Open AccessModified dot assay with increased sensitivity: Detection of small amounts of immunoglobulin molecules and the importance of different detection systems
Author(s) -
Lavanchy Daniel,
Stroun Jacques,
Frei Philippe C.
Publication year - 1990
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/jcla.1860040404
Subject(s) - avidin , chemistry , streptavidin , bovine serum albumin , antiserum , substrate (aquarium) , peroxidase , monoclonal antibody , biotin , antibody , chromatography , microbiology and biotechnology , biochemistry , enzyme , biology , immunology , ecology
The original method of the dot assay described by Hawkes et al. enabled the detection of 100 pg of immunoglobulin molecules. We compared different enzyme detection systems based on the avidin‐biotin complex, in conjunction with different substrates. Three commercial kits (Vectastain‐ABC, Streptavidin Bridge, and Streptavidin‐Peroxidase Complex) based on the same technique were also included in the comparison. In addition, we tested the Aureo‐Probe kit, which is based on gold‐ and silver‐enhanced staining. Various incubation times, blocking solutions (horse serum, bovine serum albumin, and milk), and commercial antihuman Ig antisera or monoclonal antibodies were tested. The greatest sensitivity was achieved by using the peroxidase‐avidin‐biotin complex together with 4‐chloro‐1‐naphtol as substrate, which detected 1 pg of immunoglobulin molecules. Equal sensitivity was also shown by the Aureo‐Probe system.