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Functional expression, characterization and application of the human S100A4 protein.

Wang D, Zhang J, Liu Z, Chen Y, Xu C, Zhang Z, Liu X, Wu L, Zhou X, Meng X, Li H, Liu H, Jiang Z, Wang T - Mol Med Rep (2014)

Bottom Line: Four hybridoma cell lines, which secreted mAbs specifically against human S100A4 protein, were obtained.One of the four mAbs, namely 2A12D10B2, recognized human S100A4 as indicated by immunohistochemical staining of human gastric carcinoma specimens and recombinant S100A4 was functionally expressed in E. coli.The mAbs of recombinant S100A4 were suitable for detecting S100A4 expression in human tissues and for investigating the subsequent clinical applications of the protein.

View Article: PubMed Central - PubMed

Affiliation: Performance Medicine Laboratory, Institute of Health and Environmental Medicine, Tianjin 300050, P.R. China.

ABSTRACT
Preparations utilizing monoclonal antibodies against S100A4 provide useful tools for functional studies to investigate the clinical applications of the human S100A4 protein. In the present study, human S100A4 protein was expressed in Escherichia coli (E. coli) BL21 (DE3), successfully purified by diethylaminoethyl cellulose anion-exchange chromatography and identified by western blot analysis. Soluble S100A4 bioactivity was confirmed by Transwell migration and invasion assays in the human HeLa cell line. Monoclonal antibodies (mAbs) were generated utilizing the standard hybridoma method and were validated by enzyme-linked immunosorbent assay and western blot analysis. The antibody was then used to examine human gastric carcinoma specimens by immunohistochemistry. Recombinant S100A4 was functionally expressed in E. coli and promoted the migration and invasion of HeLa cells. Four hybridoma cell lines, which secreted mAbs specifically against human S100A4 protein, were obtained. One of the four mAbs, namely 2A12D10B2, recognized human S100A4 as indicated by immunohistochemical staining of human gastric carcinoma specimens and recombinant S100A4 was functionally expressed in E. coli. The mAbs of recombinant S100A4 were suitable for detecting S100A4 expression in human tissues and for investigating the subsequent clinical applications of the protein.

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Related in: MedlinePlus

Plotted standard curve based on SPR data. SPR, surface plasmon resonance; Req, response of equilibrium.
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f4-mmr-11-01-0175: Plotted standard curve based on SPR data. SPR, surface plasmon resonance; Req, response of equilibrium.

Mentions: To calculate the association rate constant, 2A12D10B2 mAb was appropriately diluted in PBS and analyzed by SPR at several concentrations (Fig. 4). The equilibrium dissociation constant (KD) for the 2A12D10B2 clone was also calculated. KDs were determined independently by Kinetic Evaluation 5.0 software. The KD of 2A12D10B2 was ~4.72×10−8 mol/l.


Functional expression, characterization and application of the human S100A4 protein.

Wang D, Zhang J, Liu Z, Chen Y, Xu C, Zhang Z, Liu X, Wu L, Zhou X, Meng X, Li H, Liu H, Jiang Z, Wang T - Mol Med Rep (2014)

Plotted standard curve based on SPR data. SPR, surface plasmon resonance; Req, response of equilibrium.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4237081&req=5

f4-mmr-11-01-0175: Plotted standard curve based on SPR data. SPR, surface plasmon resonance; Req, response of equilibrium.
Mentions: To calculate the association rate constant, 2A12D10B2 mAb was appropriately diluted in PBS and analyzed by SPR at several concentrations (Fig. 4). The equilibrium dissociation constant (KD) for the 2A12D10B2 clone was also calculated. KDs were determined independently by Kinetic Evaluation 5.0 software. The KD of 2A12D10B2 was ~4.72×10−8 mol/l.

Bottom Line: Four hybridoma cell lines, which secreted mAbs specifically against human S100A4 protein, were obtained.One of the four mAbs, namely 2A12D10B2, recognized human S100A4 as indicated by immunohistochemical staining of human gastric carcinoma specimens and recombinant S100A4 was functionally expressed in E. coli.The mAbs of recombinant S100A4 were suitable for detecting S100A4 expression in human tissues and for investigating the subsequent clinical applications of the protein.

View Article: PubMed Central - PubMed

Affiliation: Performance Medicine Laboratory, Institute of Health and Environmental Medicine, Tianjin 300050, P.R. China.

ABSTRACT
Preparations utilizing monoclonal antibodies against S100A4 provide useful tools for functional studies to investigate the clinical applications of the human S100A4 protein. In the present study, human S100A4 protein was expressed in Escherichia coli (E. coli) BL21 (DE3), successfully purified by diethylaminoethyl cellulose anion-exchange chromatography and identified by western blot analysis. Soluble S100A4 bioactivity was confirmed by Transwell migration and invasion assays in the human HeLa cell line. Monoclonal antibodies (mAbs) were generated utilizing the standard hybridoma method and were validated by enzyme-linked immunosorbent assay and western blot analysis. The antibody was then used to examine human gastric carcinoma specimens by immunohistochemistry. Recombinant S100A4 was functionally expressed in E. coli and promoted the migration and invasion of HeLa cells. Four hybridoma cell lines, which secreted mAbs specifically against human S100A4 protein, were obtained. One of the four mAbs, namely 2A12D10B2, recognized human S100A4 as indicated by immunohistochemical staining of human gastric carcinoma specimens and recombinant S100A4 was functionally expressed in E. coli. The mAbs of recombinant S100A4 were suitable for detecting S100A4 expression in human tissues and for investigating the subsequent clinical applications of the protein.

Show MeSH
Related in: MedlinePlus