Preparation of anti-recombinant soluble tissue factor monoclonal antibody by trace injecting antigen into spleen

Purpose: To prepare the anti-recombinant soluble tissue factor monoclonal antibody to develop an ELISA method which can be used to detect the tissue factor (TF); and to give a reference for going deep into the research and clinical application of TF. Methods: We used TF to immunize Balb/c mice by injecting TF into the spleen directly. Then the spleen cells were separated and fused with mouse myeloma cells (SP2/0) to obtain the hybridoma. The hybridoma was cloned and got 3 clones named as 1-C3, 2-G9, and 3-H9 which could stably secrete TF, and then they were embedded in mice bodies to induce ascites. TF was purified by protein A affinity chromatograph column and labeled with HRP. At last, normal humans' plasma TF levels were determined by ELISA developed from the antibody. Results: The purified TF identified as IgG1 was a single band about M, 150 000 by SDS-PAGE analysis and Western blot showed that it could specifically recognize TF. Normal humans plasma TF was about (130 ± 80) pg/mL as measured by ELISA. Conclusions: We have prepared monoclonal antibody against TF and developed ELISA assays for TF successfully. The research will be useful for TF purification, measurements and clinical study.