CD133 antigen, also known as prominin-1, is a glycoprotein that in humans is encoded by the PROM1 gene. It is a member of pentaspan transmembrane glycoproteins, which specifically localize to cellular protrusions. When embedded in the cell membrane, the membrane topology of prominin-1 is such that the N-terminus extends into the extracellular space and the C-terminus resides in the intracellular compartment. The protein consists of five transmembrane segments, with the first and second segments and the third and fourth segments connected by intracellular loops while the second and third as well as fourth and fifth transmembrane segments are connected by extracellular loops. CD133 is the most commonly used marker for isolation of cancer stem cell (CSC) population from different tumors, mainly from various gliomas and carcinomas. CD133+ melanoma cells are considered a subpopulation of CSC and play a critical role in recurrence. Moreover, CD133+ melanoma cells are immunogenic and can be used as an antimelanoma vaccination. In mice the vaccination with CD133+ melanoma cells mediated strong anti-tumor activity that resulted in the eradication of parental melanoma cells. In addition, it has also been shown that CD133+ melanoma cells preferentially express the RNA helicase DDX3X . As DDX3X also is an immunogenic protein, the same anti-melanoma vaccination strategy can be employed to give therapeutic antitumor immunity in mice.
Background References
1. Kim MY et al. Accumulation of low-dose BIX01294 promotes metastatic potential of U251 glioblastoma cells. Oncol Lett 13:1767-1774 (2017).
2. Xi G et al. Targeting CD133 improves chemotherapeutic efficacy of recurrent pediatric pilocytic astrocytoma following prolonged chemotherapy. Mol Cancer 16:21 (2017).
Western blot analysis of CD133 on different lysates with Mouse anti-CD133 antibody (HA601062) at 1/1,000 dilution.
Lane 1: Caco-2 cell lysate Lane 2: HT-29 cell lysate Lane 3: HeLa cell lysate (negative) Lane 4: NCCIT cell lysate
Lysates/proteins at 20 µg/Lane.
Predicted band size: 97 kDa Observed band size: 120 kDa
Exposure time: 10 seconds; ECL: K1801;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA601062) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Mouse IgG - HRP Secondary Antibody (HA1006) at 1/50,000 dilution was used for 1 hour at room temperature.
Western blot analysis of CD133 on different lysates with Mouse anti-CD133 antibody (HA601062) at 1/5,000 dilution.
Lane 1: HT-29 cell lysate Lane 2: SW480 cell lysate Lane 3: NCCIT cell lysate
Lysates/proteins at 10 µg/Lane.
Predicted band size: 97 kDa Observed band size: 120 kDa
Exposure time: 2 minutes;
10% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA601062) at 1/5,000 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Mouse IgG - HRP Secondary Antibody (HA1006) at 1:100,000 dilution was used for 1 hour at room temperature.
Immunohistochemical analysis of paraffin-embedded human colon carcinoma tissue with Mouse anti-CD133 antibody (HA601062) at 1/1,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA601062) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
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