Production and characterization of monoclonal antibodies against human prostate specific antigen

ABSTRACT

Prostate Specific Antigen [PSA] is an important laboratory marker for diagnosis of prostatic cancer. Thus, development of diagnostic tools specific for PSA plays an important role in screening, monitoring and early diagnosis of prostate cancer. In this paper, the production and characterization of a panel of murine monoclonal antibodies [mAbs] against PSA have been presented. Balb/c mice were immunized with PSA, which was purified from seminal plasma. Splenocytes of hyperimmunized mice were extracted and fused with Sp2/0 cells. By adding selective HAT medium, hybridoma cells were established and positive clones were selected by ELISA after four times of cloning. The isotypes of produced mAbs were determined by ELISA and then purified from ascitic fluids using Hi-Trap protein G column. The reactivities of the mAbs were examined with the purified PSA and seminal plasma by ELISA and western blot techniques. Furthermore, the reactivities of the mAbs were assessed in Prostate Cancer [PC alpha], Benign Prostatic Hyperplasia [BPH] and brain cancer tissues by Immunohistochemistry [IHC]. Five anti-PSA mAbs [clones 2G2-B2, 2F9-F4, 2D6-E8, IgG1/K] and clones [2C8-E9, 2G3-E2, IgG2 alpha/K] were produced and characterized. All mAbs, except 2F9-F4 detected the expression of PSA in PCa and BPH tissues and none of them reacted with PSA in brain cancer tissue in IHC. Besides, all mAbs could detect a protein band around 33 kD alpha in human seminal plasma in western blot. These mAbs can specifically recognize PSA and may serve as a component of PSA diagnostic kit in various biological fluids