Heat serum inactivation as a mandatory procedure for antiactin antibody detection in cell culture

Cançado, Eduardo Luiz Rachid; Vilas-Boas, Lucy Santos; Abrantes-Lemos, C.P.; Novo, Neil Ferreira; Porta, Gilda; Silva, L C Da; Laudanna, Antônio Atílio

doi:10.1002/hep.510230525

Cited by 38 publications

(20 citation statements)

References 31 publications

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“…In a previous study from our group using fibroblasts grown on glass slides, only heat serum inactivation was demonstrated to be efficient. A preincubation of fibroblasts with a calcium-chelating solution did not improve AAA identification, if heat serum inactivation was not concomitantly performed [3]. The discrepancy between the results of the Swedish studies and ours was probably due to a misinterpretation by the authors, because they had also diluted the serum samples in the same calciumchelating solution, in which they had preincubated the substrate.…”

Section: Discussioncontrasting

confidence: 76%

“…This schedule was used not only for fluorescein-labelled AAA reactivity (in green) but also for rhodamine-labelled F-actin (in red). All sera reactive to AAA were titred to extinction dilution, except those which were reactive with unmodified serum (technique A), because of the well-known worse, and, even negative results at lower dilutions in this condition [3].…”

Section: Methodsmentioning

confidence: 99%

“…Indirect immunofluorescence using human fibroblasts is one of the preferred methods for anti-actin antibody identification (AAA). However, in low dilutions (<1:80), heat serum inactivation is a necessary procedure for eliminating a thermolabile serum factor which interferes with AAA reactivity [3]. This interfering factor could be interacting with F-actin, the most probable target antigen of SMA in AIH, through an actin-severing protein [4].…”

Section: Introductionmentioning

confidence: 99%

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Thermolabile and Calcium-dependent Serum Factor Interferes with Polymerized Actin, and Impairs Anti-actin Antibody Detection

Cançado

Abrantes-Lemos

Vilas-Boas

et al. 2001

Journal of Autoimmunity

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Section: Discussioncontrasting

confidence: 76%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Thermolabile and Calcium-dependent Serum Factor Interferes with Polymerized Actin, and Impairs Anti-actin Antibody Detection

Cançado

Abrantes-Lemos

Vilas-Boas

et al. 2001

Journal of Autoimmunity

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“…As described in patients aVected by AH, 34 AAA detected in CD seem to be specific for polymerised actin, and IF of fibroblasts or HEp-2cells represents the most widely used method of AAA detection. 34 35 In fact only a few of our patients, in the CD and AH groups, reacted with unpolymerised actin in the western blotting and ELISA experiments.…”

Section: Discussionmentioning

confidence: 99%

Immune Reaction Against Cytoskeleton in Coeliac Disease

Clemente

Frau

Brusco

et al. 1999

Journal of Pediatric Gastroenterology & Nutrition

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Background-The cytoskeleton actin network of intestinal microvilli has been found to be rapidly impaired after gluten challenge in coeliac disease (CD). The aim of this study was to investigate the presence of an immune reaction towards cytoskeleton structures such as actin filaments in CD. Methods-Eighty three antiendomysial antibody positive CD patients (52 children and 31 adults) were studied at our outpatient clinics from 1996 to 1998 using indirect immunofluorescence, ELISA, and western blotting for antiactin (AAA) and antitissue transglutaminase (TGA) antibodies before and after a gluten free diet (GFD). Sixteen patients with smooth muscle antibody positive autoimmune hepatitis, 21 with inflammatory bowel diseases, seven with small bowel bacterial overgrowth, and 60 healthy subjects were studied as controls. Results-Fifty nine of 83 CD patients (28/31 adults (90.3%); 31/52 children (59.6%)) were positive for IgA and/or IgG AAA. Seventy seven (92.7%) were positive for IgA TGA. IgA AAA were strongly correlated with more severe degrees of intestinal villous atrophy (p<0.0001; relative risk 86.17). After a GFD, AAA became undetectable within five months. Conclusions-Apart from the immune reaction against the extracellular matrix, we have described an immune reaction against the cytoskeleton in both children and adults with CD. As AAA are strongly associated with more severe degrees of villous atrophy, they may represent a useful serological marker of severe intestinal atrophy in CD. (Gut 2000;47:520-526)

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“…Antinuclear and anti-actin antibodies were processed by the same technique in acetone-fixed human fibroblasts as the substrate (22). Western blotting to identify the antigens reactive to antimitochondrial antibodies was carried out using commercial enzyme (Sigma, St. Louis, MO, USA) and bovine heart antigens (23).…”

Section: Autoantibody Profilementioning

confidence: 99%

Detection of human parvovirus B19 in a patient with hepatitis

Pinho

Alves

Vieira

et al. 2001

Braz J Med Biol Res

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Parvovirus B19 has been associated by some investigators with cases of severe hepatitis. The aim of the present study was to determine the presence of active parvovirus B19 infection among 129 Brazilian patients with non-A-E hepatitis. The patients were assayed for antibodies against parvovirus B19, IgM class, by ELISA. In IgM-positive cases, parvovirus B19 DNA was assayed by PCR in serum and liver tissue and parvovirus VP1 antigen in liver tissue was assayed by immunohistochemistry. Antibodies against parvovirus B19, IgM class, were detected in 3 (2.3%) of 129 patients with non-A-E hepatitis. Previous surgery and blood transfusions were reported by these 3 patients. One patient was a 56-year-old female with severe hepatitis, with antimitochondrial antibody seropositivity and submassive necrosis at liver biopsy, who responded to corticosteroid therapy. Strong evidence for active parvovirus B19 infection was found in this patient, with parvovirus B19 DNA being detected by PCR in liver tissue. Furthermore, parvovirus VP1 antigen was also detected in liver tissue by immunohistochemistry. The other two IgM-positive patients were chronic hepatitis cases, but active infection was not proven, since neither viral DNA nor antigen were detected in their liver tissues. This and other reports suggest a possible relation between parvovirus B19 infection and some cases of hepatitis.

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Heat serum inactivation as a mandatory procedure for antiactin antibody detection in cell culture

Cited by 38 publications

References 31 publications

Thermolabile and Calcium-dependent Serum Factor Interferes with Polymerized Actin, and Impairs Anti-actin Antibody Detection

Thermolabile and Calcium-dependent Serum Factor Interferes with Polymerized Actin, and Impairs Anti-actin Antibody Detection

Immune Reaction Against Cytoskeleton in Coeliac Disease

Detection of human parvovirus B19 in a patient with hepatitis

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