates prepared by Ficoll-Hypaque (F-H) centrifugation was assessed by the following criteria: (a) morphology (Wright-Giemsa stain), (b) function (phagocytosis), and (c) cytochemical staining (non-specific esterase both manually and by flow cytophotometry). Studies on F-H preparations from blood samples obtained from 50 randomly selected patients irmined by functional and cytochemical staining. Monocyte counts determined by flow cytophotometric studies of cells stained in suspension for esterase were very similar to those based on phagocytosis and manual cytochemical staining technics. It is concluded that: (1) manual Wright-Giemsa staining alone is not adequate to accurately quantitate monocyte populations in F-H concentrates, and (2) cytochemical staining and flow cytophotometry can be used for rapid and accurate quantitation of monocyte contamination in F-H mononuclear concentrates.

B-Lymphocytes

Pooled normal values, a useful technic.

Few laboratories determine their normal values, because of the work involved. This study attempted to see whether a group of laboratories, each contributing a portion of the samples, could pool their information to yield data applicable to the whole group. Each of 20 VA Hospital laboratories obtained approximately 20 fasting early morning specimens from healthy men, aged 20-29 years, during a four-week period. Routine chemistry examinations (16) were performed on Technicon AutoAnalyzers (12/60, 6/60, and AAII), and model S Coulter Counters were used for five hematology tests (381 values for each constituent). Within-day, day-to-day, and interlaboratory variance were determined in parallel-method variability studies. The data showed: (1) it is valid to pool normal values from different laboratories when non-random variables have been eliminated; (2) non-random variables can be identified by increased interlaboratory variance, whether differences such as methods and equipment are obvious or not; (3) normal values so derived agree well with those from large series performed in single laboratories.

Adult

Computer-assisted monocyte esterase assay by flow-cytophotometry.

A Wang model 2200 computer has been interfaced with the Bio/Physics Systems, Inc. model 6300 Cytograf and model 2100 Distribution Analyzer. Using a custom designed software program, in conjunction with an azo-dye technic for staining monocytes for nonspecific esterase activity, it has been possible to obtain rapid and reliable data concerning relative values for intracellular monocyte esterase activity. The method is based on measuring the axial light-loss voltage signal for each of one thousand stained monocytes. Individual stained monocytes were assigned to one of four groups (A, B, C, D), dependent upon the magnitude of the signal and were given different rating values (1, 2, 3, 4) according to their group designation. A "score" was derived for each blood sample by multiplying the percentage of cells (monocytes) in each group category by the appropriate factor and summing these values. The technic permits rapid objective assessment of intracellular nonspecific esterase activity in monocytes suspended in a mixed cell population. Both Gaussian and bi-modal patterns for monocyte esterase were observed. The latter suggests a dual monocyte population.

Computers

The application of cytochemistry to automation.

The application of cytochemical techniques for use with automated instruments introduces a variety of problems not encountered with traditional cytochemical stains prepared for human visualization. This is particularly true for flow-cytophotometers, which require that cells be stained in suspension, making it difficult to remove reagents once they are added to a staining mixture. It is much less a problem for pattern recognition instruments. The major difficulties relate to cell preparation, fixation, staining, controls and interpretation of instrumental data. The new cytochemical techniques or modifications of existing methods that are required for use with such instruments present a fresh challenge to cytochemists. The need for such methods is great and the rewards may be equivalent.

Fixatives

Assessment of monocyte esterase activity by flow cytophotometry.

An azo dye supravital method has been devised for selectively staining human monocytes in suspension for nonspecific esterase activity. Stained cells can be identified and rapidly enumerated by presenting the suspension of stained cells to the Cytograf, a flow-through cell discriminating cytophotometer. The intensity of stain is proportional to the intracellular esterase activity. By analysis of the oscilloscope display, it has been possible to obtain relative data concerning the degree of activity of monocyte nonspecific esterase activity. These observations suggest a unique approach to the measurement of intracellular enzyme activity in selected cells in a mixed population.

Autoanalysis