Bioinformatics analysis of mouse adiponectin receptor-1 and its antibody preparation / 生理学报
Acta Physiologica Sinica
; (6): 156-160, 2008.
Article
ي Zh
| WPRIM
| ID: wpr-316747
المكتبة المسؤولة:
WPRO
ABSTRACT
To establish a method for preparation of anti-mouse adiponectin receptor-1 (AdipoR-1) polyclonal antibody, the polypeptide antigen corresponding to AdipoR-1 was designed by bioinformatics analysis. The possible physicochemical property and trans-membrane structure were predicted by ExPASy and TMHMM, respectively. The antigen epitopes of mouse AdipoR-1 and its immunogenicity were analyzed by Antigenic Prediction and AntigenProfiler, respectively. According to the similarity analysis between AdipoR-1 and AdipoR-2 by Clustal W, a 16-amino acid polypeptide was designed as the antigen corresponding to AdipoR-1. To ensure the specificity of the polypeptide antigen, similarity search was run in the protein databases such as SWISS-PROT, PDB and Prosite databases. The polypeptide synthesized by solid-phase synthesis was used as immunogen to immunize rats to obtain anti-mouse AdipoR-1 polyclonal antibodies, the specificity and titer of which was identified by Western blot and indirect ELISA. The antibodies were applied to detect the AdipoR-1 expression in the muscle tissue in normal and cholesterolemic mice. The results from bioinformatics analysis showed that the similarity of amino acid sequences between AdipoR-1 and AdipoR-2 in mouse was 66%, and the designed polypeptide antigen corresponding to AdipoR-1 exhibited excellent immunogenicity (score=3.1). Using the polypeptide as antigen for immunization, anti-mouse AdipoR-1 polyclonal antibodies with high titer and good specificity were obtained. The results of Western blot demonstrated that there was no statistical difference in AdipoR-1 expression in muscle tissue between normal (1.80±0.06) and cholesterolemic mice (1.71±0.11). These results suggest that the antigen epitopes of mouse AdipoR-1 are well predicted by bioinformatics analysis, and successful preparation of the specific anti-AdipoR-1 polyclonal antibodies provides a useful tool for identification and further functional study of AdipoR-1.
النص الكامل:
1
الفهرس:
WPRIM
الموضوع الرئيسي:
Peptides
/
Enzyme-Linked Immunosorbent Assay
/
Chemistry
/
Blotting, Western
/
Amino Acid Sequence
/
Computational Biology
/
Allergy and Immunology
/
Receptors, Adiponectin
/
Antibodies
/
Antibody Specificity
نوع الدراسة:
Prognostic_studies
المحددات:
Animals
اللغة:
Zh
مجلة:
Acta Physiologica Sinica
السنة:
2008
نوع:
Article