Preparation of anti-PEG antibody expressing cell and application thereof
AssigneeKaohsiung Medical University
InventorsTian-Lu Cheng, Steven R. Roffler, Kuo-Hsiang Chuang, Ssu-Jung Lu
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Abstract
The present invention provides a method for preparing an isolated eukaryotic cell which presents an anti-polyethylene glycol (PEG) antibody on a cell membrane. The present invention also provides a method for a quantitative analysis of a polyethylene glycol (PEG) by said anti-PEG antibody expressing cell. The cell-based quantitative analysis of the present prevention could sensitively quantify free PEG and PEG-modified macromolecules (proteins, nanoparticles and liposomes) as sensitive as nano-gram level.
Claims15
A method for preparing an isolated eukaryotic cell which presents an anti-polyethylene glycol (PEG) antibody on a cell membrane, comprising:
- (a) providing a vector comprising a nucleic acid sequence for encoding the anti-PEG antibody which is fused thereto a nucleic acid sequence for encoding a transmembrane protein, wherein the nucleic acid sequence for encoding the anti-PEG antibody comprises at least a nucleic acid sequence of a heavy chain variable region (VH) and a nucleic acid sequence of a light chain ion (VL) and the nucleic acid sequence for encoding the anti-PEG antibody is selected from the group consisting of (i) a 3-3 antibody consisting of the VH encoded by SEQ ID NO:
- 5 and the VL encoded by SEQ ID NO:
- 6;
- (ii) a 6-3 antibody consisting of the VH encoded by SEQ ID NO:
- 7 and the VL encoded by SEQ ID NO:
- 8;
- (iii) a 15-2 antibody consisting of the VH encoded by SEQ ID NO:
- 9 and the VL encoded by SEQ ID NO:
- 10;
- and combinations thereof;
- (b) transfecting the vector into an eukaryotic cell, thereby enabling the eukaryotic cell to express the anti-PEG antibody and the transmembrane protein;
- and
- (c) isolating the eukaryotic cell which presents the anti-PEG antibody on the cell membrane, wherein the anti-PEG antibody presents in a consistent direction on the cell membrane and the anti-PEG antibody has an extracellularly presented antigen binding domain.
A method for a quantitative analysis of a polyethylene glycol (PEG), comprising:
- (a) providing an isolated eukaryotic cell which presents an anti-PEG antibody on a cell membrane, wherein the anti-PEG antibody presents in a consistent direction on the cell membrane and has an extracellularly presented antigen binding domain, wherein the anti-PEG antibody comprises at least a heavy chain variable region (VH) and a light chain variable region (VL), and the isolated eukaryotic cell comprises a vector comprising a nucleic acid sequence for encoding the anti-PEG antibody which is fused thereto a nucleic acid sequence for encoding a transmembrane protein, and the nucleic acid sequence for encoding the anti-PEG antibody is selected from the group consisting of (i) a 3-3 antibody consisting of the VH encoded by SEQ ID NO:
- 5 and the VL encoded by SEQ ID NO:
- 6:
- (ii) a 6-3 antibody consisting of the VH encoded by SEQ ID NO:
- 7 and the VL encoded by SEQ ID NO:
- 8;
- (iii) a 15-2 antibody consisting of the VH encoded by SEQ ID NO:
- 9 and the VL encoded by SEQ ID NO:
- 10;
- and combinations thereof;
- (b) contacting the anti-PEG antibody on the cell membrane of the eukaryotic cell with a specimen suspected containing a PEG to form a specific binding complex between the anti-PEG antibody and the PEG in the specimen;
- and
- (c) measuring a concentration of the PEG in the specimen by detecting an amount of the binding complex between the PEG in the specimen and the anti-PEG antibody on the cell membrane of the eukaryotic cell.
Dependencies are inferred from the claim text.