Objective To investigate the retardation effect of voltage gated potassium channel Kv1 5 on growth of gastric cancer cells SGC7901. Methods By using restriction enzymes of Hind Ⅲ and Kpn I, cDNA encoding Kv1 5 was reversely constructed into eukaryotic expression vector pcDNA3 1. SGC7901 cells were transfected with the recombinants using LipofectAMINE2000. Stable clones of transfectants were obtained after selection by G418 The growth of cells was monitored by cell growth curve and cell colony formation. The effect of Kv1 5 protein on cell cycle was examined by flow cytometry. Expression of Cyclin D1 protein was detected by Western blot. Results The antisense was found to effectively inhibit the expression of Kv1 5 protein in the transfectants. The growth and colony formation of transfectants were significantly reduced as compared with controls. The cell cycle review showed retardation of transfectants with Kv1 5 antisense in the G 1 phase. Expression of Cyclin D1 protein was decreased in Kv1 5 antisense transfectants. Conclusion The antisense of Kv1 5 has effect of retardation on growth of gastric cancer cells SGC7901

A review of cell traction forces (CTFs) measurement based on Biological MiCro Electromechanical Systems (BioMEMS) microposts matrix is presented.CTFs are exerted by cells and ansmitted to the underly-ing substrate through focal adhesions and close contacts.which is essential for cells movement.Cells probe the mechanicaI compliance of the exlracellular mabix (ECM) in part by locally deforming it with nanonewton-scale traction forces.Precision measurement of CTFs is significant for many researches such as call biology and tissue engineering and so on.Enabled by the advancement in BioMEMS technology,surface treated high aspeect ratio Polydimethyisiloxane(PDMS)micropos matrix devices,which serve as BioMEMS sensom for de-tecting cellular nanoforces and studying in vitro cell mechanics,have been developed.Closely spaced vartical microposts matrixes were designed to encourage cells to attach and spread across multiple microposts,and to bend the microposts like vertical cantilevers as the cells locomote on the surface.Using this dense and dis-crete matrix of microposts rather than a convanfional continuous substrate,CTFs can be directly measured and quantified by processing the microscopy images of the deformations of microposts.The resolution of the force was in tens of nN/μm scale.At first,the conventional CTFs measurement methods were concisely summa-rized.Then BioMEMS microposts matrix method was described in detail,including principle and fabfication process,Surface treatment and cell expedment results.Furthermore,high aspect ratio structure collapse prob-lem was investigated.

OBJECTIVETo generate phage-displayed anti-idiotypic antibody single chain variable fragments (anti-Id ScFv) to MG7 monoclonal antibody (McAb) directed against gastric carcinoma so as to lay a foundation for developing anti-Id ScFv vaccine of the cancer.

METHODSBalb/c mice were immunized i.p. with MG7 McAb conjugated with keyhole limpet hemocyanin (KLH), and mRNA was isolated from the spleens of the immunized mice. Heavy and light chain (VH and VL) genes of antibody were amplified separately and assembled into ScFv genes with a linker DNA by PCR. The ScFv genes were ligated into the phagemid vector pCANTAB5E and the ligated sample was transformed into competent E. coli TG1. The transformants were infected with M13K07 helper phage to yield recombinant phages displaying ScFv on the tips of M13 phage. After 4 rounds of panning with MG7, the MG7-positive clones were selected by ELISA from the enriched phages. The types of the anti-Id ScFv displayed on the selected phage clones were preliminarily identified by competition ELISA.

RESULTSThe VH, VL and ScFv DNAs were about 340 bp, 320 bp and 750 bp respectively. Twenty-four MG7-positive clones were selected from 60 enriched phage clones, among which 5 displayed beta or gamma type anti-Id ScFv.

CONCLUSIONThe anti-Id ScFv to MG7 McAb can be successfully selected by recombinant phage antibody technique, which paves a way for the study of prevention and cure of gastric carcinoma by using anti-Id ScFv.

Animals ; Antibodies, Anti-Idiotypic ; biosynthesis ; genetics ; Antibodies, Monoclonal ; genetics ; immunology ; Bacteriophages ; genetics ; Cloning, Molecular ; Immunoglobulin Fragments ; biosynthesis ; genetics ; Immunoglobulin Heavy Chains ; biosynthesis ; genetics ; Immunoglobulin Light Chains ; biosynthesis ; genetics ; Immunoglobulin Variable Region ; biosynthesis ; genetics ; Mice ; Mice, Inbred BALB C ; RNA, Messenger ; genetics ; Stomach Neoplasms ; immunology ; Vaccines, DNA ; genetics ; immunology