| Summary: | 碩士 === 國立成功大學 === 生物化學研究所 === 85 === Prothymosin a (ProTa) is an unusual acidic protein (PI=3.5)
that widely distributes in mammalian tissue. It is a kind of
thymic hormones associated with cell proliferation and has some
immunomodulating properties. The relation of ProTa to
proliferation has been well documented. ProTa mRNA expression is
correlated to the proliferative activity of T-cells. In serum
deprived NIH3T3 cells when they were stimulated to proliferate,
ProTa mRNA levels were also induced. An increased concentration
of ProTa in cancer tissue was reported. ProTa can restore the
human mixed lymphocytic response (MLR) in patients of autoimmune
diseases and enhance human and murine major histocompatibility
complex (MHC) class II surface antigen expression. However, the
mechanisms and real biological functions of ProTa are not very
clear. In order to further study its biochemical properties, we
produce monoclonal antibodies against human ProTa. The
recombinant human ProTa proteins were obtained from the E. coli
strain BL21(DE3)pLysS transformed with ProTa plasmid and
purified by high salt/phenol extraction and recovered by
enthanol precipitation. The SDS contamination in purification
procedure was eliminated by re-extraction with n-butanol. After
amino acid analysis and biological activity assay, we immunized
female BALB/c mice with recombinant human ProTa proteins. The
splenocytes were fused with NS-1 myeloma cells to prepare
hybridomas. Six hybridoma cell lines which produced monoclonal
antibodies against ProTa were obtained. Using one of the anti-
human ProTa monoclonal antibodies, we developed a competitive
ELISA system to quantify ProTa amount in urine samples of
bladder cancer patients. The results indicate a significant
difference of ProTa amount in urine between bladder cancer
patients and normal healthy people.
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