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Plasmid DNA Preparation> | Vaccination Against Tumor Vascular CD99 125660452), optimized for protein expression in bacteria (Genscript) (333 bp), was inserted downstream the bacterial truncated thioredoxin (TRXtr) (26) sequence (192 bp) containing an N-terminal 4xGS-linker sequence and His-tag (6xhistidine) into a pET21a (+) vector (Novagen; EMD Chemicals, Gibbstown, NJ, USA).... Murine CD99 extracellular part protein sequence (aa27-137) (36): ASDDFNLGDALEDPNMKPTPKAPTPKKPSGGFDLED ALPGGGGGGAGEKPGNRPQPDPKPPRPHGDSGGISDSDL ADAAGQGGGGAGRRGSGDEGGHGGAGGAEPEGTPQ For construction of the pET21a-mCD99extracellular plasmid the extracellular part of murine CD99 was PCR amplified from the original pET21a-mCD99 vector (Genscript) using the following primers: Nde1-His-extracellmCD99fw 5′ -TAT-CAT-ATG-CAC-CAC-CAC-CAC-CAC-CAC- GCA-AGC-GAT-GAT-TTT-A-3′ Xho1-3x-stop-excellmCD99rev 5′ -TAT-CTC-GAG-CTA-TTA-TCA-ACC-CTG-CGG- GGT-ACC-TTC-CGG-TTC-3′ After purification and restriction with NdeI and XhoI the mCD99extracellular sequence was ligated into a pET21a vector. | Get A Quote |
CD99 (MIC2; single-chain type-1 glycoprotein) is a heavily O-glycosylated transmembrane protein (32 kDa) present on leukocytes and activated endothelium. Expression of CD99 on endothelium is important in lymphocyte diapedesis. CD99 is a diagnostic marker for Ewing's Sarcoma (EWS), as it is highly expressed by these tumors. It has been reported that CD99 can affect the migration, invasion and metastasis of tumor cells. Our results show that CD99 is also highly expressed in the tumor vasculature of most solid tumors. Furthermore, we found that in vitro CD99 expression in cultured endothelial cells is induced by starvation. Targeting of murine CD99 by a conjugate vaccine, which induced antibodies against CD99 in m... More