Preferential labeling of alpha-amino N-terminal groups in peptides by biotin: application to the detection of specific anti-peptide antibodies by enzyme immunoassays.
Experimental conditions (pH 6.5, 24 h reaction, peptide:biotin ratio 1:5) were defined for preferential incorporation of the biotin molecule in the N-terminal alpha-amino group of peptides. This strategy could be helpful in numerous applications when an entire peptide chain must remain accessible for antibody or receptor binding. We illustrate this advantage in a solid-phase enzyme immunoassay designed to detect antibodies specific for bovine beta-lactoglobulin present in rabbit or human sera. This test involves synthetic peptides biotinylated in different positions and immobilized on a solid phase. The use of biotin/streptavidin interactions permitted more efficient detection of specific anti-peptide antibodies than solid phases prepared using conventional passive-adsorption techniques. The highest levels of antibody binding were measured when biotinylation occurred at the N-terminal extremity of immobilized peptides.
Sélo
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Laboratoire Associé INRA-CEA d'Immuno Allergie Alimentaire, SPI Bât 136, Gif sur Yvette, France.
Négroni
L
L
Créminon
C
C
Grassi
J
J
Wal
J M
JM
D000595:Amino Acid Sequence; D000818:Animals; D001710:Biotin / Q000737:chemistry*; D002417:Cattle; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007124:Immunoenzyme Techniques; D007782:Lactoglobulins / Q000276:immunology; D008894:Milk Proteins / Q000276:immunology; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*; D011817:Rabbits
D007782:Lactoglobulins; D008894:Milk Proteins; D010455:Peptides; D001710:Biotin