[Preparation and assessment of a monoclonal antibody against mouse NLRP3].
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi
; 40(4): 354-361, 2024 Apr.
Article
in Zh
| MEDLINE
| ID: mdl-38710518
ABSTRACT
Objective To prepare a monoclonal antibody (mAb) against mouse NOD-like receptor family pyrin domain-containing 3 (NLRP3) and assess its specificity. Methods A gene fragment encoding mouse NLRP3 exon3 (Ms-N3) was inserted into the vector p36-G3-throhFc to construct a recombinant plasmid named Ms-N3-throhFc. This plasmid was then transfected into HEK293F cells for eukaryotic expression. NLRP3-/- mice were immunized with Ms-N3 protein purified using a protein A chromatography column, and splenocytes from the immunized mice were fused with SP2/0 myeloma cells to generate hybridoma cells. Specific mAbs against murine NLRP3 from hybridoma cells were screened using ELISA and immunofluorescence assay(IFA). Results The Ms-N3-throhFc recombinant plasmid was successfully constructed and exhibited stable expression in HEK293F cells. Twelve hybridoma cell lines were initially screened using ELISA. IFA revealed that the mAb secreted by the 9-B8-3-2-C5 cell line specifically recognized the native form of mouse NLRP3 protein. The heavy and light chain subtypes of this mAb were identified as IgM and κ, respectively. Conclusion A monoclonal antibody against mouse NLRP3 has been successfully prepared.
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Collection:
01-internacional
Database:
MEDLINE
Main subject:
NLR Family, Pyrin Domain-Containing 3 Protein
/
Antibodies, Monoclonal
Limits:
Animals
/
Humans
Language:
Zh
Journal:
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi
Journal subject:
ALERGIA E IMUNOLOGIA
Year:
2024
Document type:
Article
Affiliation country:
China