Erroneous immunohistochemical application of monoclonal antibody EG2 to detect cellular activation

Jahnsen, F; Halstensen, TrondS.; Brandtzæg, Per

doi:10.1016/s0140-6736(94)90336-0

Cited by 15 publications

(9 citation statements)

References 4 publications

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“…Furthermore, recent studies cast doubt on the reliability of this antibody in detecting activated eosinophils (Nakajima et al, submitted). 42,43 Taken together, our data provide direct biochemical and morphological evidence that eosinophils in PBC are not only increased in number but are also functionally activated.…”

Section: Discussionsupporting

confidence: 58%

Ursodeoxycholic acid inhibits eosinophil degranulation in patients with primary biliary cirrhosis

Yamazaki¹,

Suzuki²,

Nakamura³

et al. 1999

Hepatology

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Eosinophilia is a distinctive feature of primary biliary cirrhosis (PBC), especially in its early stages. Intriguingly, treatment with ursodeoxycholic acid (UDCA) ameliorates eosinophilia as well as liver tests in patients with PBC. It remains unknown, however, whether eosinophils in PBC patients are functionally activated and whether UDCA inhibits eosinophil activation. In the present study, we systematically examined eosinophil dynamics in the blood and liver in patients with stage I to II PBC before and after UDCA treatment. We determined serum concentrations of eosinophil granule proteins (major basic protein [MBP] and eosinophil-derived neurotoxin [EDN]) by radioimmunoassay and quantitated eosinophil degranulation using computer-assisted morphometry after MBP immunohistochemistry. Before UDCA treatment, patients with PBC (n ‫؍‬ 25) showed significantly higher circulating eosinophil counts (P F .05) and serum concentrations of MBP (P F .0005) and EDN (P F .02) compared with patients with chronic viral hepatitis (n ‫؍‬ 22), autoimmune hepatitis (n ‫؍‬ 10), and obstructive jaundice (n ‫؍‬ 12). Four-week UDCA treatment significantly reduced blood eosinophil counts (P F .0001) and serum MBP (P F .0001) and EDN (P F .0001) levels in PBC patients. MBP immunohistochemistry and computer-assisted quantitative morphometry showed infiltration and degranulation of eosinophils in the portal tract in patients with PBC and significant reductions in the number of sites and the area occupied by extracellular MBP deposits after UDCA treatment for 2 years (P F .02) but not in placebo-treated patients. Our results suggest that eosinophils in patients with PBC are not only increased in number, but also release granule proteins, and that UDCA treatment inhibits this eosinophil activation/degranulation. (HEPATOL-OGY 1999;30:71-78.)

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Section: Discussionsupporting

confidence: 58%

Ursodeoxycholic acid inhibits eosinophil degranulation in patients with primary biliary cirrhosis

Yamazaki¹,

Suzuki²,

Nakamura³

et al. 1999

Hepatology

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“…Subsequently, EG2 has been widely employed to identify activated eosinophils in tissues; over 150 citations in the literature have reported on this use. Recent reports, however, demonstrated that EG2 reactivity was highly dependent on sample preparation and fixation, and these reports have questioned the use of EG2 as a specific marker for activated eosinophils [19][20][21][22]. In this study, therefore, we examined the reactivity of EG1 and EG2 for ECP and EDN by RIA, Western blot, and immunofluorescence analyses to further assess the ability of EG2 to distinguish between resting and activated eosinophils.…”

Section: Discussionmentioning

confidence: 98%

“…19 for a list of these articles]; indeed, a Medline search for EG2 revealed over 150 citations. However, more recent reports have disputed the ability of EG2 to discriminate between resting and activated eosinophils [19][20][21][22].…”

Section: Introductionmentioning

confidence: 99%

Reactivity of monoclonal antibodies EG1 and EG2 with eosinophils and their granule proteins

Nakajima

Loegering

Kita

et al. 1999

Journal of Leukocyte Biology

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Use of the murine monoclonal antibodies EG1 and EG2 has been based on the assumption that EG2 recognizes activated eosinophils. We examined the reactivity of EG1 and EG2 with eosinophil cationic protein (ECP), eosinophil-derived neurotoxin (EDN), and stimulated and nonstimulated eosinophils from normal donors. By radioimmunoassay, EG1 recognized only ECP, whereas EG2 recognized both ECP and EDN. By Western blot, EG1 reacted with ECP, EG2 reacted with both ECP and EDN, but EG2 could not distinguish between lysates of stimulated and nonstimulated eosinophils. By immunofluorescence, EG1 and EG2 at 20 g/mL stained 95-100% of nonstimulated eosinophils, regardless of fixative; EG1 and EG2 at 0.1 g/mL stained 61-90% of acetone-and paraformaldehyde-fixed and only 5-21% of methanol-fixed nonstimulated eosinophils. Thus, the reactivity of EG1 and EG2 with eosinophils depends on the method of fixation and antibody concentration; and EG2, in contrast to previous reports, cannot reliably discriminate between resting and activated eosinophils. J. Leukoc. Biol. 66: 447-454; 1999.

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“…In contrast to EG1, which recognizes both the stored and secreted forms of ECP (22), EG2 is thought to recognize only active and secreted forms of ECP and has been taken to identify activated eosinophils (22), although this is controversial (23). In the current study, EG2-positive cells were consistently fewer in number than EG1-positive cells, indicating that these two markers are not exactly synonymous, and our data suggest that salmeterol had more effect on total eosinophils than on EG2-positive cells.…”

Section: Discussionmentioning

confidence: 99%

An Antiinflammatory Effect of Salmeterol, a Long-acting β₂ Agonist, Assessed in Airway Biopsies and Bronchoalveolar Lavage in Asthma

Ward

Thien

et al. 1999

Am J Respir Crit Care Med

135

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The addition of long-acting beta(2) agonists to inhaled corticosteroid (ICS) therapy in symptomatic patients with asthma improves clinical status more than increasing the dose of ICS. It has been suggested that these benefits could be at the cost of an increase in airway inflammation, but few histopathological studies have been performed in the relevant group. In a double-blind, parallel-group, placebo-controlled study, we randomly assigned 50 symptomatic patients with asthma who were receiving ICS (range, 100 -500 microgram/d) to 12 wk of supplementary treatment with salmeterol (50 microgram twice daily) or fluticasone (100 microgram twice daily) or placebo. Bronchial biopsies and BAL were obtained from 45 patients before and after treatment and analyzed. After treatment with salmeterol there was no deterioration of airway inflammation as assessed by mast cells, lymphocytes, or macrophages in BAL or biopsies, but rather a significant fall in EG1-positive eosinophils in the lamina propria (from a median 18.3 to 7.6 cells/mm, p = 0.01), which was not seen after treatment with fluticasone. The only cellular effect of added fluticasone was a decrease in BAL lymphocyte activation. There was a concurrent improvement in clinical status, more marked with salmeterol than with increased ICS. Thus, adding salmeterol to ICS is not associated with increased "allergic" airway inflammation, but conversely with a complementary antieosinophil effect.

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Erroneous immunohistochemical application of monoclonal antibody EG2 to detect cellular activation

Cited by 15 publications

References 4 publications

Ursodeoxycholic acid inhibits eosinophil degranulation in patients with primary biliary cirrhosis

Ursodeoxycholic acid inhibits eosinophil degranulation in patients with primary biliary cirrhosis

Reactivity of monoclonal antibodies EG1 and EG2 with eosinophils and their granule proteins

An Antiinflammatory Effect of Salmeterol, a Long-acting β₂ Agonist, Assessed in Airway Biopsies and Bronchoalveolar Lavage in Asthma

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Erroneous immunohistochemical application of monoclonal antibody EG2 to detect cellular activation

Cited by 15 publications

References 4 publications

Ursodeoxycholic acid inhibits eosinophil degranulation in patients with primary biliary cirrhosis

Ursodeoxycholic acid inhibits eosinophil degranulation in patients with primary biliary cirrhosis

Reactivity of monoclonal antibodies EG1 and EG2 with eosinophils and their granule proteins

An Antiinflammatory Effect of Salmeterol, a Long-acting β2 Agonist, Assessed in Airway Biopsies and Bronchoalveolar Lavage in Asthma

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Product

Resources

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An Antiinflammatory Effect of Salmeterol, a Long-acting β₂ Agonist, Assessed in Airway Biopsies and Bronchoalveolar Lavage in Asthma