Nucleic Acid Extraction from Residual Aptima Clinical Samples: Evaluation of Automated Platform To Enhance Workflow

摘要

Previously it has been shown that Aptima collection buffer is not compatible with use on other platforms, resulting in invalid tests or major loss of sensitivity (3, 4). This is not desirable for supplementary testing. Extraction by automated methods is convenient for large-volume testing but has not been published previously. We evaluated the performance of MagNA Pure 96 (Roche Diagnostics) for nucleic acid extraction from residual Aptima collection buffer from clinical samples previously tested with the AC2 assay. Nucleic acid was extracted from 344 nongenital (throat, rectum, eye) samples from adult patients (138 N. gonorrhoeae positive [NG+]/C. trachomatis negative [CT?], 154 NG?/CT+, 16 NG+/CT+, 20 NG?/CT?) with the MagNA Pure 96 Universal Pathogen 200 protocol and the MagNA Pure 96 DNA and Viral DNA small-volume kit with a final elution of 50 μl. Five microliters of extracted nucleic acid was then tested for N. gonorrhoeae by using porA and opa targets by using LightCycler 480 with Probe MasterMix (Roche) and the Panther Aptima AC2 and Aptima Neisseria gonorrhoeae (AGC) assays (5). Samples that were CT+ with AC2 were tested by real-time PCR by using an LGV pmpH target (6). A human gene target (RNase P) used as an internal control (IC) was detected in all 344 samples, confirming successful extraction. Twenty known negative samples were interspersed with positives to detect sample cross-contamination. Negative samples were concordant on all assays.