Grancim B is a serine protease found primarily in cytotoxic granules of immune cells, cytotoxic T lymphocytes, and natural killer cells. It is a key mediator of triggering cell death in target cells, as a part of immune response. However, its expression is not limited to effector cells of the immune system. In many pathological conditions, its expression is increased in numerous tissues. For this reason, it is a suitable biological marker, useful for the purposes of diagnosing and understanding the course of various diseases. Bioluminescence is a special type of chemiluminescence observed in different types of organisms. It is an enzymatic reaction where the enzyme luciferase converts the substrate luciferin into an oxyluciferin. This releases energy in the form of light. Due to the very good ratio between ambient noise and reaction signal, it is suitable for analytical monitoring of a wide range of biological processes in vitro and in vivo. Substrate modifications of the bioluminescence reaction allow the development of probes for the purposes of visualization of processes both in tissue and in laboratory animals. In the master's thesis, we prepared a strain of P. pastoris, which expresses recombinant granzyme B. We confirmed the successful expression and secretion of active granzyme B into the medium. Recombinant granzyme B was compared with native and commercial recombinant granzyme B. We developed an in vitro bioluminescence imaging method for granzyme B detection. The selectivity of the bioluminescent probe Z-AAD-NHLuc for granzyme B was determined with the prepared recombinant granzyme B and compared to granzymes A, D, H, G and K
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