Related Experiment Video
Updated: Jul 10, 2026
Profiling of Estrogen-regulated MicroRNAs in Breast Cancer Cells
Published on: February 21, 2014
A quantitative analysis of N-myc downstream regulated gene 2 (NDRG 2) in human tissues and cell lysates by
Mi-Young Park1, Seung-Chul Choi, Hayang Sin Lee
1Stem Cell Research Center, KRIBB, Daejeon 305-806, South Korea.
Background:
N-myc downstream regulated gene 2 (NDRG2) belongs to the NDRG family, which is comprised of 4 members, NDRG1-4. Recently, NDRG2 was reported as a new candidate for a tumor suppressor gene. We developed a reverse-phase protein microarray assay to access NDRG2 levels in human tissue specimens and cell lines.
Methods:
We synthesized recombinant NDRG2 protein and produced monoclonal antibodies (mAb) to the NDRG2 protein. We selected 2 hybridomas producing mAb that specifically recognize the NDRG2 protein. To determine the NDRG2 concentration, the samples of serially-diluted NDRG2 protein, cell lysate, or tissue lysate were spotted onto a nitrocellulose membrane-coated slide glass and allowed to react with the mAb to the NDRG2 protein. The reaction was followed by additional incubation with biotin-linked anti-mouse IgG and horseradish peroxidase (HRP)-conjugated streptavidin, subsequently. The addition of dimethylaminobenzidine induced color development, which was measured using the GenePix program. We determined the NDRG2 concentration in various tissue specimens and cell lines using the new protein microarray technique.
Results:
The dose-response relationship between NDRG2 and color intensity showed linearity in a range 0-10 ng/ml and a sensitivity of 50 pg/ml. The NDRG2 concentrations in the liver tissue lysates of patients with hepatocellular carcinoma (52.0+21.5 ng/mg) were significantly diminished as compared with those in the normal liver tissues (549.6+94.6 ng/mg). The results of the assay showed good agreement with those of Western blot analysis.
Conclusions:
The protein microarray is a highly sensitive and accurate method, and can adopted to assess specific proteins in human tissues or cell lines, particularly in the field of cancer and pathological research.
Insights
A new protein microarray assay accurately measures N-myc downstream regulated gene 2 (NDRG2) levels. NDRG2 was significantly lower in liver cancer tissues, supporting its role as a tumor suppressor.
Area of Science:
- Molecular Biology
- Biochemistry
- Oncology
Background:
- N-myc downstream regulated gene 2 (NDRG2) is a potential tumor suppressor.
- The NDRG family includes NDRG1-4.
- NDRG2 levels in human tissues require accurate assessment.
Purpose of the Study:
- To develop a reverse-phase protein microarray assay for quantifying NDRG2.
- To evaluate NDRG2 expression in human tissue specimens and cell lines.
Main Methods:
- Synthesized recombinant NDRG2 protein and developed specific monoclonal antibodies (mAbs).
- Utilized a protein microarray technique involving mAb incubation, biotin-linked secondary antibodies, and HRP-conjugated streptavidin.
- Measured color development using GenePix software to determine NDRG2 concentrations.
Main Results:
- The assay demonstrated linearity (0-10 ng/ml) and high sensitivity (50 pg/ml).
- NDRG2 concentrations were significantly reduced in hepatocellular carcinoma liver tissues compared to normal liver tissues.
- Assay results correlated well with Western blot analysis.
Conclusions:
- The developed protein microarray is a sensitive and accurate tool for assessing NDRG2.
- This method is applicable for protein analysis in human tissues and cell lines.
- The findings support NDRG2's role in cancer and pathological research.
