Past Issue

Volume 12, Supplement 1,Winter 2011 (Presented at The 1st International Student Congress) Pages: 80-80

P-90: Production Polyclonal Antibody Against Surface Markers of Human Adipose-Tissue Mesenchymal Stem Cells Derived from Breast Cancer Patient

Objective: Mesenchymal Stem Cells (MSCs) have enormous potential for the regeneration and differentiation to cartilage, bone and other tissues derived from embryonic layers. Despite extensive research, there is still no specific marker that reliably identifies hMSCs from other stem cells. Therefore, interest has been directed toward recognition of specific cell surface markers of MSCs. The objective of this study was to produce polyclonal antibodies against MSCs for recognition of specific surface markers of MSCs. Materials and Methods: MSCs isolated from breast fat tissue from a breast cancer patient and normal breast. These cells were cultured and expanded in MBM,DMEM-LG containing 10% FBS,10% FFP.Several features of the MSCs were investigated, including morphology characteristics, microscopic features and expression various antigens with flowcytometry method. Polyclonal antibody produced through subcutaneous injection of MSCs into rabbits. Rabbits’ sera were screened for antibodies against surface markers of MSCs, using ELISA method. Crossreactivity of antibodies were tested with PBMC, bone marrow derived MSCs and several cell lines. Finally molecular size of determinant which were specifically recognized by antibodies were investigated, using SDS-PAGE and immunoblotting methods. Results: Our MSCs were positive for CD105, CD166, and CD44 surface antigens (known markers of MSCs), but were negative for CD45,CD14,CD34 surface antigens. Purified anti human MSCs polyclonal antibodies exhibited high reactivity with MSCs, but these had low reactivity with PBMC and other cell lines. Polyclonal antibodies recognized four specific bands with MW of 19,150,180,210 KD in immunoblotting of membrane proteins from human adipose tissue derived MSCs. Conclusion: We recognized several protein bands that were predominantly reacted with MSCs by probing membrane fractions of MSCs with antirabbit antibodies. Characterization of these proteins is underway