Abstract
We have produced and characterized six urine monoclonal antibodies to human apolipoprotein A-I named A-I-9, A-I-12, A-I-15, A-I-16, A-I-19, and A-I-57. All monoclonal antibodies were specific for apolipoprotein A-I and bound between 55% and 100% of 125I-labeled high density lipoproteins (HDL) in a fluid phase radioimmunoassay. All antibodies possessed a higher affinity to apoA-I in HDL than to free, delipidated apoA-I. Two of them, particularly A-I-12 and A-I-15, which were directed to the same or very close epitopes on the molecule, recognized very poorly the delipidated protein. Binding of apoA-I to phospholipid restored the immunoreactivity of the monoclonal antibodies ot the protein suggesting that lipids play and important role in determining the immunochemical structure of apoA-I. Using CNBr fragments and synthetic peptides, the epitopes for the antibodies were mapped as follows: A-I-19, CNBr fragment 1; A-I-12 and 15, CNBr fragment 2; A-I-9 and A-I-16, CNBr fragment 3; A-I-57, CNBr fragment 4. Antibody A-I-57 failed to recognize a mutant form of apoA-I, A-I(Milano) (Arg173 → Cys) by immunoblotting and by competitive radioimmunoassay demonstrating that substitution of a single amino acid in human apoA-I may cause the loss of an antigenic determinant.
| Original language | English |
|---|---|
| Pages (from-to) | 375-384 |
| Number of pages | 10 |
| Journal | Journal of Lipid Research |
| Volume | 31 |
| Issue number | 3 |
| Publication status | Published - 1990 |
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Keywords
- apolipoprotein A-I(Milano)
- conformational changes
- delipated apolipoprotein A-I
- high density lipoprotein
ASJC Scopus subject areas
- Endocrinology
Cite this
Immunochemical characterization of six monoclonal antibodies to human apolipoprotein A-I : Epitope mapping and expression. / Marcovina, S.; Fantappie, S.; Zoppo, A.; Franceschini, G.; Catapano, A. L.
In: Journal of Lipid Research, Vol. 31, No. 3, 1990, p. 375-384.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Immunochemical characterization of six monoclonal antibodies to human apolipoprotein A-I
T2 - Epitope mapping and expression
AU - Marcovina, S.
AU - Fantappie, S.
AU - Zoppo, A.
AU - Franceschini, G.
AU - Catapano, A. L.
PY - 1990
Y1 - 1990
N2 - We have produced and characterized six urine monoclonal antibodies to human apolipoprotein A-I named A-I-9, A-I-12, A-I-15, A-I-16, A-I-19, and A-I-57. All monoclonal antibodies were specific for apolipoprotein A-I and bound between 55% and 100% of 125I-labeled high density lipoproteins (HDL) in a fluid phase radioimmunoassay. All antibodies possessed a higher affinity to apoA-I in HDL than to free, delipidated apoA-I. Two of them, particularly A-I-12 and A-I-15, which were directed to the same or very close epitopes on the molecule, recognized very poorly the delipidated protein. Binding of apoA-I to phospholipid restored the immunoreactivity of the monoclonal antibodies ot the protein suggesting that lipids play and important role in determining the immunochemical structure of apoA-I. Using CNBr fragments and synthetic peptides, the epitopes for the antibodies were mapped as follows: A-I-19, CNBr fragment 1; A-I-12 and 15, CNBr fragment 2; A-I-9 and A-I-16, CNBr fragment 3; A-I-57, CNBr fragment 4. Antibody A-I-57 failed to recognize a mutant form of apoA-I, A-I(Milano) (Arg173 → Cys) by immunoblotting and by competitive radioimmunoassay demonstrating that substitution of a single amino acid in human apoA-I may cause the loss of an antigenic determinant.
AB - We have produced and characterized six urine monoclonal antibodies to human apolipoprotein A-I named A-I-9, A-I-12, A-I-15, A-I-16, A-I-19, and A-I-57. All monoclonal antibodies were specific for apolipoprotein A-I and bound between 55% and 100% of 125I-labeled high density lipoproteins (HDL) in a fluid phase radioimmunoassay. All antibodies possessed a higher affinity to apoA-I in HDL than to free, delipidated apoA-I. Two of them, particularly A-I-12 and A-I-15, which were directed to the same or very close epitopes on the molecule, recognized very poorly the delipidated protein. Binding of apoA-I to phospholipid restored the immunoreactivity of the monoclonal antibodies ot the protein suggesting that lipids play and important role in determining the immunochemical structure of apoA-I. Using CNBr fragments and synthetic peptides, the epitopes for the antibodies were mapped as follows: A-I-19, CNBr fragment 1; A-I-12 and 15, CNBr fragment 2; A-I-9 and A-I-16, CNBr fragment 3; A-I-57, CNBr fragment 4. Antibody A-I-57 failed to recognize a mutant form of apoA-I, A-I(Milano) (Arg173 → Cys) by immunoblotting and by competitive radioimmunoassay demonstrating that substitution of a single amino acid in human apoA-I may cause the loss of an antigenic determinant.
KW - apolipoprotein A-I(Milano)
KW - conformational changes
KW - delipated apolipoprotein A-I
KW - high density lipoprotein
UR - http://www.scopus.com/inward/record.url?scp=0025220767&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0025220767&partnerID=8YFLogxK
M3 - Article
VL - 31
SP - 375
EP - 384
JO - Journal of Lipid Research
JF - Journal of Lipid Research
SN - 0022-2275
IS - 3
ER -