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Antibody against bacterial lectins as a tool preventing lung infections of cystic fibrosis patients
Bolshova, Arina ; Hodek, Petr (advisor) ; Kubíčková, Božena (referee)
Cystic fibrosis is an autosomal recessive disease that is caused by mutation of the CFTR protein ("cystic fibrosis transmembrane conductance regulator"). The CFTR protein is an ion channel for chloride and bicarbonate ions transmission. This defect leads to the formation of dense mucus in lungs that causing breathing problems and chronic bacterial infections. Passive immunization by administering chicken antibodies produced against pathogen can be used as a prophylaxis for patients with cystic fibrosis. Immunoglobulins Y reduce adhesion of bacteria to lung epithelial cells and prevent infection. This strategy is a good alternative to classical antibiotic therapy because it targets the pathogen and doesn't affect normal bacterial microflora. At first the chicken antibodies produced after immunization of the experimental hen by Burkholderia cenocepacia lectin (BC2L-A) were isolated and further purified on an affinity column with covalently bound lectin. The affinity purified fraction was 9 times concentrated compared to original fraction as far as the content of the specific anti-lectin antibody is concerned. The amount of affinity purificated antibodies is approx. 0.2% of the total isolated immunoglobulins. Their immunoreactivity has been verified using recombinant lectin and the lectin produced by...
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Study of lectin-antibody interaction by using surface plasmon resonance
Zákopčaník, Marek ; Hodek, Petr (advisor) ; Nosková, Libuše (referee)
Cystic fibrosis is an autosomal recessive disease affecting mostly Caucasian race. The disease is caused by mutations in CFTR gene. The most affected organ system is respiratory tract, because the changes lead to insufficient natural defence function of lungs, causing the lungs being susceptible to bacterial infections, which are the most common cause of death of CF patients. Among these bacteria is also Burkholderia cepacia, which causes rapid degradation of pulmonary tissue. Eradication of this bacterial infection is complicated by a number of antibiotic resistances and biofilm formation. Chicken antibodies against bacterial lectins are potential means of preventing bacterial lung infection of patients with CF. Using the affinity chromatography with BC2L-A immobilized column a specific antibody fraction was obtained. The immunoreactivity of the mixed antibodies, affinity purified antibodies and unbounded fraction was compared by ELISA. The comparison of immunoreactivity has emerged that the concentration of the specific antibodies has grown more than 10 times. Interaction of lectin-antibody was studied with SPR method. In the first arrangement, the binding strength of the active lectin to glycosylation of the unbounded fraction antibodies was measured. The dissociation constant of this...
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Antibody against lectin PA-IIL as a tool preventing bacterial infections of cystic fibrosis patients
Vašková, Michaela ; Hodek, Petr (advisor) ; Nosková, Libuše (referee)
Cystic fibrosis is an autosomal recessive disease caused by mutation of the CFTR gene. This mutation results in the damage of protein with ion channel function. As a consequence of this damage, electrolyte equilibrium is impaired. The respiratory tract is affected the most because of the mucus thickening and changes in the glycosylation of lung epithelium cell surface structures. These changes lead to an insufficient defense function of lungs and greater susceptibility to bacterial infections. The most common pathogen of respiratory tract of CF patients is Pseudomonas aeruginosa. This bacterium contains numerous virulence factors and has the ability to form a biofilm that protects it against the host defense mechanisms and antibiotic effect. Antibiotic treatment is also complicated by the development of bacterium resistance. Chicken antibodies have a considerable potential as a tool for preventing bacterial lung infections of CF patients. The influence of specific and nonspecific antibodies on the adherence of Pseudomonas aeruginosa (PAK, ST 1763) ex vivo was monitored on a model system of lung epithelia cells from a CF patient (CuFi-1) and a healthy individual (NuLi-1). Bacterial and pulmonary cells were labeled with PKH fluorescent dyes to allow their spectrofluorimetric determination. It has...
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