Detection of Loop-Mediated Isothermal Amplification Reaction by Turbidity Derived from Magnesium Pyrophosphate Formation

Mori, Yasuyoshi; Nagamine, Kentaro; Tomita, Norihiro; Notomi, Tsugunori

doi:10.1006/bbrc.2001.5921

Cited by 1,523 publications

(1,142 citation statements)

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“…The degree of amplification was detected via a byproduct of the reaction, pyrophosphate. 15 Upon precipitation of magnesium pyrophosphate, the resulting change in turbidity was in turn correlated to CK19 mRNA copy number/lL of the original lysate via a standard curve that was established beforehand with 3 calibrators containing different CK19 mRNA concentrations. A standard positive control sample containing 5000 copies/ lL of CK19 mRNA and a negative control sample not containing any CK19 mRNA were used for validation in every assay.…”

Section: Osna Assaymentioning

confidence: 99%

Intraoperative molecular assay for sentinel lymph node metastases in early stage breast cancer

Osako

Iwase

Kimura

et al. 2011

Cancer

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BACKGROUND: Conventional histopathological examination is limited in measuring accurate total metastatic volume in a lymph node. Recently, a molecular-based procedure to detect lymph node metastases, one-step nucleic acid amplification (OSNA) assay, has been developed. OSNA assay can assess a whole lymph node and yields semiquantitative results. The authors compared the performance in intraoperative detection of sentinel lymph node metastases with OSNA assay using a whole lymph node versus routine frozen section (FS) histology with a 2 mm-sectioned lymph node. METHODS: Subjects comprised 531 consecutive patients diagnosed with OSNA assay and 618 consecutive patients diagnosed with FS histological examination. The authors compared the sentinel lymph node-positive rate between the OSNA and FS cohorts, and investigated characteristics of patients for whom OSNA could detect metastases but FS could not. OSNA (þ) was defined as micrometastasis, and OSNA (þþ) and (þI) were defined as macrometastasis. RESULTS: OSNA assay detected more cases of sentinel lymph node metastases than FS histology (OSNA 121 of 531, 22.8% vs FS 109 of 618, 17.6%; P ¼ .036), particularly micrometastases (46 of 531, 8.7% vs 28 of 618, 4.5%; P ¼ .0064). There was no difference in macrometastasis detection between OSNA and FS (75 of 531, 14.1% vs 81 of 618, 13.1%; P ¼ .68). OSNA detected more metastases than FS in postmenopausal patients (77 of 302, 25.5% vs 43 of 351, 12.3%; P < .0001), and in tumors without fat invasion (23 of 156, 14.7% vs 6 of 151, 4.0%; P ¼ .012) or lymphovascular invasion (67 of 395, 17.0% vs 45 of 458, 9.8%; P ¼ .042). CONCLUSIONS: Intraoperative OSNA assay detects more sentinel lymph node metastases, particularly micrometastases, than does FS histology. OSNA assay can also detect more metastases in postmenopausal patients or from less aggressive primary tumors compared with FS histology. Cancer 2011;117:4365-

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Section: Osna Assaymentioning

confidence: 99%

Intraoperative molecular assay for sentinel lymph node metastases in early stage breast cancer

Osako

Iwase

Kimura

et al. 2011

Cancer

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“…Moreover, this method can be carried out with simple systems and the LAMP reaction can be monitored in real-time through measurement of turbidity, which is correlated with the production of magnesium pyrophosphate, by means of an inexpensive photometer (Mori et al, 2001).…”

Section: Introductionmentioning

confidence: 99%

Development and evaluation of a loop-mediated isothermal amplification assay for rapid detection of Mycoplasma pneumoniae

Saito

Misawa

Moriya

et al. 2005

Journal of Medical Microbiology

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A loop-mediated isothermal amplification (LAMP) assay for the rapid detection of Mycoplasma pneumoniae was developed and evaluated. The assay specifically amplified only M. pneumoniae sequences, and no cross-reactivity was observed for other Mycoplasma species or respiratory bacterial species. The detection limit for this assay was found to be 2 3 10 2 copies, corresponding to 2-20 colour changing units of M. pneumoniae in 1 h, as observed in a real-time turbidimeter and electrophoretic analysis. The accuracy of the LAMP reaction was confirmed by restriction endonuclease analysis as well as direct sequencing of the amplified product. The assay was applied to 95 nasopharyngeal swab samples collected from patients or from healthy individuals, and compared to a real-time PCR assay in-house. A concordance of 100 % was observed between the two assays. The LAMP assay is easy to perform, shows a rapid reaction and is inexpensive. It may therefore be applied in the routine diagnosis of M. pneumoniae infection in the clinical laboratory.

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“…The combination of isothermal conditions and the Bst polymerase in the LAMP assay showed resistance to inhibitors that usually hinder the later stages of conventional PCR (Khan et al, 2012;Mori et al, 2001). LAMP reagents and Bst polymerase are also relatively stable even if kept at 25 and 37 uC (Thekisoe et al, 2009), and only a conventional heating block is required to perform this assay.…”

Section: Discussionmentioning

confidence: 99%

Loop-mediated isothermal amplification for rapid molecular detection of Enterocytozoon bieneusi in faecal specimens

Nasarudin

Zainudin

Bernadus

et al. 2015

Journal of Medical Microbiology

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A loop-mediated isothermal amplification (LAMP) assay was developed to detect Enterocytozoon bieneusi DNA for the first time from human faecal specimens. Four primers specific for Enterocytozoon bieneusi were designed corresponding to small subunit rRNA gene sequences and tested on 100 human faecal specimens. Thirty-nine of the faecal specimens (39 %) were confirmed positive for Enterocytozoon bieneusi by LAMP compared with 33 % by PCR and 32 % by light microscopy. LAMP yielded 94 % sensitivity and 88 % specificity compared with microscopy (sensitivity 48 %, specificity 76 %). No significant differences in positive detection of Enterocytozoon bieneusi were found among the three methods (P.0.05). However, LAMP has shown a substantial agreement with PCR (k50.78) and fair agreement was demonstrated between microscopy and PCR (k50.25). In conclusion, the LAMP assay proved to be useful as a simplified, rapid, sensitive and specific alternative molecular screening tool in the diagnosis of Enterocytozoon bieneusi in faecal specimens.

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Detection of Loop-Mediated Isothermal Amplification Reaction by Turbidity Derived from Magnesium Pyrophosphate Formation

Cited by 1,523 publications

References 12 publications

Intraoperative molecular assay for sentinel lymph node metastases in early stage breast cancer

Intraoperative molecular assay for sentinel lymph node metastases in early stage breast cancer

Development and evaluation of a loop-mediated isothermal amplification assay for rapid detection of Mycoplasma pneumoniae

Loop-mediated isothermal amplification for rapid molecular detection of Enterocytozoon bieneusi in faecal specimens

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