|Opis:||The celiac disease is one of the most common autoimmune disorders, occurring when gluten is ingested by genetically predisposed individuals; alongside environmental factors, a genetic predisposition contributes to the development of the celiac disease. Most of the individuals, suffering from this illness share either the HLA-DQ2 or HLA-DQ8 human leukocyte antigen genotype – the heterodimeric proteins, found on the surface of antigen-presenting cells. Several medical examinations must be performed before a diagnosis can be made. Among them are: small intestine biopsy, controlled trial of gluten free diet, serological tests, and genetic testing.
To determine whether an individual is prone to the disease, The European Society of Paediatric Gastroenterology, Hepatology and Nutrition (ESPGHAN) has recommended identifying the HLA-DQ2 and HLA-DQ8 genotypes, present in affected individuals. Based on this information, we have decided to compare two genetic diagnostic methods: The sequence specific primer polymerase chain reaction (SSP PCR), and real-time polymerase chain reaction (rt-PCR). The first is a polymerase chain reaction (PCR) where sequence specific primers – oligonucleotides are used, while the latter is a PCR, where the quantity of a product is measured in between every cycle of the reaction by measuring fluorescence. With the first method, Olerup diagnostics kit was used, while Eligene diagnostics kit was used with the second.
The analysis was conducted on six samples, of which four belonged to hospital patients, admitted for celiac disease. The results of both diagnostic tests do not match. We have concluded that the results, provided by the Eligene diagnostic kit, are in accordance with the medical diagnoses, while the same conclusions could not be made for the results, provided by Olerup. A comparison of the difficulty of implementation of individual detection kits has determined that the rt-PCR method is less complex to implement and more user friendly than the SSP PCR method, as it requires less preparation. The results can easily be interpreted by a single individual. Here, the only disadvantage is that a more expensive real-time machine is required. Diagnostics has proven more time consuming, using the Olerup kit; its implementation requires more time to prepare. It is conducted in two stages, the PCR reaction and the agarose gel fragment analysis. Even though the software equipment needed for the analysis is highly developed, the analysis itself is difficult to perform and requires professional assistance. If we were to choose between the two kits and methods, we would have chosen the rt-PCR method, using the Eligene kit. |