Immunocytological and Preliminary Immunohistochemical Studies of Prothymosin α, a Human Cancer–associated Polypeptide, With a Well-characterized Polyclonal Antibody

Abstract

Prothymosin α (ProTα) is a nuclear polypeptide of great biological and, possibly clinical, importance, because its expression levels have been associated with early diagnosis/prognosis of human cancer. It is therefore interesting to raise easily available and cost-effective antibodies that would be applied to develop reliable ProTα immunodiagnostics. In this study, New Zealand white rabbits and laying hens were parallel immunized against intact ProTα or the synthetic fragments ProTα[1-28], ProTα[87-109], and ProTα[101-109], all conjugated to keyhole limpet hemocyanin (KLH). The corresponding antibodies G and Y were immunochemically evaluated in parallel with ELISA and Western blot systems and applied to fluorescence immunocytology experiments using various cancer cell lines and normal cells. The antibody G raised against ProTα[101-109]/KLH had excellent functional characteristics in the Western blot and immunocytology experiments, where the fluorescent signal was almost exclusively shown in the cell nucleus independently of the cells assayed. The above antibody has been applied to preliminary IHC staining of human cancer prostate tissues, leading to a high percentage of clearly and intensively stained nuclei in the adenocarcinoma tissue; this antibody can be further used in cancer tissue immunostaining and in research concerning the role of ProTa in tumorigenesis.

Solid-Phase Synthesis of a Biotin Derivative and its Application to the Development of Anti-Biotin Antibodies

“…The above results were confirmed by suitably treating HeLa cells, so as to remove their cytoplasm, and then staining the cell nuclei. Contrary to that obtained with the anti-BAL antiserum and with streptavidin, clear nuclear staining (data not shown here) was observed with the positive control, i.e., a wellcharacterized rabbit antiserum recognizing the nuclear polypeptide prothymosin alpha [17].…”

“…A rabbit polyclonal antibody that recognizes the nuclear polypeptide prothymosin alpha [17] was used as a positive control for nuclear staining.…”

Prothymosin alpha: a ubiquitous polypeptide with potential use in cancer diagnosis and therapy

“…By immunohistochemistry and immunoblotting, they showed that proT levels consecutively increased as prostate tissue progressed from normal epithelium, through prostatic intraepithelial neoplasia to carcinoma [56]. Particularly, in prostate cancer lesions, intense nuclear staining for proT was also reported [57]. Subsequent complementary research investigated proT expression in human bladder cancer using tissue microarrays [58].…”

“…Subsequent complementary research investigated proT expression in human bladder cancer using tissue microarrays [58]. A statistically signiWcant enhancement of proT expression, also from the clinical aspect, was Colon mRNA in tissue c-myc expression [45,52] Gastric Protein in tissue - [63] Head and neck Protein in tissue Tumor recurrence [62] Liver mRNA in tissue; protein in tissue c-myc expression; poor prognosis [46,60] Lung mRNA in tissue Poor prognosis [48] Neuroblastoma mRNA in tissue N-myc expression [47] Pituitary Protein in tissue Tumor recurrence [61] Prostate Protein in tissue Cancer cell diVerentiation; tumor progression [56,57] Rectal mRNA in tissue EVectiveness of radiotherapy [51] Rhabdomyosarcoma mRNA in tissue - [49] Thyroid mRNA in tissue; protein in tissue and serum…”

4-Hexylresorcinol inhibits transglutaminase-2 activity and has synergistic effects along with cisplatin in KB cells

“…the antibody for tg-2 was purchased from santa cruz Biotechnology (santa cruz, CA, USA) for fluorescence immunocytochemistry analysis. Fluorescence immunocytochemistry was done using a previously published method (14). 4',6-Diamidino-2-phenylindole (DAPI) was used for count staining.…”