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New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination

Loop mediated isothermal amplification (LAMP) assay, a promising diagnostic test, has been developed for detection of different pathogens of human as well as animals. Various positive points support its use as a field level test but the major problem is product cross contamination leading to false p...

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Autores principales: Karthik, K., Rathore, Rajesh, Thomas, Prasad, Arun, T.R., Viswas, K.N., Dhama, Kuldeep, Agarwal, R.K.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2014
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4472950/
https://www.ncbi.nlm.nih.gov/pubmed/26150945
http://dx.doi.org/10.1016/j.mex.2014.08.009
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author Karthik, K.
Rathore, Rajesh
Thomas, Prasad
Arun, T.R.
Viswas, K.N.
Dhama, Kuldeep
Agarwal, R.K.
author_facet Karthik, K.
Rathore, Rajesh
Thomas, Prasad
Arun, T.R.
Viswas, K.N.
Dhama, Kuldeep
Agarwal, R.K.
author_sort Karthik, K.
collection PubMed
description Loop mediated isothermal amplification (LAMP) assay, a promising diagnostic test, has been developed for detection of different pathogens of human as well as animals. Various positive points support its use as a field level test but the major problem is product cross contamination leading to false positive results. Different methods were adopted by various researchers to control this false positive amplification due to cross contamination but all have their own advantages and disadvantages. A new closed tube LAMP assay based on agar dye capsule was developed in the present study and this technique has some advantages over the other closed tube technique. • Agar at the concentration of 1.5% was used to sandwich SYBR green dye I with the aid of intradermal syringe. This agar dye capsule was placed over the LAMP reaction mixture before it was amplified. • To eliminate the hazardous nature of Ultra Violet (UV) light during result visualization of LAMP products, the present study demonstrates the use of Light Emitting Diode (LED) lights for result visualization. • LAMP was carried out for Brucella species detection using this modified techniques yielding good results without any cross contamination and LED showed similar fluorescence compared to UV.
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spelling pubmed-44729502015-07-06 New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination Karthik, K. Rathore, Rajesh Thomas, Prasad Arun, T.R. Viswas, K.N. Dhama, Kuldeep Agarwal, R.K. MethodsX Article Loop mediated isothermal amplification (LAMP) assay, a promising diagnostic test, has been developed for detection of different pathogens of human as well as animals. Various positive points support its use as a field level test but the major problem is product cross contamination leading to false positive results. Different methods were adopted by various researchers to control this false positive amplification due to cross contamination but all have their own advantages and disadvantages. A new closed tube LAMP assay based on agar dye capsule was developed in the present study and this technique has some advantages over the other closed tube technique. • Agar at the concentration of 1.5% was used to sandwich SYBR green dye I with the aid of intradermal syringe. This agar dye capsule was placed over the LAMP reaction mixture before it was amplified. • To eliminate the hazardous nature of Ultra Violet (UV) light during result visualization of LAMP products, the present study demonstrates the use of Light Emitting Diode (LED) lights for result visualization. • LAMP was carried out for Brucella species detection using this modified techniques yielding good results without any cross contamination and LED showed similar fluorescence compared to UV. Elsevier 2014-08-27 /pmc/articles/PMC4472950/ /pubmed/26150945 http://dx.doi.org/10.1016/j.mex.2014.08.009 Text en © 2014 The Authors http://creativecommons.org/licenses/by/3.0/ This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/3.0/).
spellingShingle Article
Karthik, K.
Rathore, Rajesh
Thomas, Prasad
Arun, T.R.
Viswas, K.N.
Dhama, Kuldeep
Agarwal, R.K.
New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination
title New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination
title_full New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination
title_fullStr New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination
title_full_unstemmed New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination
title_short New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination
title_sort new closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4472950/
https://www.ncbi.nlm.nih.gov/pubmed/26150945
http://dx.doi.org/10.1016/j.mex.2014.08.009
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