Region-selective labeling of antibodies as determined by electrospray ionization-mass spectrometry (ESI-MS).
The electrospray ionization-mass spectrometry (ESI-MS) analysis of three sets of monoclonal antibody-acridinium-9-carboxamide conjugates is described. The conjugates (nine total) were enzymatically digested using papain and the resulting fragments [Fc heavy chain, Fab, or F(ab')(2)] were analyzed using liquid chromatography/ESI-MS. The average number of labels per fragment were calculated using Sigma nx%, where n is the number of acridinium molecules covalently bound to the fragment and x% is the percent relative area of the corresponding peaks in the mass spectrum. When these values were normalized against the molecular weight of their respective region, antibody-dependent labeling patterns were observed. For antibodies T (anti-L-T(4)) and F (anti-FITC), there was a preference for conjugation of the Fab region over the Fc region. For antibody B (anti-biotin), the trend was reversed.
Adamczyk
M
M
Department of Chemistry (9NM), Abbott Diagnostics Division, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, Illinois 60064-6016, USA. maciej.adamczyk@abbott.com
Gebler
J
J
Shreder
K
K
Wu
J
J
D000906:Antibodies / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods*
D000906:Antibodies; D007140:Immunoglobulin Fab Fragments; D007141:Immunoglobulin Fc Fragments