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Prokaryotic expression of MLAA-34 and generation of a novel human ScFv against MLAA-34 by phage display technology
Yang Zhang1, Pengyu Zhang1, Aili He1
1Department of Hematology, The Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi 710004, P.R. China.
Abstract:
MLAA-34 is a newly identified monocytic leukemia-associated antigen that is overexpressed in acute monocytic leukemia specifically, thus providing a novel target for the therapy of acute monocytic leukemia. In this study, we first expressed MLAA-34 protein in Escherichia coli (E.coli) BL21 (DE3) cells and purified it by nickel ion affinity chromatography with high purity (>90%). Then, MLAA-34 was used as antigen for biopanning anti-MLAA-34 single chain antibody fragment (ScFv) from a fully human ScFv library, and a high affinity ScFv named MA1 was selected by phage-ELISA. Finally, after expression of MA1, we found that MA1 can specifically bind with U937 MLAA-34 positive cells, and the binding affinity of MA1 was at the nanomolar level. Furthermore, inhibition of U937 cell proliferation indicated that the novel antibody MA1 has the potential to be used as a therapeutic agent for acute monocytic leukemia.
Insights
Researchers identified MLAA-34 as a novel target for acute monocytic leukemia therapy. They developed a high-affinity antibody fragment (MA1) that specifically binds to cancer cells and inhibits their proliferation.
Area of Science:
- Oncology
- Immunology
- Biotechnology
Background:
- MLAA-34 is a novel monocytic leukemia-associated antigen.
- Overexpression of MLAA-34 is specific to acute monocytic leukemia.
- MLAA-34 represents a potential therapeutic target for acute monocytic leukemia.
Purpose of the Study:
- To express and purify MLAA-34 protein.
- To develop a high-affinity single-chain antibody fragment (ScFv) targeting MLAA-34.
- To evaluate the therapeutic potential of the developed antibody in acute monocytic leukemia.
Main Methods:
- MLAA-34 protein expression in E. coli and purification via nickel ion affinity chromatography.
- Biopanning a fully human ScFv library against MLAA-34.
- Selection of high-affinity ScFv (MA1) using phage-ELISA.
- Assessment of MA1 binding specificity and affinity to U937 cells.
Main Results:
- High-purity MLAA-34 protein (>90%) was successfully expressed and purified.
- A high-affinity ScFv, named MA1, was selected.
- MA1 demonstrated specific binding to MLAA-34 positive U937 cells at nanomolar affinity.
- MA1 inhibited U937 cell proliferation.
Conclusions:
- The novel antibody MA1 specifically targets MLAA-34 on acute monocytic leukemia cells.
- MA1 exhibits high binding affinity and inhibits cancer cell proliferation.
- MA1 holds significant potential as a therapeutic agent for acute monocytic leukemia.