Characteristics of the m 2000 Automated Sample Preparation and Multiplex Real-Time PCR System for Detection of Chlamydia trachomatis and Neisseria gonorrhoeae

Abstract

We evaluated a new real-time PCR-based prototype assay for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae developed by Abbott Molecular Inc. This assay is designed to be performed on an Abbott m2000 real-time instrument system, which consists of an m2000sp instrument for sample preparation and an m2000rt instrument for real-time PCR amplification and detection. The limit of detection of this prototype assay was determined to be 20 copies of target DNA for both C. trachomatis and N. gonorrhoeae, using serially diluted linearized plasmids. No cross-reactivity could be detected when 55 nongonococcal Neisseria isolates and 3 non-C. trachomatis Chlamydia isolates were tested at 1 million genome equivalents per reaction. Concordance with the Roche Amplicor, BDProbeTec ET, and Gen-Probe APTIMA Combo 2 tests was assessed using unlinked/deidentified surplus clinical specimens previously analyzed with these tests. For C. trachomatis, concordance for positive results ranged from 93.7% to 100%, while concordance for negative results ranged from 98.2% to 100%. For N. gonorrhoeae, concordance for positive and negative results ranged from 91.4% to 100% and 99.3% to 100%, respectively. A workflow analysis of the prototype assay was conducted to obtain information on throughput under laboratory conditions. At 48 samples/run, the time to first result for both C. trachomatis and N. gonorrhoeae was 4.5 h. A total of 135 patient specimens could be analyzed in 8.9 h, with 75 min of hands-on time. This study demonstrated the technical and clinical feasibility of the new Abbott real-time PCR C. trachomatis/N. gonorrhoeae assay.

Rapid and Specific Detection of Amantadine-Resistant Influenza A Viruses with a Ser31Asn Mutation by the Cycling Probe Method

“…However, in the cycling probe method, it is not possible to adopt a one-step real-time PCR, because it includes RNase H and the template could not achieve a sufficient length of cDNA during the reverse transcription. Other common controls are cDNA (two-step method), but plasmid preparations have higher degrees of stability and reproducibility (20,23,28). Various methods has been used to examine virus isolates for their amantadine susceptibilities, such as ELISA (4), plaque reduction assay (13), the TCID 50 /0.2-ml method (24), PCR-RFLP analysis (21,31), and DNA sequencing (24).…”

Evaluation of the Abbott m 2000 RealTi m e Human Immunodeficiency Virus Type 1 (HIV-1) Assay for HIV Load Monitoring in South Africa Compared to the Roche Cobas AmpliPrep-Cobas Amplicor, Roche Cobas AmpliPrep-Cobas TaqMan HIV-1, and BioMerieux NucliSENS EasyQ HIV-1 Assays

“…Controls are also supplied with each batch, and the number of controls is also the same if 48 or 96 tests are performed. This assay showed overall good performance and potential for use with samples from populations predominantly infected with HIV-1 subtype C. It is reliable, minimizes contamination through the use of automation, and has a rapid turnaround time (with few requirements for timeconsuming manual steps); and the assay has the potential to be expanded for use for the detection of other organisms: hepatitis C virus (43), Chlamydia trachomatis, and Neisseria gonorrhoeae (22).…”

Immiscible Phase Nucleic Acid Purification Eliminates PCR Inhibitors with a Single Pass of Paramagnetic Particles through a Hydrophobic Liquid

“…The purified DNA was amplified using the Abbott RealTime CT/NG assay in a 50-l reaction volume. 21 Amplification reactions were performed in the Abbott Molecular m2000rt instrument (Abbott Park, IL).…”