What you need to know when you go with the flow: pitfalls in the use of flow cytometry.
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Cell Separation↗
Biomedical subjects
M Pallis
Publications and source records attributed to M Pallis.
An electron microscopic study of muscle capillary wall thickening in systemic lupus erythematosus.
Capillary vessel wall thickening may contribute to the muscle fibre atrophy and impaired pulmonary gas transfer notable in SLE/MCTD. Using electron microscopy we measured the thickness of unilaminar capillary basement membranes (CBM) and quantified basement membrane laminae and pericyte layers in the skeletal muscle biopsies of 31 patients with SLE, including 14 with MCTD, and 11 controls. Capillary basement membrane thickness was significantly increased in SLE compared with controls (P < 0.01) and was significantly associated with the level of C3dg (P < 0.01). Patients with MCTD had significantly greater layering of pericytes and laminae compared with patients with SLE alone (P < 0.0001 for both variables). Multiple stepwise regression analysis indicated that ANA titre was a good predictor of pericyte layering (R = 0.63, P = 0.003) and of basement membrane lamination (P < 0.0001), with steroid therapy having a significant negative association with the number of laminae (P < 0.02). A negative association between KCO and the number of layers of basement membrane (R = 0.51, P = 0.01) was documented. However, multiple regression analysis revealed no association between parameters of thickened vessel walls and the presence of muscle fibre atrophy.
Adult↗
Quantitative analysis of lymphocyte CD11a using standardized flow cytometry.
CD11a is the alpha-subunit of the leucocyte adhesion and costimulation molecule LFA-1. We have refined the measurement of lymphocyte CD11a density with a FACScan using commercially available fluorescent beads for standardization and fluorescein-conjugated antibody to CD11a of known fluorescein: protein ratio. The fluorescence intensity of CD11a on peripheral blood CD4+ and CD8+ lymphocytes was measured in 60 healthy subjects. We demonstrated linear correlation between age and mean CD11a density (r = 0.47 for CD4+ cells, r = 0.71 for CD8+ cells). We established that there is a consistent logical cut off point at 4.3 x 10(3) bound antibody molecules between low-expressing and high-expressing subsets of CD8+ cells and we then investigated whether the variation in lymphocyte CD11a expression in healthy subjects was sufficiently small for the application of this method to the detection of abnormal groups or individuals. Analysis of the CD11a high subsets has high statistical power (> 99% in 60 subjects to detect a 25% difference) and good precision (< 4% differences). The advantages of the method for comparative studies of cell surface accessory molecules are discussed. We have also evaluated a frozen cell line for quality control, and demonstrated up-regulation of CD11a density on CD4+ and CD8+ cells measured in three patients with infectious mononucleosis.
Adolescent↗
Distribution of cell adhesion molecules in skeletal muscle from patients with systemic lupus erythematosus.
OBJECTIVE: To investigate the pathophysiology of perivascular mononuclear cell infiltrates observed in skeletal muscle from patients with systemic lupus erythematosus (SLE). METHODS: Immunocytochemical techniques were used to examine frozen 5 microns sections from the quadriceps needle muscle biopsy specimens of 14 patients with SLE (including seven with infiltrates) for the expression of the cytokine inducible adhesion molecules intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin. RESULTS: Vessels in 6/7 SLE biopsy specimens with perivascular infiltrates expressed VCAM-1 at a density of > 2 vessels/mm2 in contrast with 0/7 SLE biopsy specimens without infiltrates and 1/6 control specimens. In contrast, E-selectin expression was increased ( > 0.3 positive vessels/mm2) in SLE biopsy specimens compared with control tissue regardless of the presence of perivascular infiltration. VCAM-1 and ICAM-1 were also noted on extravascular cells in the infiltrates, being particularly prominent on the intermuscle fibre dendritic processes of CD68+ HLA-DR+ cells. CONCLUSION: Expression of these cytokine inducible adhesion molecules may be important in the migration of mononuclear cells into skeletal muscle and may be involved in intercellular interactions within the tissue.
Adult↗
Nailfold capillary density as a possible indicator of pulmonary capillary loss in systemic lupus erythematosus but not in mixed connective tissue disease.
Nailfold capillary density was measured in 24 patients with systemic lupus erythematosus (SLE), 14 with mixed connective tissue disease (MCTD) and 21 healthy subjects. Pulmonary function tests were performed on all subjects and needle muscle biopsies on 12 patients with SLE and 9 with MCTD. A significant correlation was documented between nailfold capillary density and pulmonary gas transfer (KCO) in patients with SLE (p less than 0.001) but not in patients with MCTD. This suggests that in SLE poor gas transfer may be dependent on alveolar capillary loss and that nailfold capillary density may be a good indicator of alveolar capillary density. There was no significant correlation between skeletal muscle fiber atrophy and nailfold capillary density in SLE or MCTD. Additional studies to optimize the nailfold capillary counting method are described.
Adolescent↗