RESUMO
Bone mineral density (BMD) reference data of non-Caucasian women is scarce but greatly needed for African-American women. The objective of this study was to establish a metacarpal normative reference database for African-American women using digital X-ray radiogrammetry (DXR) and hand radiographs and compare these values to existing Caucasian data. Two hundred and fifty healthy African-American women between the ages of 20 and 79 years old, 14 of whom were excluded, were recruited to participate from four different clinical sites. The study population was recruited in approximately equal number into the following groups: 20-29, 30-39, 40-49, 50-59, 60-69 and 70-79 years of age. A radiograph was acquired of each subject's non-dominant hand. The radiographs were scanned and analyzed using radiogrammetric techniques, and the BMD, MCI (Metacarpal Index), bone width and cortical thickness were calculated. The regression curve that best fit the data was a second order polynomial. The BMD and MCI of young adult women (20-40 years of age) were used to calculate T-score parameters. The young reference BMD and MCI with their associated standard deviations were found to be 0.6045 g/cm2+/-0.0529 g/cm2 and 0.5096 and 0.0792, respectively. However, the MCI was found to be approximately 2.5% lower (-0.0118) compared to Caucasian women. The African-American metacarpal BMD was found to be 3.5% (0.0207 g/cm2) higher across all ages when compared to existing Caucasian reference data acquired in a similar way. The differences were found to be entirely due to larger bone size, cortical diameter and bone width in the African-American women.
Assuntos
Negro ou Afro-Americano , Densidade Óssea/fisiologia , Ossos Metacarpais/fisiologia , Absorciometria de Fóton/métodos , Adulto , Idoso , Bases de Dados Factuais , Feminino , Humanos , Ossos Metacarpais/anatomia & histologia , Ossos Metacarpais/diagnóstico por imagem , Pessoa de Meia-Idade , Valores de Referência , População BrancaRESUMO
There has been increasing interest in attempts to harness the body's normal inflammatory response mediated through the eicosanoid pathway to treat tumors. Accumulating data indicate that the growth of several different cancers is modulated by a group of pro-inflammatory bioactive lipids, the best known of which are the eicosanoids. Eicosanoid pathway constituents modulate cell function in several important ways, and an agent that activates PLA(2) and up-regulates LTB(4) levels could be expected to be an effective cytotoxic tumor agent, especially if it stimulated NK cells. PLAP is a 28-kDa polypeptide that is a member of the WD-repeat protein, G-protein-transducin superfamily. The pro-inflammatory properties of PLAP have been elucidated using a number of different approaches. PLAP has been found in inflamed tissues and synovial fluid from patients with rheumatoid arthritis. Based on knowledge of PLAP as a pro-inflammatory agent, its capacity to modulate the immune response and the role of the inflammatory and immune responses in immune surveillance, the role of PLAP in cancer therapy was explored. Significant tumor regression was observed 72 hours following a single treatment with PLAP in an animal air pouch model of glioma. PEG-PLAP treatment increased the life expectancy of animals with Lewis lung cancer, and in preliminary studies in MTVL breast tumors in mice, PLAP treatment resulted in a similar increase in life expectancy. These findings suggest that PLAP holds promise as a potential therapy for cancer, and warrants further study.
RESUMO
An ideal form of cancer therapy is the harnessing of innate immunity to eradicate spontaneously arising clones of malignant cells. To date, attempts to develop effective immunotherapies have met with limited success. Prostaglandins and leukotrienes, collectively known as eicosanoids, are important mediators of immune and inflammatory responses. Harnessing these compounds could be a method to treat cancers. Eicosanoids are formed after cleavage of fatty acids from phospholipids by phospholipase enzymes. We have previously described, characterized and cloned a naturally occurring mammalian activator of phospholipase A2. Injection of a 24 amino acid peptide from this phospholipase A2 activating protein (PLAP), resulted in induction of an acute inflammatory response, and a concomitant regression of gliomas in rats. Administration of 500 micrograms of this protein resulted in a 50% decrease of the tumor mass within 72 h. Tumor regression coincided with a greater than twenty-fold increase in levels of prostaglandin E2(PGE2) and leukotriene B4(LTB4), and a marked infiltration of natural killer(NK) cells. These data suggest that activation of phospholipase A2 and modulation of the eicosanoid biosynthetic pathway may provide a novel therapeutic strategy for the successful treatment of malignant tumors of the nervous system.
Assuntos
Glioma/tratamento farmacológico , Glioma/enzimologia , Inflamação/enzimologia , Fosfolipases A/efeitos dos fármacos , Fosfolipases A/metabolismo , Proteínas/farmacologia , Sequência de Aminoácidos , Animais , Divisão Celular/efeitos dos fármacos , Divisão Celular/fisiologia , Dinoprostona/metabolismo , Relação Dose-Resposta a Droga , Feminino , Glioma/patologia , Inflamação/induzido quimicamente , Inflamação/metabolismo , Leucotrieno B4/metabolismo , Dados de Sequência Molecular , Necrose , Transplante de Neoplasias , Fosfolipases A2 , Ratos , Ratos Wistar , Coloração e Rotulagem/métodosRESUMO
OBJECTIVE: To demonstrate expression of transforming growth factor beta (TGF beta) and basic fibroblast growth factor (bFGF) by cultured rheumatoid arthritis (RA) synovial cells and to investigate their role as synovial cell mitogens. METHODS: Polypeptide growth factors were detected and identified by immunocytochemical staining and Western blot analysis. Messenger RNA (mRNA) transcripts encoding TGF beta and bFGF were identified by polymerase chain reaction analysis. The influence of neutralizing growth factor monoclonal antibodies (MAb) on RA synovial cell growth was investigated. TGF beta bioactivity was determined by Mv1Lu assay. RESULTS: Lysates of RA, as compared with normal, synovial cells contained greater amounts of TGF beta and bFGF. Western blot analysis identified a single TGF beta band (MW approximately 25 kd) in each of the cell lysates examined. Western blot analysis using MAb DE6 identified a doublet of bFGF bands (MW approximately 18.0 kd) in normal synovial cell lysates and 4 bFGF bands (MW approximately 18.0, 22.0, 22.6, and 25.2 kd) in RA synovial cell lysates. RA and normal synovial cells expressed mRNA transcripts encoding TGF beta 1 but not TGF beta 2, and FGF-2 (basic FGF). Additional mRNA transcripts encoding FGF-5 and FGF-7 were expressed by RA, but not normal, synovial cells in culture. In contrast to MAb 1D11.16, which caused a dose-dependent decrease in RA synovial cell growth, MAb DG2 (up to 100 micrograms/ml) had no effect on cell growth. CONCLUSION: RA and normal synovial cells cultured in serum-free medium express TGF beta 1 and native bFGF. However, only RA synovial cells in culture express higher molecular weight isoforms of bFGF. TGF beta 1 appears to regulate synovial cell growth in vitro through an external autocrine loop. Despite expression of high-affinity bFGF receptors on cultured synovial cells, the mechanisms by which bFGF modulates synovial cell growth are unknown.
Assuntos
Fator 2 de Crescimento de Fibroblastos/metabolismo , Membrana Sinovial/patologia , Fator de Crescimento Transformador beta/metabolismo , Artrite Reumatoide/patologia , Sequência de Bases , Células Cultivadas , Fator 2 de Crescimento de Fibroblastos/genética , Fator 2 de Crescimento de Fibroblastos/farmacologia , Expressão Gênica , Humanos , Imuno-Histoquímica , Sondas Moleculares/genética , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Membrana Sinovial/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/farmacologiaRESUMO
Cytokines have been implicated in the regulation of eicosanoid synthesis and synovial cell proliferation. To further define these mechanisms, we have compared the effects of basic fibroblast growth factor and platelet-derived growth factor on cell growth, prostaglandin E2 (PGE2) production and phospholipase A2 enzyme activity in long-term cultures of synovial cells from rheumatoid arthritis (RA) patients capable of proliferating in serum-free medium. Compared with serum-free medium alone, RA synovial cell growth was significantly enhanced by adding either basic fibroblast growth factor (bFGF) or platelet-derived growth factor (PDGF) to the culture medium. Growing RA synovial cells for 14 days in serum-free medium plus bFGF caused them to spontaneously release significant amounts of PGE2, an effect not seen if cells were grown in serum-free medium alone, or serum-free medium plus PDGF. Enhanced release of PGE2 occurred when arachidonic acid was added to bFGF but not PDGF-treated RA synovial cells, suggesting that bFGF increased cyclooxygenase enzyme activity in these cells. Moreover, phospholipase A2 (PLA2) enzyme activity was found to be significantly greater in RA synovial cells grown for 14 days in serum-free medium containing bFGF alone, or bFGF plus interleukin 1 beta (IL-1 beta) compared with cells grown in either serum-free medium alone, or serum-free medium plus PDGF. Similarly, bFGF plus IL-1 beta-stimulated release of PLA2 activating protein, a novel mammalian phospholipase stimulator found in high concentrations in RA synovial fluid.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Artrite Reumatoide/fisiopatologia , Divisão Celular/efeitos dos fármacos , Dinoprostona/metabolismo , Fator 2 de Crescimento de Fibroblastos/administração & dosagem , Interleucina-1/administração & dosagem , Fosfolipases A/metabolismo , Proteínas/metabolismo , Membrana Sinovial/citologia , Artrite Reumatoide/patologia , Células Cultivadas , Sinergismo Farmacológico , Ativação Enzimática/efeitos dos fármacos , Humanos , Técnicas In Vitro , Fosfolipases A2 , Fator de Crescimento Derivado de Plaquetas/farmacologia , Prostaglandina-Endoperóxido Sintases/metabolismoRESUMO
In vitro, rheumatoid arthritis (RA) synovial cells display several of the characteristics of neoplastic and virally transformed cells. The recent observation that synovial cell cultures, derived from collagenase digests of synovial membranes from RA patients, proliferate in serum-free medium suggests that these cells have the capacity to synthesize those factors essential for their growth. Direct immunocytochemical staining and Western analysis have identified transforming growth factor-beta (TGF-beta) band and basic fibroblast growth factor (FGF) in the cytoplasm of RA and normal synovial cells in long-term culture. Greater amounts of each growth factor were found in RA, as compared with normal synovial cell lysates. Western analysis identified a single TGF-beta band in RA and normal synovial cell lysates. Four bands were identified by Western analysis on RA synovial cell lysates probed with monoclonal antibodies recognizing bFGF, whereas only two bands (which co-migrated with human native recombinant bFGF) were identified in normal cell lysates probed with these antibodies. Gene expression analysis using PCR identified mRNA transcripts encoding TGF-beta 1 and FGF-2 (bFGF), but not TGF-beta 2 in all cell cultures studied. Taken together, these data indicate that cultured synovial cells co-express TGF-beta 1 and multiple isoforms of hFGF. These data further strengthen the concept that both polypeptide growth factors are involved in the regulation of synovial cell growth.
Assuntos
Substâncias de Crescimento/fisiologia , Líquido Sinovial/citologia , Animais , Artrite Reumatoide/etiologia , Artrite Reumatoide/patologia , Western Blotting , Divisão Celular , Humanos , Peptídeos/fisiologiaRESUMO
When stimulated with increasing amounts of interleukin 1 beta (IL 1 beta) rheumatoid arthritis (RA), as compared with osteoarthritis (OA), synovial cells grown in RPMI plus fetal bovine serum (FBS), released significantly more prostaglandin E2 (PGE2) (p less than 0.05; paired t test, two-tailed). PGE2 release by IL 1 beta-stimulated RA synovial cells grown for 14 days in serum-free RPMI was significantly less than that released by the same cells grown in medium plus 10% FBS (p less than 0.03; two-tailed). Since these data suggest that growth factors present in FBS may augment the effects of IL 1 beta, experiments were conducted to study the influence of four polypeptide growth factors--transforming growth factor-beta (TGF-beta), platelet-derived growth factor (PDGF), epidermal growth factor (EGF), and basic fibroblast growth factor (bFGF), on IL 1 beta-induced release of PGE2 by cultured RA synovial cells. Both EGF and bFGF significantly enhanced IL 1 beta-induced release of PGE2 (p less than 0.05; paired t test, one-tailed), while PDGF was synergistic with IL 1 beta, significantly increasing release of PGE2 by these cultured cells (p less than 0.02; two-tailed). No such effect was seen when TGF-beta was added to the culture medium. Taken together, these data lend support to the concept that within the synovial micro-environment small quantities of individual growth factors may potentiate the effects of IL 1 beta to amplify intra-articular inflammation.
Assuntos
Artrite Reumatoide/fisiopatologia , Dinoprostona/metabolismo , Substâncias de Crescimento/farmacologia , Interleucina-1/farmacologia , Células Cultivadas , Fator de Crescimento Epidérmico/farmacologia , Fator 2 de Crescimento de Fibroblastos/farmacologia , Humanos , Técnicas In Vitro , Osteoartrite/fisiopatologia , Fator de Crescimento Derivado de Plaquetas/farmacologia , Membrana Sinovial/fisiopatologia , Fator de Crescimento Transformador beta/farmacologiaRESUMO
Rheumatoid arthritis (RA), and not osteoarthritis (OA) synovial cells proliferate in serum-free medium, a finding that suggests that, in vitro, RA synovial cells may be stimulated to grow by the continuous autocrine production of at least one polypeptide growth factor. Adding monoclonal antibody 1D11.16, or rabbit polyclonal anti-tumor growth factor beta (anti-TGF-beta) antibodies (both neutralizing antibodies to TGF-beta 1 and TGF-beta 2) to RA synovial cells, in culture, caused a significant reduction in cell growth, an effect not seen when other growth factor antibodies (platelet-derived growth factor [PDGF], epidermal growth factor [EGF], or EGF receptor) were added to the culture medium. Taken together, these data are consistent with the concept that RA synovial cell growth in vitro is driven endogenous TGF-beta. Moreover, when EGF was added to the culture medium, this caused the numbers of RA, and not OA, synovial cells to increase significantly. This finding suggests that RA synovial cells are in G1 phase of the cell cycle; an effect that could be mediated by endogenous TGF-beta.
Assuntos
Artrite Reumatoide/patologia , Membrana Sinovial/patologia , Fator de Crescimento Transformador beta/fisiologia , Divisão Celular/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Humanos , Técnicas In Vitro , Osteoartrite/patologiaRESUMO
Rheumatoid factor (RF) test results reported in the College of American Pathologists' surveys for 1983-1985 lacked inter-laboratory reliability and mutual validity. Using the 4 most popular commercial kits for RF testing, participating laboratories consistently identified as "positive" or "negative" all but the weakly positive samples. A wide range of titers was reported on qualitative testing, however. One popular kit using a modified sheep red blood cell agglutination technique yielded results that differed markedly from those with other kits. Investigators apparently have paid little attention to these discrepancies. In Arthritis & Rheumatism, from 1983 to 1985, over 50% of the articles that referred directly or indirectly to RFs omitted details of RF methodology. Until a reliable RF test is adopted, it is essential that such methodologic information be specified.
Assuntos
Fator Reumatoide/análise , Humanos , Laboratórios , Métodos , Kit de Reagentes para Diagnóstico/normas , Padrões de ReferênciaRESUMO
Cardiac function was investigated in men with ankylosing spondylitis (AS) age 21-65 years who had no cardiorespiratory symptoms or known abnormalities of heart or lungs. Chest radiographs and standard electrocardiograms were normal in 73 of 74 subjects. In echocardiographs of 30 men, left atrial size and left ventricular cavity size and wall thickness were normal. Minor abnormalities in the valve roots were present in 3 older men. Early diastolic abnormalities of the left ventricle were demonstrated in 16 of 30 subjects. This finding was confirmed by repetition of the echocardiography a year later in 15 subjects and by comparison of 11 probands with their healthy brothers. Myocardial tissue obtained at necropsy from 28 AS patients without ischaemic or valvular heart disease or hypertension was studied. A mild, diffuse increase of interstitial connective tissue was seen but there was no inflammatory change or amyloid. Computerised image analysis showed 30.7% interstitial reticulin compared with 17.7% in age/sex matched controls (p less than 0.0001).
Assuntos
Contração Miocárdica , Espondilite Anquilosante/fisiopatologia , Adulto , Idoso , Ecocardiografia , Feminino , Histocitoquímica , Humanos , Masculino , Pessoa de Meia-Idade , Miocárdio/ultraestrutura , Espondilite Anquilosante/genética , Espondilite Anquilosante/patologiaRESUMO
Flurbiprofen has been compared with phenylbutazone in a double-blind study involving 33 patients with acute gout. Patients received either flurbiprofen 400 mg daily for 48 h followed by 200 mg daily, or phenylbutazone 800 mg daily for 48 h followed by 400 mg daily. The drugs were of comparable efficacy, while side-effects were uncommon and relatively mild. Flurbiprofen appears to be a satisfactory alternative to phenylbutazone in the management of acute gouty arthritis.
Assuntos
Flurbiprofeno/uso terapêutico , Gota/tratamento farmacológico , Fenilbutazona/uso terapêutico , Propionatos/uso terapêutico , Adulto , Idoso , Artrite/tratamento farmacológico , Ensaios Clínicos como Assunto , Método Duplo-Cego , Flurbiprofeno/efeitos adversos , Humanos , Pessoa de Meia-Idade , Fenilbutazona/efeitos adversos , Distribuição AleatóriaRESUMO
A modification of the glutaraldehyde-osmium tetroxide-tannic acid-uranyl acetate (GOTU) fixation procedure is described which allows human leucocytes to be examined subsequently by either transmission electron microscopy (TEM) or scanning electron microscopy (SEM).
Assuntos
Taninos Hidrolisáveis , Leucócitos/ultraestrutura , Taninos , Artrite Reumatoide/patologia , Fixadores , Humanos , Microscopia Eletrônica , Microscopia Eletrônica de VarreduraRESUMO
Changes in normal polymorphonuclear leucocyte (PMN) motility after membrane-binding and internalisation of IgG aggregates (a model of soluble immune complexes) have been studied by the micropore filter assay. The results have confirmed that IgG aggregates stimulate as well as inhibit PMN chemotaxis. These effects are dependent on the size and concentration of the IgG aggregates in solution as well as the length of time of incubation. Stimulated chemotaxis was observed in a small subset of the whole PMN population which was apparent only when cell distribution through the filters was analysed. These results indicate the need for caution when drawing conclusions about PMN function from results obtained by these assay techniques.
Assuntos
Complexo Antígeno-Anticorpo/imunologia , Quimiotaxia de Leucócito , Imunoglobulina G/imunologia , Humanos , Cinética , Filtros Microporos , Neutrófilos/imunologiaRESUMO
This study has investigated the chemotactic activity of polymorphonuclear cells (PMNs) isolated from the blood of patients with either articular rheumatoid arthritis (RA) or RA with extra-articular manifestations. A double fluorochrome immunofluorescent staining test has been employed to identify cell-associated immunoglobulins, probably immune complexes. The results suggest an inverse relationship between PMN chemotaxis and staining for cell-associated immunoglobulins, either surface bound or internalised. PMNs from RA patients showed reduced chemotaxis, and this was further reduced when RA PMNs were incubated for 30 minutes in autologous serum. A similar reduction in chemotaxis of normal PMNs occurred after incubation in RA sera. Preincubation of both RA and normal PMNs in RA serum (but not normal serum) resulted in an increase in the number of cells in which cell-associated immunoglobulins were demonstrable. This further reduction in RA PMN chemotaxis after exposure to autologous serum, together with an increase in immunoglobulin staining, may indicate selection of certain PMNs at the time of venepuncture due to cell margination. Such a selection process would call for a re-evaluation of previous studies of RA PMN function in relation to the disease process.
Assuntos
Artrite Reumatoide/imunologia , Quimiotaxia de Leucócito , Complexo Antígeno-Anticorpo/análise , Feminino , Humanos , Imunoglobulinas/análise , Corpos de Inclusão/imunologia , Masculino , Neutrófilos/imunologia , Receptores de Antígenos de Linfócitos B/análiseRESUMO
The use of therapeutic ultrasound within the Health Service is widespread and growing. The most common indication is to reduce inflammation. We have tested the influence of ultrasound on a model of acute inflammation in the rat, and we have found a complete absence of any anti-inflammatory action.
Assuntos
Inflamação/terapia , Terapia por Ultrassom , Animais , Flurbiprofeno/uso terapêutico , Inflamação/tratamento farmacológico , Irritantes , Poliuretanos , Ratos , Ratos Endogâmicos , PeleRESUMO
The validity of the antidromic method in the measurement of median and ulnar sensory nerve conduction is determined. Analysis of our results show that both the orthodromic and antidromic methods provide consistent results. Both methods were accurate in diagnosing Carpal Tunnel Syndrome (CTS), with an upper limit of normal for distal sensory latency of 4.5 msecs motor, and 4.0 msecs sensory in the median nerve. Variation with age and sex was studied. Latency when measured by either method showed no variation. Amplitude showed a consistent decrease with age. No sex difference was detected. We conclude that the antidromic stimulatory method is accurate, reproducible, and convenient, and therefore is at least as good if not better than the orthodromic method in the context of a busy routine electrodiagnostic clinic.
Assuntos
Síndrome do Túnel Carpal/diagnóstico , Condução Nervosa , Adolescente , Adulto , Fatores Etários , Síndrome do Túnel Carpal/fisiopatologia , Eletrodos , Eletromiografia/métodos , Potenciais Evocados , Feminino , Humanos , Masculino , Nervo Mediano/fisiopatologia , Pessoa de Meia-Idade , Fatores Sexuais , Nervo Ulnar/fisiopatologiaRESUMO
The aim of this study was to determine whether polymorphonuclear leukocyte chemotaxis, adhesion, and electrophoretic mobility were altered in inflammatory bowel disease and whether such alterations could be related to prior ingestion of immune complexes. Polymorphonuclear leukocytes from patients with ulcerative colitis and Crohn's disease showed significantly impaired stimulated migration (P less than 0.05), increased adhesiveness (P less than 0.01 in ulcerative colitis, P less than 0.001 in Crohn's disease), and reduced electrophoretic mobility (P less than 0.02 in ulcerative colitis, P less than 0.001 in Crohn's disease) compared with healthy controls. The disease control of patients with rheumatoid arthritis demonstrated reduced stimulated migration (P less than 0.025) but normal adhesion. Preincubating normal cells in inflammatory bowel disease sera suggested that the altered migration and adhesion were due to circulating serum factors. Circulating immune complexes, detected by the C1q PEG binding assay, were present in 12.5% of patients with ulcerative colitis and 30% with Crohn's disease. Direct immunofluorescence of polymorphonuclear leukocytes suggested binding and/or ingestion of complexes in 57% of patients with ulcerative colitis, and 67% with Crohn's disease. There was a direct correlation between positive immunofluorescence and impaired cell migration in ulcerative colitis (P less than 0.05), but no such relationship was found in the other parameters of polymorph function.
Assuntos
Colite Ulcerativa/fisiopatologia , Doença de Crohn/fisiopatologia , Neutrófilos/fisiologia , Complexo Antígeno-Anticorpo , Artrite Reumatoide/fisiopatologia , Adesão Celular , Quimiotaxia de Leucócito , Humanos , Imunoeletroforese , FagocitoseRESUMO
A discriminating direct immunofluorescent test has been used to identify immunoglobulin inclusions in polymorphonuclear leucocytes (PMNs) isolated from the blood of patients with rheumatoid arthritis. These inclusions are thought to represent phagocytosed immune complexes, since normal PMNs incubated in RA sera known to contain raised levels of immune complexes developed similar immunoglobulin inclusions. Inclusions did not develop in normal PMNs incubated in normal serum. No correlation was found between the percentage of either RA blood PMNs with immunoglobulin inclusions or normal PMNs developing inclusions after incubation in RA sera, and levels of immune complexes in the corresponding sera. Using heat-aggregated IgG as a laboratory model of immune complexes, a simple relationship has been demonstrated between the uptake of IgG aggregates by normal PMNs and the concentrations of IgG aggregates in the test solutions over a concentration range of 12.5-200 micrograms . ml-1. These results indicate that the C1q- PEG test gives no measure of the actual amounts of immune complexes available in serum for phagocytosis.
Assuntos
Complexo Antígeno-Anticorpo/imunologia , Artrite Reumatoide/imunologia , Imunoglobulinas/imunologia , Corpos de Inclusão/imunologia , Neutrófilos/imunologia , Artrite Reumatoide/sangue , Artrite Reumatoide/patologia , Imunofluorescência , Humanos , Imunoglobulina G/imunologia , MétodosRESUMO
Discrimination between immunoglobulin on the surface of polymorphonuclear leucocytes and immunoglobulin internalised as a result of phagocytosis has been achieved by direct immunofluorescent staining with fluorescein and rhodamine antiglobulin conjugates of identical specificity applied sequentially to live cells in suspension and after fixation on microscope slides. Surface bound antigen is recognised by double staining whereas internalised material is stained only by the conjugate applied to the fixed preparation. This differential staining procedure was evaluated in a model system in which normal PMNs were exposed to small (less than 20 s) and large (greater than 90 s) aggregates of human IgG.