Construction and identification of the recombinant hFcεRIα/RBL-2H3 cells

Plasmid. 2018 Jun:98:31-36. doi: 10.1016/j.plasmid.2018.09.004. Epub 2018 Sep 10.

Abstract

IgE/FcεRI signal pathway plays a crucial role in triggering allergic reactions, and there is no cross-recognition between IgE and FcεRI in human and rats. In order to obtain the hFcεRIα/ RBL-2H3 cell line, total RNA was extracted from U937 cells, and the human FcεRIα gene was obtained by RT-PCR technology. Then the amplified product was digested and inserted into the pIRES2-EGFP vector. After the plasmid was transfected into the RBL-2H3 cells using lipofectamine, and the RBL-2H3 cell lines of stable expression were screened by G418. The transfection efficiency reached 60.45% with optimizing transfection parameters. The last the expression of hFcεRIα was detected by RT-PCR, western blotting and fluorescent microscopy. The present results demonstrated that the pIRES2-EGFP-hFcεRIα vector was constructed and a stable cell line of hFcεRIα/ RBL-2H3 cells was established successfully. This cell line is promising tools for further research on the pathogenesis and drug development of allergic diseases.

Keywords: Allergic diseases; IgE; RBL-2H3 cells; hFcεRIα.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line, Tumor
  • Humans
  • Immunoglobulin E / genetics
  • Immunoglobulin E / metabolism*
  • Leukemia, Basophilic, Acute / genetics
  • Leukemia, Basophilic, Acute / metabolism*
  • Leukemia, Basophilic, Acute / pathology
  • Plasmids / genetics*
  • Rats
  • Receptors, IgE / genetics
  • Receptors, IgE / metabolism*
  • Transfection / methods*
  • Tumor Cells, Cultured

Substances

  • Receptors, IgE
  • Immunoglobulin E