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Analysis of serological cross-reactivity between antibodies against BK polyomavirus variants
Caisová, Helena ; Horníková, Lenka (advisor) ; Roubalová, Kateřina (referee)
BK polyomavirus (BKPyV), which asymptomatically infects about 80 % of the human population, can reactivate in immunosuppressed patients after kidney transplantation and cause nephropathy. In the European population, there are predominantly two BKPyV sub- types, BKPyV-I and BKPyV-IV, which behave as different serotypes. Recipients who receive a kidney from a donor positive for a different subtype are at a greater risk of graft rejection. So far, it is impossible to distinguish between these two subtypes using a serological ELISA test as serum antibodies against these two subtypes cross-react. This work aimed to iden- tify epitopes on the major antigenic protein VP1, which are responsible for the binding of cross-reacting and/or subtype-specific antibodies. The identification of such epitopes could further lead to improvement in pre-transplant diagnostics and better management of pati- ents after transplantation. Therefore, two types of antigens composed of the VP1 protein of the BKPyV-IV subtype with introduced mutations characteristic of BKPyV-I in the DE and EF loops of VP1 were prepared and tested using an antigen competition assay with a panel of human sera. The results showed that the DE loop region is an immunogenic epitope that binds specific antibodies for BKPyV subtype I. Conversely, a mutation in...
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Laboratory diagnostics of Hepatitis B
JANŮ, Denisa
Hepatitis B is an inflammatory viral disease of the liver. The etiological agent is a DNA virus from the Hepadnaviridae family. It is transmitted through the blood and secretions of an infected person. The greatest danger of the disease lies in the transition to chronicity. Chronic infection carries a high risk of developing liver cirrhosis and hepatocellular carcinoma. Hepatitis B is widespread throughout the world. The most reliable prevention is vaccination and avoidance of risky behavior and contact with persons with risky behavior, especially drug users and promiscuous persons. The aim of this bachelor thesis is to get acquainted with the disease of viral hepatitis B and to describe the current possibilities of diagnostics of hepatitis B markers using the COBAS e601 device from ROCHE by electrochemiluminescent immunoassay and quantification of viral DNA on the COBAS 4800 from ROCHE by real time PCR and evaluation of results using both methods. The set of examined samples comes from the laboratory of infectious serology and virology of the company VIDIA-DIAGNOSTIKA. The results show that HBsAg is the most investigated marker of hepatitis B. Positive findings make up only 0.67%. The ratio of HBsAg positive men and women in 2019 was almost balanced. Most HBsAg positive patients were aged 30-40 years. I also examined the prevalence of anti-HBc Ig total antibodies. The positivity of anti-HBc Ig total was confirmed in 4.27% of examinations. The ratio of anti-HBc Ig total positive men and women in 2019 was also almost balanced and the most positive were at the age of 30-40 years.
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Toxocara canis
Žáková, Barbora ; Voxová, Barbora (advisor) ; Vejsová, Marcela (referee)
The aim of this work is to compare seroprevalence against Toxocara of patients from FN HK between 2017 and 2021. Finally these results are compared with the results of the study of PharmDr. Barbora Voxová and Čermáková from 2009-2013. The methods of investigation were chosen indirect serological. It was testing by ELISA methods on present of IgG antibodies in the serum in both of studies (from 2009-2013 and 2017-2021). If the presence of antigen in serum was confirmed, an avidity test was made. This avidity test was made by modified ELISA method using of urea. During period 2017 to 2021 a total of 609 patients were examined (282 men and 327 women). The largest group of patients consisted of patients aged 50-60, the smallest group of patients was over 80 years. 7 % of patients had positive result (47 % were men and 53 % were women). The most positive (11) were in the age group 70-80 years, the least (0) in the age group 0-10 years. 31 % of positive had low avidity, 8 % had borderline avidity and 61 % had high avidity. During period 2009 to 2013 a 1083 patients were examined. 10 % of patients had positive result (53 % were men and 47 % were women). 11 % positive had low avidity and 89 % had high avidity. The number of examinations decreased rapidly between the two studies. In theyears 2009-2013 there...
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Molecular and serologic diagnosis of infections caused by Trichobilharzia
Vaščiková, Michaela ; Horák, Petr (advisor) ; Ditrich, Oleg (referee)
Cercariae of the genus Trichobilharzia can penetrate not only the skin of definitive hosts (ducks), but they are also able to penetrate the skin of accidental hosts (mammals). As a result of the penetration, the inflammatory response known as cercarial dermatitis appears. The goal of our thesis is to detect parasite DNA in the serum and cerebrospinal fluid of infected ducks, and also in the serum of infected mice. By using PCR with primers designed for a tandem repeated sequence, we were able to detect 1 femtograms of parasite DNA isolated from sera of infected ducks. We were able to amplify parasite DNA only from 16 samples of sera and cerebrospinal fluid of the infected ducks, but we were not able to do so with the serum of mice. Sera of infected mice were also tested by ELISA and Western blot. The homogenates of T. regenti (TRhc), T. szidati (TShc) and S. mansoni (SMhc) cercariae were selected as an antigen. The results showed progressive increase in the level of IgM antibody from 10 days after 1st infection and also increase of the level of IgG from the 2nd infection. 10 days after the 4th infection, the level of IgM and IgG gradually declined, but the level of antibodies 100 days after the 4th infection was still higher if compared to uninfected mice. Results from Western blot analysis...
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Molecular and serologic diagnosis of infections caused by Trichobilharzia
Vaščiková, Michaela ; Horák, Petr (advisor) ; Ditrich, Oleg (referee)
Cercariae of the genus Trichobilharzia can penetrate not only the skin of definitive hosts (ducks), but they are also able to penetrate the skin of accidental hosts (mammals). As a result of the penetration, the inflammatory response known as cercarial dermatitis appears. The goal of our thesis is to detect parasite DNA in the serum and cerebrospinal fluid of infected ducks, and also in the serum of infected mice. By using PCR with primers designed for a tandem repeated sequence, we were able to detect 1 femtograms of parasite DNA isolated from sera of infected ducks. We were able to amplify parasite DNA only from 16 samples of sera and cerebrospinal fluid of the infected ducks, but we were not able to do so with the serum of mice. Sera of infected mice were also tested by ELISA and Western blot. The homogenates of T. regenti (TRhc), T. szidati (TShc) and S. mansoni (SMhc) cercariae were selected as an antigen. The results showed progressive increase in the level of IgM antibody from 10 days after 1st infection and also increase of the level of IgG from the 2nd infection. 10 days after the 4th infection, the level of IgM and IgG gradually declined, but the level of antibodies 100 days after the 4th infection was still higher if compared to uninfected mice. Results from Western blot analysis...
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Molecular biology methods and their use for HLA antigen polymorphism determination
Brožová, Jitka ; Slavčev, Antonij (advisor) ; Trošan, Peter (referee)
HLA (Human Leukocyte Antigen) is a gene segment localized on the short arm of chromosome 6. This segment consists of three regions, I. and II. region encode HLA class I and HLA class II genes, in the III. region there are the complement genes, TNFα and other genes that are not evolutionary and functionally associated with HLA genes. HLA class I and class II genes encode cell-surface glycoproteins which in their binding sites present peptides to cells of the immune system. HLA genes are associated with many autoimmune and infectious diseases, although mostly the mechanisms of these associations remain unclear. HLA function as strong transplantation antigens, thus they represent a major obstacle in allogeneic organ transplantation and transplantation of hematopoietic cells. To prevent rejection of the transplanted organ, a match between the recipient and donor in the HLA antigens is necessary. Determination of donor and recipient matching is performed by HLA typing techniques. Nowadays for HLA typing in most laboratories in Europe and the USA are used methods of molecular biology. Among these methods, three main techniques based on the polymerase chain reaction (PCR) are applied, these are PCR-sequence-specific primers, PCR-sequence-specific oligonucleotide probes and PCR-sequencing-based typing....
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