Microtiter assay for colorimetric detection of in vitro amplified Chlamydia trachomatis sequences.
An integrated diagnostic method for colorimetric detection of polymerase chain reaction (PCR) products in a microtitre format is described. The amplified material is labelled by using modified PCR primers introducing a biotin molecule and a lac operator handle. Positive samples are identified, after immobilization onto streptavidin-coated microtitre wells, using a reporter fusion protein consisting of the Escherichia coli lac repressor and beta-galactosidase. The analysis of the PCR products is thus, from a practical point of view, identical with the enzyme-linked immunosorbent assay (ELISA) using the enzymatic activity of beta-galactosidase. Here, we show that the assay can be used for rapid and simple detection of Chlamydia trachomatis in an automated format. The reported analysis of 90 urethral patient samples, using this microtitre assay, indicates that this technique is more sensitive and more specific than culture technique.
Lundeberg
J
J
Department of Biochemistry and Biotechnology, Royal Institute of Technology, Stockholm, Sweden.
Bondesson
L
L
Hedrum
A
A
Grillner
L
L
Stark
M
M
von Krogh
G
G
Uhlén
M
M
D001483:Base Sequence; D002478:Cells, Cultured; D002690:Chlamydia Infections / Q000175:diagnosis* / Q000382:microbiology / Q000652:urine; D002692:Chlamydia trachomatis / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D003001:Cloning, Molecular; D003124:Colorimetry / Q000379:methods*; D004269:DNA, Bacterial / Q000032:analysis*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D008297:Male; D008832:Microchemistry; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity
D004269:DNA, Bacterial