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Free and complexed prostate-specific antigen (PSA) : in vitro stability, epitope map, and development of immunofluorometric assays for specific and sensitive detection of free PSA and PSA-alpha 1-antichymotrypsin complex

Pettersson, K ; Piironen, T ; Seppälä, M ; Liukkonen, L ; Christensson, A LU ; Matikainen, M T ; Suonpää, M ; Lövgren, T and Lilja, H LU (1995) In Clinical Chemistry 41(10). p.8-1480
Abstract

Generation of 15 monoclonal antibodies (MAbs) allowed construction of epitope maps and specific two-site immunofluorometric assays for free prostate-specific antigen (PSA) and PSA complexed with alpha 1-antichymotrypsin (ACT). Close correlation of PSA concentrations obtained with the use of different assays of free PSA suggested extensive similarity in immunodetection of free PSA in serum. Assays of the PSA-ACT complex overestimated the concentration of PSA-ACT in serum because of nonspecific adsorbance of ACT or cathepsin G-complexed ACT to the solid phase. This interference was substantially decreased in the presence of heparin. In studying the stability of purified PSA and PSA-ACT complexes formed in vitro, we found that the free PSA... (More)

Generation of 15 monoclonal antibodies (MAbs) allowed construction of epitope maps and specific two-site immunofluorometric assays for free prostate-specific antigen (PSA) and PSA complexed with alpha 1-antichymotrypsin (ACT). Close correlation of PSA concentrations obtained with the use of different assays of free PSA suggested extensive similarity in immunodetection of free PSA in serum. Assays of the PSA-ACT complex overestimated the concentration of PSA-ACT in serum because of nonspecific adsorbance of ACT or cathepsin G-complexed ACT to the solid phase. This interference was substantially decreased in the presence of heparin. In studying the stability of purified PSA and PSA-ACT complexes formed in vitro, we found that the free PSA was stable during storage for 4 weeks at 35 degrees C, whereas PSA-ACT complexes largely dissociated in these conditions. The instability of PSA-ACT complexes was counteracted by storage at low temperatures, by adjusting the pH of the storage buffer between 6.8 and 7.4, and through addition of 100-1000-fold molar excess of native ACT. The ease of calibration and the accuracy of free PSA assays in comparison with assays of the PSA-ACT complex suggest that measurements of free to total PSA most accurately reflect the inverse of the proportion of PSA complexed to ACT in serum.

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Please use this url to cite or link to this publication:
author
organization
publishing date
type
Contribution to journal
publication status
published
keywords
Adsorption, Animals, Antibodies, Monoclonal, Cathepsin G, Cathepsins/blood, Drug Stability, Epitope Mapping, False Positive Reactions, Fluoroimmunoassay/methods, Humans, Male, Mice, Mice, Inbred BALB C, Prostate-Specific Antigen/blood, Sensitivity and Specificity, Serine Endopeptidases, alpha 1-Antichymotrypsin/blood
in
Clinical Chemistry
volume
41
issue
10
pages
9 pages
publisher
American Association for Clinical Chemistry
external identifiers
  • pmid:7586521
  • scopus:0028840275
ISSN
0009-9147
language
English
LU publication?
yes
id
3d2c9456-ae30-4fc5-bcf5-bce4e3df5779
alternative location
http://clinchem.aaccjnls.org/content/41/10/1480
date added to LUP
2019-05-16 14:07:23
date last changed
2020-01-13 01:48:22
@article{3d2c9456-ae30-4fc5-bcf5-bce4e3df5779,
  abstract     = {<p>Generation of 15 monoclonal antibodies (MAbs) allowed construction of epitope maps and specific two-site immunofluorometric assays for free prostate-specific antigen (PSA) and PSA complexed with alpha 1-antichymotrypsin (ACT). Close correlation of PSA concentrations obtained with the use of different assays of free PSA suggested extensive similarity in immunodetection of free PSA in serum. Assays of the PSA-ACT complex overestimated the concentration of PSA-ACT in serum because of nonspecific adsorbance of ACT or cathepsin G-complexed ACT to the solid phase. This interference was substantially decreased in the presence of heparin. In studying the stability of purified PSA and PSA-ACT complexes formed in vitro, we found that the free PSA was stable during storage for 4 weeks at 35 degrees C, whereas PSA-ACT complexes largely dissociated in these conditions. The instability of PSA-ACT complexes was counteracted by storage at low temperatures, by adjusting the pH of the storage buffer between 6.8 and 7.4, and through addition of 100-1000-fold molar excess of native ACT. The ease of calibration and the accuracy of free PSA assays in comparison with assays of the PSA-ACT complex suggest that measurements of free to total PSA most accurately reflect the inverse of the proportion of PSA complexed to ACT in serum.</p>},
  author       = {Pettersson, K and Piironen, T and Seppälä, M and Liukkonen, L and Christensson, A and Matikainen, M T and Suonpää, M and Lövgren, T and Lilja, H},
  issn         = {0009-9147},
  language     = {eng},
  number       = {10},
  pages        = {8--1480},
  publisher    = {American Association for Clinical Chemistry},
  series       = {Clinical Chemistry},
  title        = {Free and complexed prostate-specific antigen (PSA) : in vitro stability, epitope map, and development of immunofluorometric assays for specific and sensitive detection of free PSA and PSA-alpha 1-antichymotrypsin complex},
  url          = {http://clinchem.aaccjnls.org/content/41/10/1480},
  volume       = {41},
  year         = {1995},
}