A simplified procedure for the preparation of antibodies to serum fibronectin

Zardi, Luciano; Siri, Annalisa; Bárbara, Cristina; Cosulich, Elisabetta; Viale, Giovanna; Santi, Leonardo

doi:10.1016/0022-1759(80)90169-6

Cited by 56 publications

(22 citation statements)

References 16 publications

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“…The preparation was rinsed three times with PBS, and a drop of FITCconjugated antiserum to fibronectin was deposited on the tissue section. Fluorescence was observed in a fluorescent light microscope after incubation for 30 min with the antibody, subsequent rinsing with PBS, and exposure to 90010 glycerol [31]. Immunofluorescent fibronectin was observed after incubation of lung tissue sections with cystic fibrosis bronchial secretions.…”

Section: Assessment Of Proteolysis Of Cellular Fibronectinmentioning

confidence: 99%

Fibronectin-Cleaving Activity in Bronchial Secretions of Patients with Cystic Fibrosis

Suter

Schaad²,

Morgenthaler³

et al. 1988

Journal of Infectious Diseases

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In cystic fibrosis, colonization of the airways with Pseudomonas aeruginosa follows colonization with Staphylococcus aureus and is related to accelerated deterioration of pulmonary function. Because P. aeruginosa adheres better to cell surfaces devoid of fibronectin, we searched for fibronectin-cleaving activity in bronchial secretions and saliva from 24 patients with cystic fibrosis who were followed up for 4.5 y and from two control groups. Proteolytic activity against 12sI-labeled fibronectin was continuously present in cysticfibrosis bronchial secretions; significantly higher fibronectin-cleaving activity was found in older vs. younger patients, in patients with advanced disease stages determined by a five-stage scoring system, and in those colonized with P. aeruginosa. The fibronectin-cleaving activity was due to neutrophil elastase and cathepsin G. Cystic fibrosis bronchial secretions had proteolytic activity against surface fibronectin of airway mucosal cells. Thus fibronectin-cleaving activity of bronchial secretions rather than of saliva may favor P. aeruginosa colonization of the upper respiratory tract in individuals with cystic fibrosis.Most individuals with cystic fibrosis who survive beyond the neonatal period die from respiratory failure, which is the consequence of chronic, progressively destructive bronchitis [1][2][3]. Clinical and pathological observations show extensive tissue destruction in the airways, which first causes obstruction and obliteration of small airways and thereafter destruction of the walls of large airways [4,5]. Early in the disease course the bacterial pathogens frequently found in the bronchial secretions are Staphylococcus aureus and Haemophilus influenzae. Later the airways of these subjects are invariably colonized with Pseudomonas aeruginosa, an event that initiates an acclerated deterioration of lung functions [3] as wellas an excessive systemic immune response [6]. These pathogens cannot be eliminated from the airways by antimicrobial therapy, and because the deleterious effect of P. aeruginosa infection in individuals with cystic fibrosis is well established, the identification of factors favoring colonization of the airways with P. aeruginosa is important. Coloniza-

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Section: Assessment Of Proteolysis Of Cellular Fibronectinmentioning

confidence: 99%

Fibronectin-Cleaving Activity in Bronchial Secretions of Patients with Cystic Fibrosis

Suter

Schaad²,

Morgenthaler³

et al. 1988

Journal of Infectious Diseases

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“…Fibronectin was purified from pooled human plasma by affinity chromatography on gelatin-Sepharose 4B (Pharmacia Fine Chemicals, Piscataway, NJ) [17]. Plasma proteins that bound nonspecifically to the matrix were first removed with a column packed with unmodified Sepharose [18]. Plasma proteins that were weakly bound to the gelatin-Sepharose were washed off with 1 M urea.…”

Section: Implantation Ofmta Coverslips Into Guinea Pigsmentioning

confidence: 99%

“…:;-Q x fibronectin. Rabbit antibodies to human fibronectin were prepared essentially according to Zardi et al [18]. Antiserum from immunized rabbits was adsorbed onto insolubilized fibronectin-free plasma proteins [18] and was shown to be monospecifie by immunoelectrophoresis.…”

Section: Implantation Ofmta Coverslips Into Guinea Pigsmentioning

confidence: 99%

“…Rabbit antibodies to human fibronectin were prepared essentially according to Zardi et al [18]. Antiserum from immunized rabbits was adsorbed onto insolubilized fibronectin-free plasma proteins [18] and was shown to be monospecifie by immunoelectrophoresis. The following immunoglobulin fractions were obtained from the monospecific antiserum by ammonium sulfate precipitation: (1) total immunoglobulins (IgG); (2) inimunoglobulins depleted of antifibronectin activity, which were designated dIgG and were obtained after selective removal of antibodies to fibronectin by affinity chromatography on immobilized fibronectin; and (3) affinity-purified antibodies to fibronectin, which were designated anti-FN IgG and were obtained by elution of the immunoglobulins previously adsorbed on the fibronectin-Sepharose gel.…”

Section: Implantation Ofmta Coverslips Into Guinea Pigsmentioning

confidence: 99%

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Foreign Body Infection: Role of Fibronectin as a Ligand for the Adherence of Staphylococcus aureus

Vaudaux¹,

Suzuki²,

Waldvogel³

et al. 1984

Journal of Infectious Diseases

144

118

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Foreign bodies made of polymethylmethacrylate coverslips were implanted subcutaneously into guinea pigs, were explanted four weeks later, and were tested for in vitro adherence of Staphylococcus aureus strain Wood 46. In the presence of serum, the level of staphylococcal adherence to explanted coverslips was 20 times higher than that of adherence to unimplanted coverslips. Adherence to explanted coverslips was caused by fibronectin deposits on the foreign body surface and was inhibited in a dose-related fashion by specific antibodies to fibronectin.

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“…Indirect immunofluorescence staining with anti-fibronectin antibodies (a kind gift from Dr L. Zardi, Institute of Oncology, University of Genoa, Italy ;Zardi et al, 1980) was carried out on IAR-6 and IAR-6-1 liver cell lines, as described by Yamada (1978). Cell cultures at logarithmic phase of growth were washed x 4-5 with warm culture medium and fixed in 4% buffered formaldehyde solution for 30 min at room temperature.…”

Section: Methodsmentioning

confidence: 99%

Surface proteins in normal and transformed rat liver epithelial cells in culture

Bannikov¹,

Vincent²,

Montesano³

1980

Br J Cancer

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Summary.-The pattern of surface proteins of different types of normal and transformed rat liver cells have been studied in culture by means of lactoperoxidasecatalysed iodination procedures, followed by SDS-gel electrophoresis. The cells examined were primary cultures of epithelial liver cells, long-term cultures of epithelial liver cells, in vitro transformed epithelial liver cell lines and liver tumourcell lines; mesenchymal cells from liver and skin were also examined. The principal surface proteins of primary cultures of epithelial cells from adult or neonatal rats had components with mol. wts of 140,000-160,000, 100,000 and 40,000-70,000. A band that had the same position as fibronectin from mesenchymal cells was also present and this band, as well as other iodinated components, were less sensitive to trypsin than fibroblastic fibronectin. A similar pattern of iodinated proteins was seen in long-term cultures of epithelial liver cells, with a great reduction in the number and intensity of the bands in the mol. wt region below 100,000. Almost all the in vitro transformed and tumour epithelial cell lines contain a protein with a mol. wt 135,000 as one of the major iodinated bands, and in contrast to the observation in transformed fibroblasts, the fibronectin was retained by most of these transformed cell lines.

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A simplified procedure for the preparation of antibodies to serum fibronectin

Cited by 56 publications

References 16 publications

Fibronectin-Cleaving Activity in Bronchial Secretions of Patients with Cystic Fibrosis

Fibronectin-Cleaving Activity in Bronchial Secretions of Patients with Cystic Fibrosis

Foreign Body Infection: Role of Fibronectin as a Ligand for the Adherence of Staphylococcus aureus

Surface proteins in normal and transformed rat liver epithelial cells in culture

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