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. 2012 Jun;31(3):209-13.
doi: 10.1089/hyb.2011.0115.

A monoclonal antibody against the catalytic domain of PTP1B

Fanwei Zeng  1 , Erru Ni, Shengyu Wang, Sha Lv, Chang Cao, Haipeng Zhang, Haiyan Lv, Xiaoli Chen, Jianghua Yan, Fanghong Luo
Affiliations

A monoclonal antibody against the catalytic domain of PTP1B

Fanwei Zeng et al. Hybridoma (Larchmt). 2012 Jun.

Abstract

Protein tyrosine phosphatase 1B (PTP1B), a member of the protein tyrosine phosphatase (PTP) family, plays a crucial role in metabolic signaling, with insulin and leptin signaling being well studied. New evidence indicates that PTP1B is also involved in cancer. In the present study, we report on the establishment of a monoclonal antibody specific for catalytic domain of PTP1B (PTP1Bc) generated through the hybridoma method. The monoclonal antibody is measured to have a titer of 4.1×10(6) against PTP1Bc in indirect ELISA. Western blot and immunofluorescent analyses indicated that this antibody can specifically combine native PTP1B in MDA-MB-231 and MDA-MB-453 cells. This monoclonal antibody against PTP1Bc can help enhance the understanding of PTP1B-related physiological and pathological mechanisms and may act as a therapeutic agent for diabetes, obesity, and cancer in the future.

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Figures

FIG. 1.
FIG. 1.
SDS-PAGE analysis for the expression and purification of PTP1Bc in DE3. M, protein marker; lane 1, DE3 culture without IPTG induction; lane 2, DE3 culture after 0.3mM IPTG induction; lane 3, supernatant of DE3 culture with 0.3 mM IPTG induction; lane 4, precipitate of DE3 culture with 0.3 mM IPTG induction; lanes 5–7, purified PTP1Bc in different concentrations.
FIG. 2.
FIG. 2.
SDS-PAGE analysis of MAb ascites and Garbu MAb. M, protein marker; lane 1, MAb ascites; lane 2, Garbu MAb.
FIG. 3.
FIG. 3.
Titration analysis of MAb ascites and Garbu MAb. 1, MAb ascites; 2, Garbu MAb; 3, mouse IgG1 as control. The coating concentration of PTP1Bc was 10 μg/mL. Each point represents the OD values with the mean±standard deviation from six determinations (n=6).
FIG. 4.
FIG. 4.
Western blot analysis with Garbu MAb. M, protein marker; lane 1, extracts of MDA-MB-231 cells; lane 2, extracts of MDA-MB-453 cells; lane 3, purified PTP1Bc fusion protein.
FIG. 5.
FIG. 5.
Immunofluorescent analysis of MDA-MB-231 and MDA-MB-453 cells with Garbu MAb.
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