Author: DOKI, Tomoyoshi; TAKANO, Tomomi; HOHDATSU, Tsutomu
Title: Development of a mouse-feline chimeric antibody against feline tumor necrosis factor-alpha Document date: 2016_6_4
ID: y0x0quha_9
Snippet: Construction and expression of chimeric mAb 2-4: The C H and C L genes and V H and V L fragments inserted in the pCR-bluntII-TOPO vectors were connected using a Bam HI linker to prepare chimeric H and L chains (Fig. S1 ), respectively. The chimeric H chain fragment was inserted into the EcoRI site of the pCDNA3.1 (+)/Neo expression vector. The chimeric L chain fragment was inserted into the HindIII/EcoRV site of the pCDNA3.1 (+)/Hygro expression .....
Document: Construction and expression of chimeric mAb 2-4: The C H and C L genes and V H and V L fragments inserted in the pCR-bluntII-TOPO vectors were connected using a Bam HI linker to prepare chimeric H and L chains (Fig. S1 ), respectively. The chimeric H chain fragment was inserted into the EcoRI site of the pCDNA3.1 (+)/Neo expression vector. The chimeric L chain fragment was inserted into the HindIII/EcoRV site of the pCDNA3.1 (+)/Hygro expression vector. FO cells were co-transfected with the H-and L-chain expression vectors with Lipofectamine 2000 (Life Technologies). The transfected FO cells were cultured in medium containing G418 (Roche Diagnostics, Basel, Switzerland) and hygromycyn B (Roche Diagnostics), to acquire a stably expressing cell line (FOCM24). FOCM24 cells were cloned twice employing the limiting dilution method.
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