Limits...
Production and Characterization of Monoclonal Antibodies against Human Prostate Specific Antigen.

Bayat AA, Ghods R, Shabani M, Mahmoudi AR, Yeganeh O, Hassannia H, Sadeghitabar A, Balay-Goli L, Noutash-Haghighat F, Sarrafzadeh AR, Jeddi-Tehrani M - Avicenna J Med Biotechnol (2015 Jan-Mar)

Bottom Line: 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 kDa 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.

View Article: PubMed Central - PubMed

Affiliation: Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, Iran.

ABSTRACT

Background: 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.

Methods: 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 (PCa), Benign Prostatic Hyperplasia (BPH) and brain cancer tissues by Immunohistochemistry (IHC).

Results: Five anti-PSA mAbs (clones: 2G2-B2, 2F9-F4, 2D6-E8, IgG1/К) and clones (2C8-E9, 2G3-E2, IgG2a/К) 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 kDa in human seminal plasma in western blot.

Conclusion: These mAbs can specifically recognize PSA and may serve as a component of PSA diagnostic kit in various biological fluids.

No MeSH data available.


Related in: MedlinePlus

Titration of anti-PSA antibody in immunized mice sera by indirect ELISA. The results indicated that both mice were immunized with PSA. However, mouse 1 had higher titer of PSA-specific antibody and was selected for fusion.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4388886&req=5

Figure 0002: Titration of anti-PSA antibody in immunized mice sera by indirect ELISA. The results indicated that both mice were immunized with PSA. However, mouse 1 had higher titer of PSA-specific antibody and was selected for fusion.

Mentions: After immunization of Balb/c mice, the titers of anti-PSA antibodies in the mice sera were detected by indirect ELISA. Mouse 1 with higher titer of anti-PSA antibody was selected for hybridoma production (Figure 2). After cell fusion, supernatants of growing hybridoma cells were screened based on their reactivity with the purified PSA by ELISA. After four times of cloning, five hybridoma cells named 2G3-E2, 2F9-F4, 2G2-B2, 2D6-E8 and 2C8-E9 were produced. All five anti-PSA mAbs were produced in ascitic fluids and purified.


Production and Characterization of Monoclonal Antibodies against Human Prostate Specific Antigen.

Bayat AA, Ghods R, Shabani M, Mahmoudi AR, Yeganeh O, Hassannia H, Sadeghitabar A, Balay-Goli L, Noutash-Haghighat F, Sarrafzadeh AR, Jeddi-Tehrani M - Avicenna J Med Biotechnol (2015 Jan-Mar)

Titration of anti-PSA antibody in immunized mice sera by indirect ELISA. The results indicated that both mice were immunized with PSA. However, mouse 1 had higher titer of PSA-specific antibody and was selected for fusion.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4388886&req=5

Figure 0002: Titration of anti-PSA antibody in immunized mice sera by indirect ELISA. The results indicated that both mice were immunized with PSA. However, mouse 1 had higher titer of PSA-specific antibody and was selected for fusion.
Mentions: After immunization of Balb/c mice, the titers of anti-PSA antibodies in the mice sera were detected by indirect ELISA. Mouse 1 with higher titer of anti-PSA antibody was selected for hybridoma production (Figure 2). After cell fusion, supernatants of growing hybridoma cells were screened based on their reactivity with the purified PSA by ELISA. After four times of cloning, five hybridoma cells named 2G3-E2, 2F9-F4, 2G2-B2, 2D6-E8 and 2C8-E9 were produced. All five anti-PSA mAbs were produced in ascitic fluids and purified.

Bottom Line: 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 kDa 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.

View Article: PubMed Central - PubMed

Affiliation: Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, Iran.

ABSTRACT

Background: 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.

Methods: 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 (PCa), Benign Prostatic Hyperplasia (BPH) and brain cancer tissues by Immunohistochemistry (IHC).

Results: Five anti-PSA mAbs (clones: 2G2-B2, 2F9-F4, 2D6-E8, IgG1/К) and clones (2C8-E9, 2G3-E2, IgG2a/К) 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 kDa in human seminal plasma in western blot.

Conclusion: These mAbs can specifically recognize PSA and may serve as a component of PSA diagnostic kit in various biological fluids.

No MeSH data available.


Related in: MedlinePlus