CrELISA: a fast and robust enzyme-linked immunosorbent assay bypassing the need for purification of recombinant protein.
A multitude of antigens has been recently identified by screening of cDNA expression libraries derived from human tumors with autologous sera. Using a phage autoantibody assay and small panels of sera derived from cancer patients or controls it has been shown that some of these antigens display cancer-associated autoantibody responses. The diagnostic and prognostic significance of these potentially cancer-related autoantibodies remains unclear until large-scale assays are developed and serological data are available for hundreds of cancer patients and controls. The major bottleneck for the development of large-scale assays are the cloning, expression and the purification of each of the respective antigens. Due to these limitations and despite the potential clinical relevance large-scale autoantibody tests are established for only a few of these tumor antigens. Here we describe an enzyme-linked immunosorbent assay, Crude lysate ELISA (CrELISA), suitable for antigens identified by expression screening based on crude lysates of antigen-expressing bacteria. This assay permits sensitive and specific autoantibody seroscreening without the need of laborious and time-consuming cloning, expression and purification of recombinant proteins. CrELISA is robust and provides a versatile high throughput procedure for the rapid evaluation of multiple antigens in large-scale serology.
Türeci
Ozlem
O
III. Department of Internal Medicine, Johannes Gutenberg University, Mainz, Germany. tureci@mail.uni-mainz.de
Luxemburger
Uli
U
Heinen
Helma
H
Mack
Ulrich
U
Sybrecht
Gerhard W
GW
Huber
Christoph
C
Sahin
Ugur
U
D000912:Antibodies, Neoplasm / Q000097:blood*; D000951:Antigens, Neoplasm / Q000032:analysis* / Q000096:biosynthesis / Q000302:isolation & purification; D001323:Autoantibodies / Q000097:blood*; D001324:Autoantigens / Q000032:analysis* / Q000096:biosynthesis / Q000302:isolation & purification; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D004926:Escherichia coli / Q000276:immunology / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000032:analysis / Q000096:biosynthesis / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000032:analysis* / Q000096:biosynthesis / Q000302:isolation & purification
D000912:Antibodies, Neoplasm; D000951:Antigens, Neoplasm; D001323:Autoantibodies; D001324:Autoantigens; C104948:CTAG1B protein, human; D008565:Membrane Proteins; D011994:Recombinant Proteins