title: Mass-tag enhanced immuno-laser desorption/ionization mass spectrometry for sensitive detection of intact protein antigens.
abstract: A new read-out method for antibody arrays using laser desorption/ionization-mass spectrometry (LDI-MS) is presented. Small, photocleavable reporter molecules with a defined mass called "mass-tags" are used for detection of immunocaptured proteins from human plasma. Using prostate specific antigen (PSA), a biomarker for prostate cancer, as a model antigen, a high sensitivity generic detection methodology based immunocapture with a primary antibody and with a biotin labeled secondary antibody coupled to mass-tagged avidin is demonstrated. As each secondary antibody can bind several avidin molecules, each having a large number of mass-tags, signal amplification can be achieved. The developed PSA sandwich mass-tag analysis method provided a limit of detection below 200 pg/mL (6 pM) for a 10 μL plasma sample, well below the clinically relevant cutoff value of 3-4 ng/mL. This brings the limit of detection (LOD) for detection of intact antigens with matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) down to levels comparable to capture by anti-peptide antibodies selected reaction monitoring (SISCAPA SRM) and enzyme linked immunosorbent assay (ELISA), as 6 pM corresponds to a maximal amount of 60 amol PSA captured on-spot. We propose the potential use of LDI (laser desorption/ionization) with mass-tag read-out implemented in a sandwich assay format for low abundant and/or early disease biomarker detection.
authors: †Meilahti Clinical Proteomics Core Facility, University of Helsinki, P.O. Box 63, FI-00014 University of Helsinki, Finland.|¶Institute of Biotechnology, University of Helsinki, P.O. Box 65, Helsinki FI-00014, Finland.|∥Unit of Systems Toxicology, Finnish Institute of Occupational Health, Topeliuksenkatu 41 b, FI-00250 Helsinki, Finland.
authors: Martina
authors: M
authors: Lorey
authors: ‡Department of Biomedical Engineering, Faculty of Engineering, Lund University, P.O. Box 118, SE-21100 Lund, Sweden.
authors: Belinda
authors: B
authors: Adler
authors: ‡Department of Biomedical Engineering, Faculty of Engineering, Lund University, P.O. Box 118, SE-21100 Lund, Sweden.
authors: Hong
authors: H
authors: Yan
authors: †Meilahti Clinical Proteomics Core Facility, University of Helsinki, P.O. Box 63, FI-00014 University of Helsinki, Finland.
authors: Rabah
authors: R
authors: Soliymani
authors: ‡Department of Biomedical Engineering, Faculty of Engineering, Lund University, P.O. Box 118, SE-21100 Lund, Sweden.
authors: Simon
authors: S
authors: Ekström
authors: §Division of Pharmaceutical Chemistry and Technology, Faculty of Pharmacy, University of Helsinki, P.O. Box 56, Viikinkaari 5 E, FI-00014 University of Helsinki, Finland.
authors: Jari
authors: J
authors: Yli-Kauhaluoma
authors: ‡Department of Biomedical Engineering, Faculty of Engineering, Lund University, P.O. Box 118, SE-21100 Lund, Sweden.|⊥Department of Biomedical Engineering, Dongguk University, 202 Science Building Phildong 3ga. Joong-Gu Seoul, Republic of Korea.
authors: Thomas
authors: T
authors: Laurell
authors: †Meilahti Clinical Proteomics Core Facility, University of Helsinki, P.O. Box 63, FI-00014 University of Helsinki, Finland.
authors: Marc
authors: M
authors: Baumann
mesh_terms: D055910:Antibodies, Immobilized / Q000737:chemistry; D004867:Equipment Design; D005260:Female; D006801:Humans; D007118:Immunoassay / Q000295:instrumentation*; D057230:Limit of Detection; D017430:Prostate-Specific Antigen / Q000032:analysis / Q000097:blood*; D040081:Protein Array Analysis / Q000295:instrumentation; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000295:instrumentation*
chemical_list: D055910:Antibodies, Immobilized; D017430:Prostate-Specific Antigen